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1
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Bresesti C, Carito E, Notaro M, Giacca G, Breggion S, Kerzel T, Mercado CM, Beretta S, Monti M, Merelli I, Canu T, Naldini L, Squadrito ML. Reprogramming liver metastasis-associated macrophages toward an anti-tumoral phenotype through enforced miR-342 expression. Cell Rep 2025; 44:115592. [PMID: 40253698 DOI: 10.1016/j.celrep.2025.115592] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/13/2024] [Revised: 01/24/2025] [Accepted: 03/29/2025] [Indexed: 04/22/2025] Open
Abstract
Upon metastatic seeding in the liver, liver macrophages, including Kupffer cells, acquire a transcriptional profile typical of tumor-associated macrophages (TAMs), which support tumor progression. MicroRNAs (miRNAs) fine-tune TAM pro-tumoral functions, making their modulation a promising strategy for macrophage reprogramming into an anti-tumoral phenotype. Here, we analyze the transcriptomic profiles of liver and splenic macrophages, identifying miR-342-3p as a key regulator of liver macrophage function. miR-342-3p is highly active in healthy liver macrophages but significantly downregulated in colorectal cancer liver metastases (CRLMs). Lentiviral vector-engineered liver macrophages enforcing miR-342-3p expression acquire a pro-inflammatory phenotype and reduce CRLM growth. We identify Slc7a11, a cysteine-glutamate antiporter linked to pro-tumoral activity, as a direct miR-342-3p target, which may be at least partially responsible for TAM phenotypic reprogramming. Our findings highlight the potential of in vivo miRNA modulation as a therapeutic strategy for TAM reprogramming, offering an approach to enhance cancer immunotherapy.
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Affiliation(s)
- Chiara Bresesti
- Vector Engineering and In vivo Tumor Targeting Unit, San Raffaele Telethon Institute for Gene Therapy, IRCCS San Raffaele Scientific Institute, 20132 Milan, Italy; Vita-Salute San Raffaele University, 20132 Milan, Italy
| | - Eleonora Carito
- Vector Engineering and In vivo Tumor Targeting Unit, San Raffaele Telethon Institute for Gene Therapy, IRCCS San Raffaele Scientific Institute, 20132 Milan, Italy; Vita-Salute San Raffaele University, 20132 Milan, Italy
| | - Marco Notaro
- Vector Engineering and In vivo Tumor Targeting Unit, San Raffaele Telethon Institute for Gene Therapy, IRCCS San Raffaele Scientific Institute, 20132 Milan, Italy
| | - Giovanna Giacca
- Vector Engineering and In vivo Tumor Targeting Unit, San Raffaele Telethon Institute for Gene Therapy, IRCCS San Raffaele Scientific Institute, 20132 Milan, Italy; Vita-Salute San Raffaele University, 20132 Milan, Italy
| | - Sara Breggion
- Vector Engineering and In vivo Tumor Targeting Unit, San Raffaele Telethon Institute for Gene Therapy, IRCCS San Raffaele Scientific Institute, 20132 Milan, Italy; Vita-Salute San Raffaele University, 20132 Milan, Italy
| | - Thomas Kerzel
- Vector Engineering and In vivo Tumor Targeting Unit, San Raffaele Telethon Institute for Gene Therapy, IRCCS San Raffaele Scientific Institute, 20132 Milan, Italy
| | - Carl Mirko Mercado
- Vector Engineering and In vivo Tumor Targeting Unit, San Raffaele Telethon Institute for Gene Therapy, IRCCS San Raffaele Scientific Institute, 20132 Milan, Italy
| | - Stefano Beretta
- BioInformatics Core, San Raffaele Telethon Institute for Gene Therapy, IRCCS San Raffaele Scientific Institute, 20132 Milan, Italy
| | - Marco Monti
- BioInformatics Core, San Raffaele Telethon Institute for Gene Therapy, IRCCS San Raffaele Scientific Institute, 20132 Milan, Italy
| | - Ivan Merelli
- BioInformatics Core, San Raffaele Telethon Institute for Gene Therapy, IRCCS San Raffaele Scientific Institute, 20132 Milan, Italy
| | - Tamara Canu
- Preclinical Imaging Facility, IRCCS San Raffaele Scientific Institute, 20132 Milan, Italy
| | - Luigi Naldini
- Vita-Salute San Raffaele University, 20132 Milan, Italy; Targeted Cancer Gene Therapy Unit, San Raffaele Telethon Institute for Gene Therapy, IRCCS San Raffaele Scientific Institute, 20132 Milan, Italy
| | - Mario Leonardo Squadrito
- Vector Engineering and In vivo Tumor Targeting Unit, San Raffaele Telethon Institute for Gene Therapy, IRCCS San Raffaele Scientific Institute, 20132 Milan, Italy; Vita-Salute San Raffaele University, 20132 Milan, Italy.
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2
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Mazza T, Aslanzadeh M, Berentsen L, Bonath F, Friedländer MR, Barragan A. Cytoskeletal alterations in neuronal cells implicate Toxoplasma gondii secretory machinery and host microRNA-containing extracellular vesicles. Sci Rep 2025; 15:12606. [PMID: 40221584 PMCID: PMC11993698 DOI: 10.1038/s41598-025-96298-8] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/19/2024] [Accepted: 03/27/2025] [Indexed: 04/14/2025] Open
Abstract
The widespread protozoan Toxoplasma gondii chronically infects neural tissue in vertebrates and is linked to various neurological and neuropsychiatric disorders in humans. However, its effects on sparsely infected neurons and on broader neural circuits remain elusive. Our study reveals that T. gondii infection disrupts cytoskeletal dynamics in SH-SY5Y neuronal cells and primary cortical neurons. Infected neuronal cells undergo significant cytomorphological changes, including retraction of dendritic extensions and alterations in microtubule and F-actin networks, across both parasite genotypes I and II. These cytoskeletal alterations were notably diminished in cells exposed to T. gondii mutants with impaired secretion via the MYR translocon, and were independent of intraneuronal parasite replication. Moreover, a bystander effect was observed, with supernatants from T. gondii-challenged cells inducing similar cytoskeletal changes in uninfected cells. Analyses of extracellular vesicles (EVs) in supernatants revealed differential expression of host microRNAs in response to infection, most notably the upregulation of miR-221-3p, a microRNA not previously associated with T. gondii. The data indicate that unidentified parasite-derived effector(s) secreted via the MYR translocon, in conjunction with MYR-independently induced EV-associated host microRNAs, mediate cytoskeletal alterations in both infected and bystander neuronal cells. The findings provide new insights into molecular mechanisms by which T. gondii infection may disrupt neural networks, shedding light on its potential role in neuronal dysregulation.
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Affiliation(s)
- Thomas Mazza
- Department of Molecular Biosciences, The Wenner-Gren Institute, Stockholm University, Stockholm, Sweden
| | - Morteza Aslanzadeh
- Science for Life Laboratory, Department of Molecular Biosciences, The Wenner-Gren Institute, Stockholm University, Stockholm, Sweden
| | - Lïse Berentsen
- Department of Molecular Biosciences, The Wenner-Gren Institute, Stockholm University, Stockholm, Sweden
| | - Franziska Bonath
- Department of Gene Technology, School of Engineering Sciences in Chemistry, Biotechnology and Health, Kungliga Tekniska Högskolan, Stockholm, Sweden
| | - Marc R Friedländer
- Science for Life Laboratory, Department of Molecular Biosciences, The Wenner-Gren Institute, Stockholm University, Stockholm, Sweden
| | - Antonio Barragan
- Department of Molecular Biosciences, The Wenner-Gren Institute, Stockholm University, Stockholm, Sweden.
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3
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Liu W, Yang H, Liu X, Cai H, Bao Y, Jiang Y, Zhou W, Yuan J, Zhang Z, Fang X. Ultrasensitive Quantification of microRNA Copy Number in Individual Extracellular Vesicles Using DNA Tetrahedron-Based Single-Molecule Imaging. Anal Chem 2025; 97:4233-4240. [PMID: 39936597 DOI: 10.1021/acs.analchem.4c07068] [Citation(s) in RCA: 2] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/13/2025]
Abstract
The ultrasensitive detection of microRNAs (miRNAs) in extracellular vesicles (EVs) can accurately reflect the progress and metastasis of miRNA-mediated intercellular communication, providing an unprecedented opportunity for liquid biopsy. However, due to the low abundance and high heterogeneity of miRNAs in EVs, the ultrasensitive quantification and establishment of a distribution model for miRNA within native EVs remain challenging. Here, we have developed a DNA tetrahedron-based single-molecule fluorescence imaging strategy to overcome this challenge. The internalization efficiency of the probe was as high as 70% without disrupting the native structure of EVs, and combined with single-molecule fluorescence imaging, we achieved in situ imaging analysis of single-copy miRNA in individual EVs without amplification for the first time. A new distribution model for miRNAs has been revealed by statistical analysis of the copy number of miRNAs in EVs across multiple cell lines, characterized by low occupancy and a heterogeneous distribution. More importantly, we found that drug resistance cancer cells promote an increase in the number of drug resistance-related miRNAs within EVs without a corresponding increase in the number of EVs secreted, providing new insights into the EV miRNA sorting mechanisms. We anticipate that this technology will rapidly advance miRNA-mediated intercellular communication based on EVs.
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Affiliation(s)
- Weifeng Liu
- Key Laboratory of Molecular Nanostructure and Nanotechnology, CAS Research/Education Center for Excellence in Molecular Sciences, Institute of Chemistry, Chinese Academy of Sciences, Beijing 100190, China
- University of Chinese Academy of Sciences, Beijing 100049, China
| | - Hongwei Yang
- Technical Institute of Physics and Chemistry, Chinese Academy of Sciences, Beijing 100190, China
| | - Xiaolong Liu
- Key Laboratory of Molecular Nanostructure and Nanotechnology, CAS Research/Education Center for Excellence in Molecular Sciences, Institute of Chemistry, Chinese Academy of Sciences, Beijing 100190, China
- University of Chinese Academy of Sciences, Beijing 100049, China
| | - Heqi Cai
- Key Laboratory of Molecular Nanostructure and Nanotechnology, CAS Research/Education Center for Excellence in Molecular Sciences, Institute of Chemistry, Chinese Academy of Sciences, Beijing 100190, China
- University of Chinese Academy of Sciences, Beijing 100049, China
| | - Yuting Bao
- Hangzhou Institute of Medicine (HIM), Chinese Academy of Sciences, Hangzhou, Zhejiang 310022, China
| | - Yifei Jiang
- Hangzhou Institute of Medicine (HIM), Chinese Academy of Sciences, Hangzhou, Zhejiang 310022, China
| | - Wei Zhou
- Hangzhou Institute of Medicine (HIM), Chinese Academy of Sciences, Hangzhou, Zhejiang 310022, China
| | - Jinghe Yuan
- Key Laboratory of Molecular Nanostructure and Nanotechnology, CAS Research/Education Center for Excellence in Molecular Sciences, Institute of Chemistry, Chinese Academy of Sciences, Beijing 100190, China
| | - Zhen Zhang
- Huairou Research Center, Institute of Chemistry, Chinese Academy of Sciences, Beijing 100190, China
| | - Xiaohong Fang
- Key Laboratory of Molecular Nanostructure and Nanotechnology, CAS Research/Education Center for Excellence in Molecular Sciences, Institute of Chemistry, Chinese Academy of Sciences, Beijing 100190, China
- University of Chinese Academy of Sciences, Beijing 100049, China
- Hangzhou Institute of Medicine (HIM), Chinese Academy of Sciences, Hangzhou, Zhejiang 310022, China
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4
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Chutipongtanate S, Kongsomros S, Cetinkaya H, Zhang X, Kuhnell D, Benefield D, Haffey WD, Wyder MA, Kwatra G, Conrey SC, Burrell AR, Langevin SM, Esfandiari L, Newburg DS, Greis KD, Staat MA, Morrow AL. Prenatal SARS-CoV-2 Infection Alters Human Milk-Derived Extracellular Vesicles. Cells 2025; 14:284. [PMID: 39996756 PMCID: PMC11853888 DOI: 10.3390/cells14040284] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/26/2025] [Revised: 02/12/2025] [Accepted: 02/13/2025] [Indexed: 02/26/2025] Open
Abstract
Human milk-derived extracellular vesicles (HMEVs) are key components in breast milk, promoting infant health and development. Maternal conditions could affect HMEV cargo; however, the impact of SARS-CoV-2 infection on HMEVs remains unknown. This study investigated the influence of SARS-CoV-2 infection during pregnancy on postpartum HMEV molecules. The median duration from SARS-CoV-2 test positivity to milk collection was 3 months. After defatting and casein micelle disaggregation, HMEVs were isolated from milk samples of nine mothers with prenatal SARS-CoV-2 and six controls by sequential centrifugation, ultrafiltration, and qEV-size exclusion chromatography. The presence of HMEV was confirmed via transmission electron microscopy. Nanoparticle tracking analysis demonstrated particle diameters of <200 nm and yields of >1 × 1011 particles per mL of milk. Western immunoblots detected ALIX, CD9, and HSP70, supporting the presence of HMEVs in the isolates. Cargo from thousands of HMEVs were analyzed using a multi-omics approach, including proteomics and microRNA sequencing, and predicted that mothers with prenatal SARS-CoV-2 infection produced HMEVs with enhanced functionalities involving metabolic reprogramming, mucosal tissue development, and immunomodulation. Our findings suggest that SARS-CoV-2 infection during pregnancy boosts mucosal site-specific functions of HMEVs, potentially protecting infants against viral infections. Further prospective studies should be pursued to reevaluate the short- and long-term benefits of breastfeeding in the post-COVID era.
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Affiliation(s)
- Somchai Chutipongtanate
- Department of Environmental and Public Health Sciences, University of Cincinnati College of Medicine, Cincinnati, OH 45267, USA; (S.K.); (H.C.); (X.Z.); (D.K.); (S.C.C.); (D.S.N.)
| | - Supasek Kongsomros
- Department of Environmental and Public Health Sciences, University of Cincinnati College of Medicine, Cincinnati, OH 45267, USA; (S.K.); (H.C.); (X.Z.); (D.K.); (S.C.C.); (D.S.N.)
- Department of Biomedical Engineering, University of Cincinnati, Cincinnati, OH 45267, USA;
| | - Hatice Cetinkaya
- Department of Environmental and Public Health Sciences, University of Cincinnati College of Medicine, Cincinnati, OH 45267, USA; (S.K.); (H.C.); (X.Z.); (D.K.); (S.C.C.); (D.S.N.)
| | - Xiang Zhang
- Department of Environmental and Public Health Sciences, University of Cincinnati College of Medicine, Cincinnati, OH 45267, USA; (S.K.); (H.C.); (X.Z.); (D.K.); (S.C.C.); (D.S.N.)
| | - Damaris Kuhnell
- Department of Environmental and Public Health Sciences, University of Cincinnati College of Medicine, Cincinnati, OH 45267, USA; (S.K.); (H.C.); (X.Z.); (D.K.); (S.C.C.); (D.S.N.)
| | - Desirée Benefield
- Center for Advanced Structural Biology, Department of Molecular & Cellular Biosciences, University of Cincinnati College of Medicine, Cincinnati, OH 45267, USA;
| | - Wendy D. Haffey
- Department of Cancer Biology, University of Cincinnati College of Medicine, Cincinnati, OH 45267, USA; (W.D.H.); (M.A.W.); (K.D.G.)
| | - Michael A. Wyder
- Department of Cancer Biology, University of Cincinnati College of Medicine, Cincinnati, OH 45267, USA; (W.D.H.); (M.A.W.); (K.D.G.)
| | - Gaurav Kwatra
- Department of Infectious Disease, Cincinnati Children’s Hospital Medical Center, Cincinnati, OH 45267, USA; (G.K.); (A.R.B.); (M.A.S.)
- Department of Clinical Microbiology, Christian Medical College, Vellore 632004, India
| | - Shannon C. Conrey
- Department of Environmental and Public Health Sciences, University of Cincinnati College of Medicine, Cincinnati, OH 45267, USA; (S.K.); (H.C.); (X.Z.); (D.K.); (S.C.C.); (D.S.N.)
| | - Allison R. Burrell
- Department of Infectious Disease, Cincinnati Children’s Hospital Medical Center, Cincinnati, OH 45267, USA; (G.K.); (A.R.B.); (M.A.S.)
| | - Scott M. Langevin
- Larner College of Medicine, University of Vermont, Burlington, VT 05405, USA;
| | - Leyla Esfandiari
- Department of Biomedical Engineering, University of Cincinnati, Cincinnati, OH 45267, USA;
| | - David S. Newburg
- Department of Environmental and Public Health Sciences, University of Cincinnati College of Medicine, Cincinnati, OH 45267, USA; (S.K.); (H.C.); (X.Z.); (D.K.); (S.C.C.); (D.S.N.)
| | - Kenneth D. Greis
- Department of Cancer Biology, University of Cincinnati College of Medicine, Cincinnati, OH 45267, USA; (W.D.H.); (M.A.W.); (K.D.G.)
| | - Mary A. Staat
- Department of Infectious Disease, Cincinnati Children’s Hospital Medical Center, Cincinnati, OH 45267, USA; (G.K.); (A.R.B.); (M.A.S.)
| | - Ardythe L. Morrow
- Department of Environmental and Public Health Sciences, University of Cincinnati College of Medicine, Cincinnati, OH 45267, USA; (S.K.); (H.C.); (X.Z.); (D.K.); (S.C.C.); (D.S.N.)
- Department of Infectious Disease, Cincinnati Children’s Hospital Medical Center, Cincinnati, OH 45267, USA; (G.K.); (A.R.B.); (M.A.S.)
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5
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Ellrott K, Wong CK, Yau C, Castro MAA, Lee JA, Karlberg BJ, Grewal JK, Lagani V, Tercan B, Friedl V, Hinoue T, Uzunangelov V, Westlake L, Loinaz X, Felau I, Wang PI, Kemal A, Caesar-Johnson SJ, Shmulevich I, Lazar AJ, Tsamardinos I, Hoadley KA, Robertson AG, Knijnenburg TA, Benz CC, Stuart JM, Zenklusen JC, Cherniack AD, Laird PW. Classification of non-TCGA cancer samples to TCGA molecular subtypes using compact feature sets. Cancer Cell 2025; 43:195-212.e11. [PMID: 39753139 PMCID: PMC11949768 DOI: 10.1016/j.ccell.2024.12.002] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 08/16/2024] [Revised: 08/26/2024] [Accepted: 12/05/2024] [Indexed: 02/12/2025]
Abstract
Molecular subtypes, such as defined by The Cancer Genome Atlas (TCGA), delineate a cancer's underlying biology, bringing hope to inform a patient's prognosis and treatment plan. However, most approaches used in the discovery of subtypes are not suitable for assigning subtype labels to new cancer specimens from other studies or clinical trials. Here, we address this barrier by applying five different machine learning approaches to multi-omic data from 8,791 TCGA tumor samples comprising 106 subtypes from 26 different cancer cohorts to build models based upon small numbers of features that can classify new samples into previously defined TCGA molecular subtypes-a step toward molecular subtype application in the clinic. We validate select classifiers using external datasets. Predictive performance and classifier-selected features yield insight into the different machine-learning approaches and genomic data platforms. For each cancer and data type we provide containerized versions of the top-performing models as a public resource.
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Affiliation(s)
- Kyle Ellrott
- Oregon Health and Science University, Portland, OR 97239, USA.
| | - Christopher K Wong
- Biomolecular Engineering Department, School of Engineering, University of California, Santa Cruz, Santa Cruz, CA 95064, USA
| | - Christina Yau
- University of California, San Francisco, Department of Surgery, San Francisco, CA 94158, USA; Buck Institute for Research on Aging, Novato, CA 94945, USA
| | - Mauro A A Castro
- Bioinformatics and Systems Biology Laboratory, Federal University of Paraná, Curitiba, PR 81520-260, Brazil
| | - Jordan A Lee
- Oregon Health and Science University, Portland, OR 97239, USA
| | | | - Jasleen K Grewal
- Canada's Michael Smith Genome Sciences Centre, BC Cancer, Vancouver, BC, Canada
| | - Vincenzo Lagani
- JADBio Gnosis DA, GR-700 13 Heraklion, Crete, Greece; Institute of Chemical Biology, Ilia State University, Tbilisi 0162, Georgia
| | - Bahar Tercan
- Institute for Systems Biology, 401 Terry Avenue North, Seattle, WA 98109, USA
| | - Verena Friedl
- Biomolecular Engineering Department, School of Engineering, University of California, Santa Cruz, Santa Cruz, CA 95064, USA
| | - Toshinori Hinoue
- Department of Epigenetics, Van Andel Institute, Grand Rapids, MI 49503, USA
| | - Vladislav Uzunangelov
- Biomolecular Engineering Department, School of Engineering, University of California, Santa Cruz, Santa Cruz, CA 95064, USA
| | - Lindsay Westlake
- The Broad Institute of Harvard and MIT, Cambridge, MA 02142, USA; Department of Medical Oncology, Dana-Farber Cancer Institute, Boston, MA 02215, USA
| | - Xavier Loinaz
- The Broad Institute of Harvard and MIT, Cambridge, MA 02142, USA
| | - Ina Felau
- Center for Cancer Genomics, National Cancer Institute, Bethesda, MD 20892, USA
| | - Peggy I Wang
- Center for Cancer Genomics, National Cancer Institute, Bethesda, MD 20892, USA
| | - Anab Kemal
- Center for Cancer Genomics, National Cancer Institute, Bethesda, MD 20892, USA
| | | | - Ilya Shmulevich
- Institute for Systems Biology, 401 Terry Avenue North, Seattle, WA 98109, USA
| | - Alexander J Lazar
- Departments of Pathology & Genomic Medicine, The University of Texas MD Anderson Cancer Center, Houston, TX 77030, USA
| | - Ioannis Tsamardinos
- JADBio Gnosis DA, GR-700 13 Heraklion, Crete, Greece; Department of Computer Science, University of Crete, GR-700 13 Heraklion, Crete, Greece; Institute of Applied and Computational Mathematics, Foundation for Research and Technology Hellas (FORTH), GR-700 13 Heraklion, Crete, Greece
| | - Katherine A Hoadley
- Department of Genetics, Lineberger Comprehensive Cancer Center, University of North Carolina at Chapel Hill, Chapel Hill, NC 27519, USA
| | - A Gordon Robertson
- Canada's Michael Smith Genome Sciences Centre, BC Cancer, Vancouver, BC, Canada
| | - Theo A Knijnenburg
- Institute for Systems Biology, 401 Terry Avenue North, Seattle, WA 98109, USA
| | | | - Joshua M Stuart
- Biomolecular Engineering Department, School of Engineering, University of California, Santa Cruz, Santa Cruz, CA 95064, USA
| | - Jean C Zenklusen
- Center for Cancer Genomics, National Cancer Institute, Bethesda, MD 20892, USA
| | - Andrew D Cherniack
- The Broad Institute of Harvard and MIT, Cambridge, MA 02142, USA; Department of Medical Oncology, Dana-Farber Cancer Institute, Boston, MA 02215, USA; Harvard Medical School, Boston, MA 02115, USA.
| | - Peter W Laird
- Department of Epigenetics, Van Andel Institute, Grand Rapids, MI 49503, USA.
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6
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Liu XM, Halushka MK. Beyond the Bubble: A Debate on microRNA Sorting Into Extracellular Vesicles. J Transl Med 2025; 105:102206. [PMID: 39647608 PMCID: PMC11842217 DOI: 10.1016/j.labinv.2024.102206] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/01/2024] [Revised: 11/26/2024] [Accepted: 11/26/2024] [Indexed: 12/10/2024] Open
Abstract
Over the past decade, a scientific field has been developed demonstrating microRNAs (miRNAs) to be actively sorted into extracellular vesicles via specific nucleotide motifs that interact with discrete RNA-binding proteins. These miRNAs are proposed to be transported into recipient cells in which they can regulate specific cellular pathways. This mechanism could have enormous potential in explaining how cells signal and regulate other cells nearby or at a distance. Tens of studies have built this theme of a regulated transport of miRNAs. However, some concerns exist about this field. Taken together, there are concerns of a lack of a consistent motif, RNA-binding protein, or preferential miRNA involved in this process. In this study, we provide an expert and extensive analysis of the field that makes the cases for and against an active sorting mechanism. We provide potential explanations on why there is a lack of agreement. Most importantly, we provide ideas on how to move this field forward with more rigor and reproducibility. It is hoped that by engaging in a scientific debate of the pros and cons of this field, more rigorous experiments can be performed to conclusively demonstrate this biological activity.
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Affiliation(s)
- Xiao-Man Liu
- The Stanley Center for Psychiatric Research, The Broad Institute of Massachusetts Institute of Technology and Harvard, Cambridge, Massachusetts
| | - Marc K Halushka
- Pathology and Laboratory Medicine Institute, Cleveland Clinic, Cleveland, Ohio.
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7
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Xuan DTM, Yeh IJ, Liu HL, Su CY, Ko CC, Ta HDK, Jiang JZ, Sun Z, Lin HY, Wang CY, Yen MC. A comparative analysis of Marburg virus-infected bat and human models from public high-throughput sequencing data. Int J Med Sci 2025; 22:1-16. [PMID: 39744175 PMCID: PMC11659840 DOI: 10.7150/ijms.100696] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 07/09/2024] [Accepted: 10/10/2024] [Indexed: 02/01/2025] Open
Abstract
Marburg virus (MARV) disease (MVD) is an uncommon yet serious viral hemorrhagic fever that impacts humans and non-human primates. In humans, infection by the MARV is marked by rapid onset, high transmissibility, and elevated mortality rates, presenting considerable obstacles to the development of vaccines and treatments. Bats, particularly Rousettus aegyptiacus, are suspected to be natural hosts of MARV. Previous research reported asymptomatic MARV infection in bats, in stark contrast to the severe responses observed in humans and other primates. Recent MARV outbreaks highlight significant public health concerns, underscoring the need for gene expression studies during MARV progression. To investigate this, we employed two models from the Gene Expression Omnibus, including kidney cells from Rousettus aegyptiacus and primary proximal tubular cells from Homo sapiens. These models were chosen to identify changes in gene expression profiles and to examine co-regulated genes and pathways involved in MARV disease progression. Our analysis of differentially expressed genes (DEGs) revealed that these genes are mainly associated with pathways related to the complement system, innate immune response via interferons (IFNs), Wnt/β-catenin signaling, and Hedgehog signaling, which played crucial roles in MARV infection across both models. Furthermore, we also identified several potential compounds that may be useful against MARV infection. These findings offer valuable insights into the mechanisms underlying MARV's pathophysiology and suggest potential strategies for preventing transmission, managing post-infection effects, and developing future vaccines.
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Affiliation(s)
- Do Thi Minh Xuan
- Faculty of Pharmacy, Van Lang University, 69/68 Dang Thuy Tram Street, Ward 13, Binh Thanh District, Ho Chi Minh City 70000, Vietnam
| | - I-Jeng Yeh
- Department of Emergency Medicine, Kaohsiung Medical University Hospital, Kaohsiung Medical University, Kaohsiung 80708, Taiwan
- Graduate Institute of Clinical Medicine, College of Medicine, Kaohsiung Medical University, Kaohsiung 80708, Taiwan
| | - Hsin-Liang Liu
- Department of Emergency Medicine, Kaohsiung Medical University Hospital, Kaohsiung Medical University, Kaohsiung 80708, Taiwan
| | - Che-Yu Su
- Department of Emergency Medicine, Kaohsiung Medical University Hospital, Kaohsiung Medical University, Kaohsiung 80708, Taiwan
| | - Ching-Chung Ko
- Department of Medical Imaging, Chi-Mei Medical Center, Tainan, Taiwan
- Department of Health and Nutrition, Chia Nan University of Pharmacy and Science, Tainan, Taiwan
- School of Medicine, College of Medicine, National Sun Yat-Sen University, Kaohsiung, Taiwan
| | - Hoang Dang Khoa Ta
- Ph.D. Program for Cancer Molecular Biology and Drug Discovery, College of Medical Science, Taipei Medical University, Taipei 11031, Taiwan
| | - Jia-Zhen Jiang
- Emergency Department, Huashan Hospital North, Fudan University, Shanghai 201508, People's Republic of China
| | - Zhengda Sun
- Kaiser Permanente, Northern California Regional Laboratories, The Permanente Medical Group, 1725 Eastshore Hwy, Berkeley, CA 94710, USA
| | - Hung-Yun Lin
- Graduate Institute of Cancer Biology and Drug Discovery, College of Medical Science and Technology, Taipei Medical University, Taipei 11031, Taiwan
- Cancer Center, Wan Fang Hospital, Taipei Medical University, Taipei 11031, Taiwan
- TMU Research Center of Cancer Translational Medicine, Taipei Medical University, Taipei 11031, Taiwan
- Traditional Herbal Medicine Research Center of Taipei Medical University Hospital, Taipei Medical University, Taipei 11031, Taiwan
- Pharmaceutical Research Institute, Albany College of Pharmacy and Health Sciences, Rensselaer, NY 12144, USA
| | - Chih-Yang Wang
- Ph.D. Program for Cancer Molecular Biology and Drug Discovery, College of Medical Science, Taipei Medical University, Taipei 11031, Taiwan
- Graduate Institute of Cancer Biology and Drug Discovery, College of Medical Science and Technology, Taipei Medical University, Taipei 11031, Taiwan
- TMU Research Center of Cancer Translational Medicine, Taipei Medical University, Taipei 11031, Taiwan
| | - Meng-Chi Yen
- Department of Emergency Medicine, Kaohsiung Medical University Hospital, Kaohsiung Medical University, Kaohsiung 80708, Taiwan
- Graduate Institute of Clinical Medicine, College of Medicine, Kaohsiung Medical University, Kaohsiung 80708, Taiwan
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8
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Mollaoglu G, Tepper A, Falcomatà C, Potak HT, Pia L, Amabile A, Mateus-Tique J, Rabinovich N, Park MD, LaMarche NM, Brody R, Browning L, Lin JR, Zamarin D, Sorger PK, Santagata S, Merad M, Baccarini A, Brown BD. Ovarian cancer-derived IL-4 promotes immunotherapy resistance. Cell 2024; 187:7492-7510.e22. [PMID: 39481380 PMCID: PMC11682930 DOI: 10.1016/j.cell.2024.10.006] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/17/2023] [Revised: 04/20/2024] [Accepted: 10/03/2024] [Indexed: 11/02/2024]
Abstract
Ovarian cancer is resistant to immunotherapy, and this is influenced by the immunosuppressed tumor microenvironment (TME) dominated by macrophages. Resistance is also affected by intratumoral heterogeneity, whose development is poorly understood. To identify regulators of ovarian cancer immunity, we employed a spatial functional genomics screen (Perturb-map), focused on receptor/ligands hypothesized to be involved in tumor-macrophage communication. Perturb-map recapitulated tumor heterogeneity and revealed that interleukin-4 (IL-4) promotes resistance to anti-PD-1. We find ovarian cancer cells are the key source of IL-4, which directs the formation of an immunosuppressive TME via macrophage control. IL-4 loss was not compensated by nearby IL-4-expressing clones, revealing short-range regulation of TME composition dictating tumor evolution. Our studies show heterogeneous TMEs can emerge from localized altered expression of cancer-derived cytokines/chemokines that establish immune-rich and immune-excluded neighborhoods, which drive clone selection and immunotherapy resistance. They also demonstrate the potential of targeting IL-4 signaling to enhance ovarian cancer response to immunotherapy.
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Affiliation(s)
- Gurkan Mollaoglu
- Icahn Genomics Institute, Icahn School of Medicine at Mount Sinai, New York, NY, USA; Precision Immunology Institute, Icahn School of Medicine at Mount Sinai, New York, NY, USA; Tisch Cancer Institute, Icahn School of Medicine at Mount Sinai, New York, NY, USA; Department of Immunology and Immunotherapy, Icahn School of Medicine at Mount Sinai, New York, NY, USA
| | - Alexander Tepper
- Icahn Genomics Institute, Icahn School of Medicine at Mount Sinai, New York, NY, USA; Precision Immunology Institute, Icahn School of Medicine at Mount Sinai, New York, NY, USA
| | - Chiara Falcomatà
- Icahn Genomics Institute, Icahn School of Medicine at Mount Sinai, New York, NY, USA; Precision Immunology Institute, Icahn School of Medicine at Mount Sinai, New York, NY, USA
| | - Hunter T Potak
- Icahn Genomics Institute, Icahn School of Medicine at Mount Sinai, New York, NY, USA; Precision Immunology Institute, Icahn School of Medicine at Mount Sinai, New York, NY, USA
| | - Luisanna Pia
- Icahn Genomics Institute, Icahn School of Medicine at Mount Sinai, New York, NY, USA; Precision Immunology Institute, Icahn School of Medicine at Mount Sinai, New York, NY, USA
| | - Angelo Amabile
- Icahn Genomics Institute, Icahn School of Medicine at Mount Sinai, New York, NY, USA; Precision Immunology Institute, Icahn School of Medicine at Mount Sinai, New York, NY, USA
| | - Jaime Mateus-Tique
- Icahn Genomics Institute, Icahn School of Medicine at Mount Sinai, New York, NY, USA; Precision Immunology Institute, Icahn School of Medicine at Mount Sinai, New York, NY, USA
| | | | - Matthew D Park
- Precision Immunology Institute, Icahn School of Medicine at Mount Sinai, New York, NY, USA; Tisch Cancer Institute, Icahn School of Medicine at Mount Sinai, New York, NY, USA; Department of Immunology and Immunotherapy, Icahn School of Medicine at Mount Sinai, New York, NY, USA
| | - Nelson M LaMarche
- Precision Immunology Institute, Icahn School of Medicine at Mount Sinai, New York, NY, USA; Tisch Cancer Institute, Icahn School of Medicine at Mount Sinai, New York, NY, USA; Department of Immunology and Immunotherapy, Icahn School of Medicine at Mount Sinai, New York, NY, USA
| | - Rachel Brody
- Department of Pathology, Molecular and Cell-based Medicine, Icahn School of Medicine at Mount Sinai, New York, NY, USA
| | - Lindsay Browning
- Laboratory of Systems Pharmacology, Harvard Medical School, Boston, MA, USA; Department of Pathology, Brigham and Women's Hospital, Harvard Medical School, Boston, MA, USA
| | - Jia-Ren Lin
- Laboratory of Systems Pharmacology, Harvard Medical School, Boston, MA, USA; Department of Systems Biology, Harvard Medical School, Boston, MA, USA; Ludwig Center at Harvard, Boston, MA, USA
| | - Dmitriy Zamarin
- Icahn Genomics Institute, Icahn School of Medicine at Mount Sinai, New York, NY, USA; Precision Immunology Institute, Icahn School of Medicine at Mount Sinai, New York, NY, USA; Tisch Cancer Institute, Icahn School of Medicine at Mount Sinai, New York, NY, USA
| | - Peter K Sorger
- Laboratory of Systems Pharmacology, Harvard Medical School, Boston, MA, USA; Department of Systems Biology, Harvard Medical School, Boston, MA, USA; Ludwig Center at Harvard, Boston, MA, USA
| | - Sandro Santagata
- Laboratory of Systems Pharmacology, Harvard Medical School, Boston, MA, USA; Department of Pathology, Brigham and Women's Hospital, Harvard Medical School, Boston, MA, USA; Department of Systems Biology, Harvard Medical School, Boston, MA, USA; Ludwig Center at Harvard, Boston, MA, USA
| | - Miriam Merad
- Precision Immunology Institute, Icahn School of Medicine at Mount Sinai, New York, NY, USA; Tisch Cancer Institute, Icahn School of Medicine at Mount Sinai, New York, NY, USA; Department of Immunology and Immunotherapy, Icahn School of Medicine at Mount Sinai, New York, NY, USA
| | - Alessia Baccarini
- Icahn Genomics Institute, Icahn School of Medicine at Mount Sinai, New York, NY, USA; Precision Immunology Institute, Icahn School of Medicine at Mount Sinai, New York, NY, USA; Department of Immunology and Immunotherapy, Icahn School of Medicine at Mount Sinai, New York, NY, USA.
| | - Brian D Brown
- Icahn Genomics Institute, Icahn School of Medicine at Mount Sinai, New York, NY, USA; Precision Immunology Institute, Icahn School of Medicine at Mount Sinai, New York, NY, USA; Tisch Cancer Institute, Icahn School of Medicine at Mount Sinai, New York, NY, USA; Department of Immunology and Immunotherapy, Icahn School of Medicine at Mount Sinai, New York, NY, USA.
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9
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Wang L, Xu W, Zhang S, Gundberg GC, Zheng CR, Wan Z, Mustafina K, Caliendo F, Sandt H, Kamm R, Weiss R. Sensing and guiding cell-state transitions by using genetically encoded endoribonuclease-mediated microRNA sensors. Nat Biomed Eng 2024; 8:1730-1743. [PMID: 38982158 DOI: 10.1038/s41551-024-01229-z] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/24/2023] [Accepted: 05/11/2024] [Indexed: 07/11/2024]
Abstract
Precisely sensing and guiding cell-state transitions via the conditional genetic activation of appropriate differentiation factors is challenging. Here we show that desired cell-state transitions can be guided via genetically encoded sensors, whereby endogenous cell-state-specific miRNAs regulate the translation of a constitutively transcribed endoribonuclease, which, in turn, controls the translation of a gene of interest. We used this approach to monitor several cell-state transitions, to enrich specific cell types and to automatically guide the multistep differentiation of human induced pluripotent stem cells towards a haematopoietic lineage via endothelial cells as an intermediate state. Such conditional activation of gene expression is durable and resistant to epigenetic silencing and could facilitate the monitoring of cell-state transitions in physiological and pathological conditions and eventually the 'rewiring' of cell-state transitions for applications in organoid-based disease modelling, cellular therapies and regenerative medicine.
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Affiliation(s)
- Lei Wang
- Department of Biological Engineering, Massachusetts Institute of Technology, Cambridge, MA, USA.
- Synthetic Biology Center, Massachusetts Institute of Technology, Cambridge, MA, USA.
- Department of Bioengineering, Northeastern University, Boston, MA, USA.
- Department of Biology, Northeastern University, Boston, MA, USA.
| | - Wenlong Xu
- Department of Biological Engineering, Massachusetts Institute of Technology, Cambridge, MA, USA
- Synthetic Biology Center, Massachusetts Institute of Technology, Cambridge, MA, USA
| | - Shun Zhang
- Department of Biological Engineering, Massachusetts Institute of Technology, Cambridge, MA, USA
- Beijing Institute for Stem Cell and Regenerative Medicine, Beijing, China
- State Key Laboratory of Organ Regeneration and Reconstruction, Institute of Zoology, Chinese Academy of Sciences, Beijing, China
| | - Gregory C Gundberg
- Department of Biological Engineering, Massachusetts Institute of Technology, Cambridge, MA, USA
- Synthetic Biology Center, Massachusetts Institute of Technology, Cambridge, MA, USA
| | - Christine R Zheng
- Department of Biological Engineering, Massachusetts Institute of Technology, Cambridge, MA, USA
| | - Zhengpeng Wan
- Department of Biological Engineering, Massachusetts Institute of Technology, Cambridge, MA, USA
| | - Kamila Mustafina
- Department of Biological Engineering, Massachusetts Institute of Technology, Cambridge, MA, USA
- Synthetic Biology Center, Massachusetts Institute of Technology, Cambridge, MA, USA
| | - Fabio Caliendo
- Department of Biological Engineering, Massachusetts Institute of Technology, Cambridge, MA, USA
- Synthetic Biology Center, Massachusetts Institute of Technology, Cambridge, MA, USA
| | - Hayden Sandt
- Department of Biological Engineering, Massachusetts Institute of Technology, Cambridge, MA, USA
- Synthetic Biology Center, Massachusetts Institute of Technology, Cambridge, MA, USA
| | - Roger Kamm
- Department of Biological Engineering, Massachusetts Institute of Technology, Cambridge, MA, USA
- Department of Mechanical Engineering, Massachusetts Institute of Technology, Cambridge, MA, USA
| | - Ron Weiss
- Department of Biological Engineering, Massachusetts Institute of Technology, Cambridge, MA, USA.
- Synthetic Biology Center, Massachusetts Institute of Technology, Cambridge, MA, USA.
- Department of Electrical Engineering and Computer Science, Massachusetts Institute of Technology, Cambridge, MA, USA.
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10
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Oesinghaus L, Castillo-Hair S, Ludwig N, Keller A, Seelig G. Quantitative design of cell type-specific mRNA stability from microRNA expression data. BIORXIV : THE PREPRINT SERVER FOR BIOLOGY 2024:2024.10.28.620728. [PMID: 39554011 PMCID: PMC11565874 DOI: 10.1101/2024.10.28.620728] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Subscribe] [Scholar Register] [Indexed: 11/19/2024]
Abstract
Limiting expression to target cell types is a longstanding goal in gene therapy, which could be met by sensing endogenous microRNA. However, an unclear association between microRNA expression and activity currently hampers such an approach. Here, we probe this relationship by measuring the stability of synthetic microRNA-responsive 3'UTRs across 10 cell lines in a library format. By systematically addressing biases in microRNA expression data and confounding factors such as microRNA crosstalk, we demonstrate that a straightforward model can quantitatively predict reporter stability purely from expression data. We use this model to design constructs with previously unattainable response patterns across our cell lines. The rules we derive for microRNA expression data selection and processing should apply to microRNA- responsive devices for any environment with available expression data.
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11
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Santiago FE, Adige T, Mahmud S, Dong X, Niedernhofer LJ, Robbins PD. miR-96-5p expression is sufficient to induce and maintain the senescent cell fate in the absence of stress. Proc Natl Acad Sci U S A 2024; 121:e2321182121. [PMID: 39325426 PMCID: PMC11459134 DOI: 10.1073/pnas.2321182121] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/22/2023] [Accepted: 08/08/2024] [Indexed: 09/27/2024] Open
Abstract
Senescence is a cell fate driven by different types of stress that results in exit from the cell cycle and expression of an inflammatory senescence-associated secretory phenotype (SASP). Here, we demonstrate that stable overexpression of miR-96-5p was sufficient to induce cellular senescence in the absence of genotoxic stress, inducing expression of certain markers of early senescence including SASP factors while repressing markers of deep senescence including LINE-1 and type 1 interferons. Stable miR-96-5p overexpression led to genome-wide changes in heterochromatin followed by epigenetic activation of p16Ink4a, p21Cip1, and SASP expression, induction of a marker of DNA damage, and induction of a transcriptional signature similar to other senescent lung and endothelial cell types. Expression of miR-96-5p significantly increased following senescence induction in culture cells and with aging in tissues from naturally aged and Ercc1-/Δ progeroid mice. Mechanistically, miR-96-5p directly suppressed expression of SIN3B and SIN3 corepressor complex constituents KDM5A and MORF4L2, and siRNA-mediated knockdown of these transcriptional regulators recapitulated the senescent phenotype. In addition, pharmacologic inhibition of the SIN3 complex suppressed senescence and SASP markers. These results clearly demonstrate that a single microRNA is sufficient to drive early senescence in the absence of genotoxic stress through targeting epigenetic and transcriptional regulators, identifying novel targets for the development of senotherapeutics.
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Affiliation(s)
- Fernando E. Santiago
- Institute on the Biology of Aging and Metabolism, University of Minnesota, Minnesota, MN55455
- Department of Biochemistry, Molecular Biology and Biophysics, University of Minnesota, Minnesota, MN55455
| | - Tanvi Adige
- Institute on the Biology of Aging and Metabolism, University of Minnesota, Minnesota, MN55455
- Department of Biochemistry, Molecular Biology and Biophysics, University of Minnesota, Minnesota, MN55455
| | - Shamsed Mahmud
- Institute on the Biology of Aging and Metabolism, University of Minnesota, Minnesota, MN55455
- Department of Genetics, Cell Biology and Development, University of Minnesota, MinnesotaMN55455
| | - Xiao Dong
- Institute on the Biology of Aging and Metabolism, University of Minnesota, Minnesota, MN55455
- Department of Genetics, Cell Biology and Development, University of Minnesota, MinnesotaMN55455
| | - Laura J. Niedernhofer
- Institute on the Biology of Aging and Metabolism, University of Minnesota, Minnesota, MN55455
- Department of Biochemistry, Molecular Biology and Biophysics, University of Minnesota, Minnesota, MN55455
| | - Paul D. Robbins
- Institute on the Biology of Aging and Metabolism, University of Minnesota, Minnesota, MN55455
- Department of Biochemistry, Molecular Biology and Biophysics, University of Minnesota, Minnesota, MN55455
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12
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Seitz H. A new perspective on microRNA-guided gene regulation specificity, and its potential generalization to transcription factors and RNA-binding proteins. Nucleic Acids Res 2024; 52:9360-9368. [PMID: 39149906 PMCID: PMC11381331 DOI: 10.1093/nar/gkae694] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/03/2024] [Revised: 07/25/2024] [Accepted: 07/31/2024] [Indexed: 08/17/2024] Open
Abstract
Our conception of gene regulation specificity has undergone profound changes over the last 20 years. Previously, regulators were considered to control few genes, recognized with exquisite specificity by a 'lock and key' mechanism. However, recently genome-wide exploration of regulator binding site occupancy (whether on DNA or RNA targets) revealed extensive lists of molecular targets for every studied regulator. Such poor biochemical specificity suggested that each regulator controls many genes, collectively contributing to biological phenotypes. Here, I propose a third model, whereby regulators' biological specificity is only partially due to 'lock and key' biochemistry. Rather, regulators affect many genes at the microscopic scale, but biological consequences for most interactions are attenuated at the mesoscopic scale: only a few regulatory events propagate from microscopic to macroscopic scale; others are made inconsequential by homeostatic mechanisms. This model is well supported by the microRNA literature, and data suggest that it extends to other regulators. It reconciles contradicting observations from biochemistry and comparative genomics on one hand and in vivo genetics on the other hand, but this conceptual unification is obscured by common misconceptions and counter-intuitive modes of graphical display. Profound understanding of gene regulation requires conceptual clarification, and better suited statistical analyses and graphical representation.
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Affiliation(s)
- Hervé Seitz
- Institut de Génétique Humaine (UMR 9002), CNRS, 141, rue de la Cardonille, 34396 Montpellier, France
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13
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Confuorti C, Jaramillo M, Plante I. Hormonal regulation of miRNA during mammary gland development. Biol Open 2024; 13:bio060308. [PMID: 38712984 PMCID: PMC11190577 DOI: 10.1242/bio.060308] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/11/2024] [Accepted: 04/30/2024] [Indexed: 05/08/2024] Open
Abstract
The mammary gland is a unique organ as most of its development occurs after birth through stages of proliferation, differentiation and apoptosis that are tightly regulated by circulating hormones and growth factors. Throughout development, hormonal cues induce the regulation of different pathways, ultimately leading to differential transcription and expression of genes involved in this process, but also in the activation or inhibition of post-transcriptional mechanisms of regulation. However, the role of microRNAs (miRNAs) in the different phases of mammary gland remodeling is still poorly understood. The objectives of this study were to analyze the expression of miRNA in key stages of mammary gland development in mice and to determine whether it could be associated with hormonal variation between stages. To do so, miRNAs were isolated from mouse mammary glands at stages of adulthood, pregnancy, lactation and involution, and sequenced. Results showed that 490, 473, 419, and 460 miRNAs are detected in adult, pregnant, lactating and involuting mice, respectively, most of them being common to all four groups, and 58 unique to one stage. Most genes could be divided into six clusters of expression, including two encompassing the highest number of miRNA (clusters 1 and 3) and showing opposite profiles of expression, reaching a peak at adulthood and valley at lactation, or showing the lowest expression at adulthood and peaking at lactation. GO and KEGG analyses suggest that the miRNAs differentially expressed between stages influence the expression of targets associated with mammary gland homeostasis and hormone regulation. To further understand the links between miRNA expression and hormones involved in mammary gland development, miRNAs were then sequenced in breast cells exposed to estradiol, progesterone, prolactin and oxytocin. Four, 38, 24 and 66 miRNAs were associated with progesterone, estradiol, prolactin, and oxytocin exposure, respectively. Finally, when looking at miRNAs modulated by the hormones, differentially expressed during mammary gland development, and having a pattern of expression that could be correlated with the relative levels of hormones known to be found in vivo, 16 miRNAs were identified as likely regulated by circulating hormones. Overall, our study brings a better understanding of the regulation of miRNAs throughout mammary gland development and suggests that there is a relationship between their expression and the main hormones involved in mammary gland development. Future studies will examine this role more in detail.
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Affiliation(s)
- Cameron Confuorti
- INRS, Centre Armand-Frappier Santé Biotechnologie, 531 boul. des Prairies, Laval, QC, H7V 1B7, Canada
| | - Maritza Jaramillo
- INRS, Centre Armand-Frappier Santé Biotechnologie, 531 boul. des Prairies, Laval, QC, H7V 1B7, Canada
| | - Isabelle Plante
- INRS, Centre Armand-Frappier Santé Biotechnologie, 531 boul. des Prairies, Laval, QC, H7V 1B7, Canada
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14
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Czubala MA, Jenkins RH, Gurney M, Wallace L, Cossins B, Dennis J, Rosas M, Andrews R, Fraser D, Taylor PR. Tissue-specific transcriptional programming of macrophages controls the microRNA transcriptome targeting multiple functional pathways. J Biol Chem 2024; 300:107244. [PMID: 38556087 PMCID: PMC11067537 DOI: 10.1016/j.jbc.2024.107244] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/17/2024] [Revised: 03/22/2024] [Accepted: 03/25/2024] [Indexed: 04/02/2024] Open
Abstract
Recent interest in the biology and function of peritoneal tissue resident macrophages (pMΦ) has led to a better understanding of their cellular origin, programming, and renewal. The programming of pMΦ is dependent on microenvironmental cues and tissue-specific transcription factors, including GATA6. However, the contribution of microRNAs remains poorly defined. We conducted a detailed analysis of the impact of GATA6 deficiency on microRNA expression in mouse pMΦ. Our data suggest that for many of the pMΦ, microRNA composition may be established during tissue specialization and that the effect of GATA6 knockout is largely unable to be rescued in the adult by exogenous GATA6. The data are consistent with GATA6 modulating the expression pattern of specific microRNAs, directly or indirectly, and including miR-146a, miR-223, and miR-203 established by the lineage-determining transcription factor PU.1, to achieve a differentiated pMΦ phenotype. Lastly, we showed a significant dysregulation of miR-708 in pMΦ in the absence of GATA6 during homeostasis and in response to LPS/IFN-γ stimulation. Overexpression of miR-708 in mouse pMΦ in vivo altered 167 mRNA species demonstrating functional downregulation of predicted targets, including cell immune responses and cell cycle regulation. In conclusion, we demonstrate dependence of the microRNA transcriptome on tissue-specific programming of tissue macrophages as exemplified by the role of GATA6 in pMΦ specialization.
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Affiliation(s)
- Magdalena A Czubala
- Systems Immunity Research Institute and Division of Infection and Immunity, Cardiff University, Cardiff, UK.
| | - Robert H Jenkins
- Systems Immunity Research Institute and Division of Infection and Immunity, Cardiff University, Cardiff, UK
| | - Mark Gurney
- Systems Immunity Research Institute and Division of Infection and Immunity, Cardiff University, Cardiff, UK
| | - Leah Wallace
- Systems Immunity Research Institute and Division of Infection and Immunity, Cardiff University, Cardiff, UK
| | - Benjamin Cossins
- Systems Immunity Research Institute and Division of Infection and Immunity, Cardiff University, Cardiff, UK
| | - James Dennis
- Systems Immunity Research Institute and Division of Infection and Immunity, Cardiff University, Cardiff, UK
| | - Marcela Rosas
- Systems Immunity Research Institute and Division of Infection and Immunity, Cardiff University, Cardiff, UK
| | - Robert Andrews
- Systems Immunity Research Institute and Division of Infection and Immunity, Cardiff University, Cardiff, UK
| | - Donald Fraser
- Systems Immunity Research Institute and Division of Infection and Immunity, Cardiff University, Cardiff, UK; Wales Kidney Research Unit, Cardiff University, Cardiff, UK
| | - Philip R Taylor
- Systems Immunity Research Institute and Division of Infection and Immunity, Cardiff University, Cardiff, UK; UK Dementia Research Institute at Cardiff, Cardiff University, Cardiff, UK.
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15
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Nkechika V, Zhang N, Belsham DD. The Involvement of the microRNAs miR-466c and miR-340 in the Palmitate-Mediated Dysregulation of Gonadotropin-Releasing Hormone Gene Expression. Genes (Basel) 2024; 15:397. [PMID: 38674332 PMCID: PMC11048885 DOI: 10.3390/genes15040397] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/25/2024] [Revised: 03/18/2024] [Accepted: 03/21/2024] [Indexed: 04/28/2024] Open
Abstract
Diets high in saturated fatty acids are associated with obesity and infertility. Palmitate, the most prevalent circulating saturated fatty acid, is sensed by hypothalamic neurons, contributing to homeostatic dysregulation. Notably, palmitate elevates the mRNA levels of gonadotropin-releasing hormone (Gnrh) mRNA and its activating transcription factor, GATA binding protein 4 (Gata4). GATA4 is essential for basal Gnrh expression by binding to its enhancer region, with Oct-1 (Oct1) and CEBP-β (Cebpb) playing regulatory roles. The pre- and post-transcriptional control of Gnrh by palmitate have not been investigated. Given the ability of palmitate to alter microRNAs (miRNAs), we hypothesized that palmitate-mediated dysregulation of Gnrh mRNA involves specific miRNAs. In the mHypoA-GnRH/GFP neurons, palmitate significantly downregulated six miRNAs (miR-125a, miR-181b, miR-340, miR-351, miR-466c and miR-503), and the repression was attenuated by co-treatment with 100 μM of oleate. Subsequent mimic transfections revealed that miR-466c significantly downregulates Gnrh, Gata4, and Chop mRNA and increases Per2, whereas miR-340 upregulates Gnrh, Gata4, Oct1, Cebpb, and Per2 mRNA. Our findings suggest that palmitate may indirectly regulate Gnrh at both the pre- and post-transcriptional levels by altering miR-466c and miR-340, which in turn regulate transcription factor expression levels. In summary, palmitate-mediated dysregulation of Gnrh and, consequently, reproductive function involves parallel transcriptional mechanisms.
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Affiliation(s)
- Vanessa Nkechika
- Department of Physiology, University of Toronto, 1 King’s College Circle, Toronto, ON M5S 1A8, Canada; (V.N.); (N.Z.)
| | - Ningtong Zhang
- Department of Physiology, University of Toronto, 1 King’s College Circle, Toronto, ON M5S 1A8, Canada; (V.N.); (N.Z.)
| | - Denise D. Belsham
- Department of Physiology, University of Toronto, 1 King’s College Circle, Toronto, ON M5S 1A8, Canada; (V.N.); (N.Z.)
- Department of Medicine, University of Toronto, 1 King’s College Circle, Toronto, ON M5S 1A8, Canada
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16
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Chen SJ, Hashimoto K, Fujio K, Hayashi K, Paul SK, Yuzuriha A, Qiu WY, Nakamura E, Kanashiro MA, Kabata M, Nakamura S, Sugimoto N, Kaneda A, Yamamoto T, Saito H, Takayama N, Eto K. A let-7 microRNA-RALB axis links the immune properties of iPSC-derived megakaryocytes with platelet producibility. Nat Commun 2024; 15:2588. [PMID: 38519457 PMCID: PMC10960040 DOI: 10.1038/s41467-024-46605-0] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/06/2023] [Accepted: 03/04/2024] [Indexed: 03/25/2024] Open
Abstract
We recently achieved the first-in-human transfusion of induced pluripotent stem cell-derived platelets (iPSC-PLTs) as an alternative to standard transfusions, which are dependent on donors and therefore variable in supply. However, heterogeneity characterized by thrombopoiesis-biased or immune-biased megakaryocytes (MKs) continues to pose a bottleneck against the standardization of iPSC-PLT manufacturing. To address this problem, here we employ microRNA (miRNA) switch biotechnology to distinguish subpopulations of imMKCLs, the MK cell lines producing iPSC-PLTs. Upon miRNA switch-based screening, we find imMKCLs with lower let-7 activity exhibit an immune-skewed transcriptional signature. Notably, the low activity of let-7a-5p results in the upregulation of RAS like proto-oncogene B (RALB) expression, which is crucial for the lineage determination of immune-biased imMKCL subpopulations and leads to the activation of interferon-dependent signaling. The dysregulation of immune properties/subpopulations, along with the secretion of inflammatory cytokines, contributes to a decline in the quality of the whole imMKCL population.
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Affiliation(s)
- Si Jing Chen
- Department of Clinical Application, Center for iPS Cell Research and Application (CiRA), Kyoto University, Kyoto, Japan
- Department of Regenerative Medicine, Graduate School of Medicine, Chiba University, Chiba, Japan
| | - Kazuya Hashimoto
- Department of Clinical Application, Center for iPS Cell Research and Application (CiRA), Kyoto University, Kyoto, Japan
| | - Kosuke Fujio
- Department of Clinical Application, Center for iPS Cell Research and Application (CiRA), Kyoto University, Kyoto, Japan
| | - Karin Hayashi
- Department of Life Science Frontiers, Center for iPS Cell Research and Application (CiRA), Kyoto University, Kyoto, Japan
| | - Sudip Kumar Paul
- Department of Regenerative Medicine, Graduate School of Medicine, Chiba University, Chiba, Japan
| | - Akinori Yuzuriha
- Department of Clinical Application, Center for iPS Cell Research and Application (CiRA), Kyoto University, Kyoto, Japan
| | - Wei-Yin Qiu
- Department of Clinical Application, Center for iPS Cell Research and Application (CiRA), Kyoto University, Kyoto, Japan
| | - Emiri Nakamura
- Department of Clinical Application, Center for iPS Cell Research and Application (CiRA), Kyoto University, Kyoto, Japan
| | | | - Mio Kabata
- Department of Life Science Frontiers, Center for iPS Cell Research and Application (CiRA), Kyoto University, Kyoto, Japan
| | - Sou Nakamura
- Department of Clinical Application, Center for iPS Cell Research and Application (CiRA), Kyoto University, Kyoto, Japan
| | - Naoshi Sugimoto
- Department of Clinical Application, Center for iPS Cell Research and Application (CiRA), Kyoto University, Kyoto, Japan
| | - Atsushi Kaneda
- Department of Molecular Oncology, Graduate School of Medicine, Chiba University, Chiba, Japan
| | - Takuya Yamamoto
- Department of Life Science Frontiers, Center for iPS Cell Research and Application (CiRA), Kyoto University, Kyoto, Japan
- Institute for the Advanced Study of Human Biology (WPI-ASHBi), Kyoto University, Kyoto, Japan
- Medical-risk Avoidance Based on iPS Cells Team, RIKEN Center for Advanced Intelligence Project (AIP), Kyoto, Japan
| | - Hirohide Saito
- Department of Life Science Frontiers, Center for iPS Cell Research and Application (CiRA), Kyoto University, Kyoto, Japan.
| | - Naoya Takayama
- Department of Regenerative Medicine, Graduate School of Medicine, Chiba University, Chiba, Japan.
| | - Koji Eto
- Department of Clinical Application, Center for iPS Cell Research and Application (CiRA), Kyoto University, Kyoto, Japan.
- Department of Regenerative Medicine, Graduate School of Medicine, Chiba University, Chiba, Japan.
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17
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Huang W, Paul D, Calin GA, Bayraktar R. miR-142: A Master Regulator in Hematological Malignancies and Therapeutic Opportunities. Cells 2023; 13:84. [PMID: 38201290 PMCID: PMC10778542 DOI: 10.3390/cells13010084] [Citation(s) in RCA: 6] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/25/2023] [Revised: 11/29/2023] [Accepted: 12/15/2023] [Indexed: 01/12/2024] Open
Abstract
MicroRNAs (miRNAs) are a type of non-coding RNA whose dysregulation is frequently associated with the onset and progression of human cancers. miR-142, an ultra-conserved miRNA with both active -3p and -5p mature strands and wide-ranging physiological targets, has been the subject of countless studies over the years. Due to its preferential expression in hematopoietic cells, miR-142 has been found to be associated with numerous types of lymphomas and leukemias. This review elucidates the multifaceted role of miR-142 in human physiology, its influence on hematopoiesis and hematopoietic cells, and its intriguing involvement in exosome-mediated miR-142 transport. Moreover, we offer a comprehensive exploration of the genetic and molecular landscape of the miR-142 genomic locus, highlighting its mutations and dysregulation within hematological malignancies. Finally, we discuss potential avenues for harnessing the therapeutic potential of miR-142 in the context of hematological malignancies.
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Affiliation(s)
- Wilson Huang
- Department of Translational Molecular Pathology, The University of Texas MD Anderson Cancer Center, Houston, TX 77030, USA; (W.H.); (G.A.C.)
| | - Doru Paul
- Division of Hematology and Medical Oncology, Department of Medicine, Weill Cornell Medicine, New York, NY 10065, USA;
| | - George A. Calin
- Department of Translational Molecular Pathology, The University of Texas MD Anderson Cancer Center, Houston, TX 77030, USA; (W.H.); (G.A.C.)
- Center for RNA Interference and Non-Coding RNAs, The University of Texas MD Anderson Cancer Center, Houston, TX 77030, USA
- Department of Leukemia, Division of Cancer Medicine, The University of Texas MD Anderson Cancer Center, Houston, TX 77030, USA
| | - Recep Bayraktar
- Center for RNA Interference and Non-Coding RNAs, The University of Texas MD Anderson Cancer Center, Houston, TX 77030, USA
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18
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Kaeffer B. Human Breast Milk miRNAs: Their Diversity and Potential for Preventive Strategies in Nutritional Therapy. Int J Mol Sci 2023; 24:16106. [PMID: 38003296 PMCID: PMC10671413 DOI: 10.3390/ijms242216106] [Citation(s) in RCA: 5] [Impact Index Per Article: 2.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/02/2023] [Revised: 11/02/2023] [Accepted: 11/06/2023] [Indexed: 11/26/2023] Open
Abstract
The endogenous miRNAs of breast milk are the products of more than 1000 nonprotein-coding genes, giving rise to mature small regulatory molecules of 19-25 nucleotides. They are incorporated in macromolecular complexes, loaded on Argonaute proteins, sequestrated in exosomes and lipid complexes, or present in exfoliated cells of epithelial, endothelial, or immune origins. Their expression is dependent on the stage of lactation; however, their detection depends on progress in RNA sequencing and the reappraisal of the definition of small RNAs. Some miRNAs from plants are detected in breast milk, opening the possibility of the stimulation of immune cells from the allergy repertoire. Each miRNA harbors a seeding sequence, which targets mRNAs, gene promoters, or long noncoding RNAs. Their activities depend on their bioavailability. Efficient doses of miRNAs are estimated to be roughly 100 molecules in the cytoplasm of target cells from in vitro and in vivo experiments. Each miRNA is included in networks of stimulation/inhibition/sequestration, driving the expression of cellular phenotypes. Three types of stress applied during lactation to manipulate miRNA supply were explored using rodent offspring: a foster mother, a cafeteria diet, and early weaning. This review presents the main mature miRNAs described from current mothers' cohorts and their bioavailability in experimental models as well as studies assessing the potential of miR-26 or miR-320 miRNA families to alter offspring phenotypes.
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Affiliation(s)
- Bertrand Kaeffer
- Nantes Université, INRAE, UMR 1280, PhAN, F-44000 Nantes, France
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19
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Yun Y, Wu R, He X, Qin X, Chen L, Sha L, Yun X, Nishiumi T, Borjigin G. Integrated Transcriptome Analysis of miRNAs and mRNAs in the Skeletal Muscle of Wuranke Sheep. Genes (Basel) 2023; 14:2034. [PMID: 38002977 PMCID: PMC10671749 DOI: 10.3390/genes14112034] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/08/2023] [Revised: 10/26/2023] [Accepted: 10/30/2023] [Indexed: 11/26/2023] Open
Abstract
MicroRNAs (miRNAs) are regarded as important regulators in skeletal muscle development. To reveal the regulatory roles of miRNAs and their target mRNAs underlying the skeletal muscle development of Wuranke sheep, we investigated the miRNA and mRNA expression profiles in the biceps femoris of these sheep at the fetal (3 months of gestation) and 3- and 15-month-old postnatal stages. Consequently, a total of 1195 miRNAs and 24,959 genes were identified. Furthermore, 474, 461, and 54 differentially expressed miRNAs (DEMs) and 6783, 7407, and 78 differentially expressed genes (DEGs) were detected among three comparative groups. Functional analysis demonstrated that the target mRNAs of the DEMs were enriched in multiple pathways related to muscle development. Moreover, the interactions among several predicted miRNA-mRNA pairs (oar-miR-133-HDAC1, oar-miR-1185-5p-MYH1/HADHA/OXCT1, and PC-5p-3703_578-INSR/ACTG1) that potentially affect skeletal muscle development were verified using dual-luciferase reporter assays. In this study, we identified the miRNA and mRNA differences in the skeletal muscle of Wuranke sheep at different developmental stages and revealed that a series of candidate miRNA-mRNA pairs may act as modulators of muscle development. These results will contribute to future studies on the function of miRNAs and their target mRNAs during skeletal muscle development in Wuranke sheep.
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Affiliation(s)
- Yueying Yun
- College of Food Science and Engineering, Inner Mongolia Agricultural University, Hohhot 010018, China; (Y.Y.); (X.H.); (X.Q.); (L.C.); (L.S.); (X.Y.)
- School of Life Science and Technology, Inner Mongolia University of Science and Technology, Baotou 014010, China
| | - Rihan Wu
- College of Biochemistry and Engineering, Hohhot Vocational College, Hohhot 010051, China;
| | - Xige He
- College of Food Science and Engineering, Inner Mongolia Agricultural University, Hohhot 010018, China; (Y.Y.); (X.H.); (X.Q.); (L.C.); (L.S.); (X.Y.)
| | - Xia Qin
- College of Food Science and Engineering, Inner Mongolia Agricultural University, Hohhot 010018, China; (Y.Y.); (X.H.); (X.Q.); (L.C.); (L.S.); (X.Y.)
| | - Lu Chen
- College of Food Science and Engineering, Inner Mongolia Agricultural University, Hohhot 010018, China; (Y.Y.); (X.H.); (X.Q.); (L.C.); (L.S.); (X.Y.)
| | - Lina Sha
- College of Food Science and Engineering, Inner Mongolia Agricultural University, Hohhot 010018, China; (Y.Y.); (X.H.); (X.Q.); (L.C.); (L.S.); (X.Y.)
| | - Xueyan Yun
- College of Food Science and Engineering, Inner Mongolia Agricultural University, Hohhot 010018, China; (Y.Y.); (X.H.); (X.Q.); (L.C.); (L.S.); (X.Y.)
| | - Tadayuki Nishiumi
- Division of Life and Food Science, Graduate School of Science and Technology, Niigata University, Niigata 950-2181, Japan
| | - Gerelt Borjigin
- College of Food Science and Engineering, Inner Mongolia Agricultural University, Hohhot 010018, China; (Y.Y.); (X.H.); (X.Q.); (L.C.); (L.S.); (X.Y.)
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20
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Zhang J, Liu L, Wei X, Zhao C, Li S, Li J, Le TD. Pan-cancer characterization of ncRNA synergistic competition uncovers potential carcinogenic biomarkers. PLoS Comput Biol 2023; 19:e1011308. [PMID: 37812646 PMCID: PMC10586676 DOI: 10.1371/journal.pcbi.1011308] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/29/2023] [Revised: 10/19/2023] [Accepted: 09/25/2023] [Indexed: 10/11/2023] Open
Abstract
Non-coding RNAs (ncRNAs) act as important modulators of gene expression and they have been confirmed to play critical roles in the physiology and development of malignant tumors. Understanding the synergism of multiple ncRNAs in competing endogenous RNA (ceRNA) regulation can provide important insights into the mechanisms of malignant tumors caused by ncRNA regulation. In this work, we present a framework, SCOM, for identifying ncRNA synergistic competition. We systematically construct the landscape of ncRNA synergistic competition across 31 malignant tumors, and reveal that malignant tumors tend to share hub ncRNAs rather than the ncRNA interactions involved in the synergistic competition. In addition, the synergistic competition ncRNAs (i.e. ncRNAs involved in the synergistic competition) are likely to be involved in drug resistance, contribute to distinguishing molecular subtypes of malignant tumors, and participate in immune regulation. Furthermore, SCOM can help to infer ncRNA synergistic competition across malignant tumors and uncover potential diagnostic and prognostic biomarkers of malignant tumors. Altogether, the SCOM framework (https://github.com/zhangjunpeng411/SCOM/) and the resulting web-based database SCOMdb (https://comblab.cn/SCOMdb/) serve as a useful resource for exploring ncRNA regulation and to accelerate the identification of carcinogenic biomarkers.
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Affiliation(s)
- Junpeng Zhang
- School of Engineering, Dali University, Dali, Yunnan, People’s Republic of China
| | - Lin Liu
- UniSA STEM, University of South Australia, Mawson Lakes, South Australia, Australia
| | - Xuemei Wei
- School of Engineering, Dali University, Dali, Yunnan, People’s Republic of China
| | - Chunwen Zhao
- School of Engineering, Dali University, Dali, Yunnan, People’s Republic of China
| | - Sijing Li
- School of Engineering, Dali University, Dali, Yunnan, People’s Republic of China
| | - Jiuyong Li
- UniSA STEM, University of South Australia, Mawson Lakes, South Australia, Australia
| | - Thuc Duy Le
- UniSA STEM, University of South Australia, Mawson Lakes, South Australia, Australia
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21
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Chen X, Sokirniy I, Wang X, Jiang M, Mseis-Jackson N, Williams C, Mayes K, Jiang N, Puls B, Du Q, Shi Y, Li H. MicroRNA-375 Is Induced during Astrocyte-to-Neuron Reprogramming and Promotes Survival of Reprogrammed Neurons when Overexpressed. Cells 2023; 12:2202. [PMID: 37681934 PMCID: PMC10486704 DOI: 10.3390/cells12172202] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/25/2023] [Revised: 08/25/2023] [Accepted: 09/01/2023] [Indexed: 09/09/2023] Open
Abstract
While astrocyte-to-neuron (AtN) reprogramming holds great promise in regenerative medicine, the molecular mechanisms that govern this unique biological process remain elusive. To understand the function of miRNAs during the AtN reprogramming process, we performed RNA-seq of both mRNAs and miRNAs on human astrocyte (HA) cultures upon NeuroD1 overexpression. Bioinformatics analyses showed that NeuroD1 not only activated essential neuronal genes to initiate the reprogramming process but also induced miRNA changes in HA. Among the upregulated miRNAs, we identified miR-375 and its targets, neuronal ELAVL genes (nELAVLs), which encode a family of RNA-binding proteins and were also upregulated by NeuroD1. We further showed that manipulating the miR-375 level regulated nELAVLs' expression during NeuroD1-mediated reprogramming. Interestingly, miR-375/nELAVLs were also induced by the reprogramming factors Neurog2 and ASCL1 in HA, suggesting a conserved function to neuronal reprogramming, and by NeuroD1 in the mouse astrocyte culture and spinal cord. Functionally, we showed that miR-375 overexpression improved NeuroD1-mediated reprogramming efficiency by promoting cell survival at early stages in HA and did not appear to compromise the maturation of the reprogrammed neurons. Lastly, overexpression of miR-375-refractory ELAVL4 induced apoptosis and reversed the cell survival-promoting effect of miR-375 during AtN reprogramming. Together, we demonstrated a neuroprotective role of miR-375 during NeuroD1-mediated AtN reprogramming.
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Affiliation(s)
- Xuanyu Chen
- Department of Neuroscience & Regenerative Medicine, Medical College of Georgia at Augusta University, Augusta, GA 30912, USA
| | - Ivan Sokirniy
- Department of Biomedical Engineering, The Pennsylvania State University, University Park, PA 16802, USA
| | - Xin Wang
- Department of Biology, The Pennsylvania State University, University Park, PA 16802, USA
| | - Mei Jiang
- Department of Neuroscience & Regenerative Medicine, Medical College of Georgia at Augusta University, Augusta, GA 30912, USA
| | - Natalie Mseis-Jackson
- Department of Neuroscience & Regenerative Medicine, Medical College of Georgia at Augusta University, Augusta, GA 30912, USA
| | - Christine Williams
- Department of Chemistry & Biochemistry, College of Science & Mathematics, Augusta University, Augusta, GA 30912, USA
| | - Kristopher Mayes
- Department of Neuroscience & Regenerative Medicine, Medical College of Georgia at Augusta University, Augusta, GA 30912, USA
| | - Na Jiang
- Department of Neuroscience & Regenerative Medicine, Medical College of Georgia at Augusta University, Augusta, GA 30912, USA
| | - Brendan Puls
- Department of Biology, The Pennsylvania State University, University Park, PA 16802, USA
| | - Quansheng Du
- Department of Neuroscience & Regenerative Medicine, Medical College of Georgia at Augusta University, Augusta, GA 30912, USA
| | - Yang Shi
- Department of Neuroscience & Regenerative Medicine, Medical College of Georgia at Augusta University, Augusta, GA 30912, USA
- Division of Biostatistics and Data Science, Department of Population Health Sciences, Medical College of Georgia at Augusta University, Augusta, GA 30912, USA
| | - Hedong Li
- Department of Neuroscience & Regenerative Medicine, Medical College of Georgia at Augusta University, Augusta, GA 30912, USA
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22
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Kan Changez MI, Mubeen M, Zehra M, Samnani I, Abdul Rasool A, Mohan A, Wara UU, Tejwaney U, Kumar V. Role of microRNA in non-alcoholic fatty liver disease (NAFLD) and non-alcoholic steatohepatitis (NASH): a comprehensive review. J Int Med Res 2023; 51:3000605231197058. [PMID: 37676968 PMCID: PMC10492500 DOI: 10.1177/03000605231197058] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/29/2023] [Accepted: 08/08/2023] [Indexed: 09/09/2023] Open
Abstract
Non-alcoholic fatty liver disease (NAFLD) is a prevalent liver condition that affects people who do not overconsume alcohol. Uncertainties exist over how microRNAs (miRNAs) in the blood and liver relate to NAFLD. The aim of this narrative review was to investigate the role of miRNAs in the onset and progression of non-alcoholic steatohepatitis (NASH) from NAFLD, and explore their potential as diagnostic tools and treatment targets for NAFLD patients. Liver miRNA-34a levels were found to accurately represent the degree of liver damage, with lower levels suggesting more damage. In patients with NAFLD and severe liver fibrosis, higher levels of miRNA-193a-5p and miRNA-378d were found. Moreover, miRNA-34a, miRNA-122, and miRNA-192 levels might aid in differentiating NASH from NAFLD. Similar to this, miRNA-21 and miRNA-27 levels in rats were able to distinguish between steatosis and steatohepatitis. High-fat diets enhanced the expression of 15 distinct miRNAs in rats, and there were substantial differences in the miRNA expression patterns between obese and lean people. The results from the present review imply that miRNA microarrays and sequencing may be helpful diagnostic tools, and miRNAs may be a possible treatment target for patients with NAFLD.
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Affiliation(s)
- Mah I Kan Changez
- Department of Medicine, Quetta Institute of Medical Sciences, Quetta, Pakistan
| | - Maryam Mubeen
- Department of Medicine, Punjab Medical College, Faisalabad, Pakistan
| | - Monezahe Zehra
- Department of Medicine, Jinnah Sindh Medical University, Karachi, Pakistan
| | - Inara Samnani
- Department of Medicine, Karachi Medical & Dental College, Karachi, Pakistan
| | | | - Anmol Mohan
- Department of Medicine, Karachi Medical & Dental College, Karachi, Pakistan
| | - Um Ul Wara
- Department of Medicine, Karachi Medical & Dental College, Karachi, Pakistan
| | - Usha Tejwaney
- Department of Pharmacy, Valley Health System, New Jersey, USA
| | - Vikash Kumar
- Department of Internal Medicine, The Brooklyn Hospital Center, New York City, NY, USA
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23
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McIlwraith EK, He W, Belsham DD. Promise and Perils of MicroRNA Discovery Research: Working Toward Quality Over Quantity. Endocrinology 2023; 164:bqad111. [PMID: 37453100 DOI: 10.1210/endocr/bqad111] [Citation(s) in RCA: 6] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 05/25/2023] [Revised: 07/07/2023] [Accepted: 07/11/2023] [Indexed: 07/18/2023]
Abstract
Since the first microRNA (miRNA) was described in 1993 in the humble worm Caenorhabditis elegans, the miRNA field has boomed, with more than 100 000 related patents filed and miRNAs now in ongoing clinical trials. Despite an advanced understanding of the biogenesis and action of miRNAs, applied miRNA research faces challenges and irreproducibility due to a lack of standardization. This review provides guidelines regarding miRNA investigation, while focusing on the pitfalls and considerations that are often overlooked in prevailing applied miRNA research. These include miRNA annotation and quantification, to modulation, target prediction, validation, and the study of circulating miRNAs.
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Affiliation(s)
- Emma K McIlwraith
- Department of Physiology, University of Toronto, Toronto, Ontario, M5S 1A8, Canada
| | - Wenyuan He
- Department of Physiology, University of Toronto, Toronto, Ontario, M5S 1A8, Canada
| | - Denise D Belsham
- Department of Physiology, University of Toronto, Toronto, Ontario, M5S 1A8, Canada
- Department of Medicine, University of Toronto, Toronto, Ontario, M5S 1A8, Canada
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24
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Chen X, Sokirniy I, Wang X, Jiang M, Mseis-Jackson N, Williams C, Mayes K, Jiang N, Puls B, Du Q, Shi Y, Li H. MicroRNA-375 is induced during astrocyte-to-neuron reprogramming and promotes survival of reprogrammed neurons when overexpressed. BIORXIV : THE PREPRINT SERVER FOR BIOLOGY 2023:2023.07.10.548401. [PMID: 37503054 PMCID: PMC10369893 DOI: 10.1101/2023.07.10.548401] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 07/29/2023]
Abstract
While astrocyte-to-neuron (AtN) reprogramming holds great promise in regenerative medicine, the molecular mechanisms that govern this unique biological process remain elusive. MicroRNAs (miRNAs), as post-transcriptional regulators of gene expression, play crucial roles during development and under various pathological conditions. To understand the function of miRNAs during AtN reprogramming process, we performed RNA-seq of both mRNAs and miRNAs on human astrocyte (HA) cultures upon NeuroD1 overexpression. Bioinformatics analyses showed that NeuroD1 not only activates essential neuronal genes to initiate reprogramming process but also induces miRNA changes in HA. Among the upregulated miRNAs, we identified miR-375 and its targets, neuronal ELAVL genes ( nELAVLs ), which encode a family of RNA-binding proteins and are also upregulated by NeuroD1. We further showed that manipulating miR-375 level regulates nELAVLs expression during NeuroD1-mediated reprogramming. Interestingly, miR-375/ nELAVLs are also induced by reprogramming factors Neurog2 and ASCL1 in HA suggesting a conserved function to neuronal reprogramming, and by NeuroD1 in the mouse astrocyte culture and spinal cord. Functionally, we showed that miR-375 overexpression improves NeuroD1-mediated reprogramming efficiency by promoting cell survival at early stages in HA even in cultures treated with the chemotherapy drug Cisplatin. Moreover, miR-375 overexpression doesn't appear to compromise maturation of the reprogrammed neurons in long term HA cultures. Lastly, overexpression of miR-375-refractory ELAVL4 induces apoptosis and reverses the cell survival-promoting effect of miR-375 during AtN reprogramming. Together, we demonstrate a neuro-protective role of miR-375 during NeuroD1-mediated AtN reprogramming and suggest a strategy of combinatory overexpression of NeuroD1 and miR-375 for improving neuronal reprogramming efficiency.
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25
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Walgrave H, Penning A, Tosoni G, Snoeck S, Davie K, Davis E, Wolfs L, Sierksma A, Mars M, Bu T, Thrupp N, Zhou L, Moechars D, Mancuso R, Fiers M, Howden AJ, De Strooper B, Salta E. microRNA-132 regulates gene expression programs involved in microglial homeostasis. iScience 2023; 26:106829. [PMID: 37250784 PMCID: PMC10213004 DOI: 10.1016/j.isci.2023.106829] [Citation(s) in RCA: 18] [Impact Index Per Article: 9.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/22/2022] [Revised: 03/13/2023] [Accepted: 05/03/2023] [Indexed: 05/31/2023] Open
Abstract
microRNA-132 (miR-132), a known neuronal regulator, is one of the most robustly downregulated microRNAs (miRNAs) in the brain of Alzheimer's disease (AD) patients. Increasing miR-132 in AD mouse brain ameliorates amyloid and Tau pathologies, and also restores adult hippocampal neurogenesis and memory deficits. However, the functional pleiotropy of miRNAs requires in-depth analysis of the effects of miR-132 supplementation before it can be moved forward for AD therapy. We employ here miR-132 loss- and gain-of-function approaches using single-cell transcriptomics, proteomics, and in silico AGO-CLIP datasets to identify molecular pathways targeted by miR-132 in mouse hippocampus. We find that miR-132 modulation significantly affects the transition of microglia from a disease-associated to a homeostatic cell state. We confirm the regulatory role of miR-132 in shifting microglial cell states using human microglial cultures derived from induced pluripotent stem cells.
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Affiliation(s)
- Hannah Walgrave
- VIB-KU Leuven Center for Brain & Disease Research, 3000 Leuven, Belgium
- KU Leuven, Department of Neurosciences, Leuven Brain Institute (LBI), 3000 Leuven, Belgium
| | - Amber Penning
- Netherlands Institute for Neuroscience, 1105 BA Amsterdam, the Netherlands
| | - Giorgia Tosoni
- Netherlands Institute for Neuroscience, 1105 BA Amsterdam, the Netherlands
| | - Sarah Snoeck
- Netherlands Institute for Neuroscience, 1105 BA Amsterdam, the Netherlands
| | - Kristofer Davie
- VIB-KU Leuven Center for Brain & Disease Research, 3000 Leuven, Belgium
- VIB-KU Leuven Center for Brain & Disease Research, Bioinformatics Core Facility, 3000 Leuven, Belgium
| | - Emma Davis
- UK Dementia Research Institute at UCL, London WC1E 6BT, UK
| | - Leen Wolfs
- VIB-KU Leuven Center for Brain & Disease Research, 3000 Leuven, Belgium
- KU Leuven, Department of Neurosciences, Leuven Brain Institute (LBI), 3000 Leuven, Belgium
| | - Annerieke Sierksma
- VIB-KU Leuven Center for Brain & Disease Research, 3000 Leuven, Belgium
- KU Leuven, Department of Neurosciences, Leuven Brain Institute (LBI), 3000 Leuven, Belgium
| | - Mayte Mars
- Netherlands Institute for Neuroscience, 1105 BA Amsterdam, the Netherlands
| | - Taofeng Bu
- VIB-KU Leuven Center for Brain & Disease Research, 3000 Leuven, Belgium
- KU Leuven, Department of Neurosciences, Leuven Brain Institute (LBI), 3000 Leuven, Belgium
| | - Nicola Thrupp
- VIB-KU Leuven Center for Brain & Disease Research, 3000 Leuven, Belgium
- KU Leuven, Department of Neurosciences, Leuven Brain Institute (LBI), 3000 Leuven, Belgium
| | - Lujia Zhou
- Discovery Neuroscience, Janssen Research and Development, Division of Janssen Pharmaceutica NV, 2340 Beerse, Belgium
| | - Diederik Moechars
- Discovery Neuroscience, Janssen Research and Development, Division of Janssen Pharmaceutica NV, 2340 Beerse, Belgium
| | - Renzo Mancuso
- Microglia and Inflammation in Neurological Disorders (MIND) Lab, VIB Center for Molecular Neurology, VIB, 2610 Antwerp, Belgium
- Department of Biomedical Sciences, University of Antwerp, 2610 Antwerp, Belgium
| | - Mark Fiers
- VIB-KU Leuven Center for Brain & Disease Research, 3000 Leuven, Belgium
- KU Leuven, Department of Neurosciences, Leuven Brain Institute (LBI), 3000 Leuven, Belgium
| | - Andrew J.M. Howden
- UK Dementia Research Institute, University of Dundee, Dundee DD1 4HN, UK
| | - Bart De Strooper
- VIB-KU Leuven Center for Brain & Disease Research, 3000 Leuven, Belgium
- KU Leuven, Department of Neurosciences, Leuven Brain Institute (LBI), 3000 Leuven, Belgium
- UK Dementia Research Institute at UCL, London WC1E 6BT, UK
| | - Evgenia Salta
- Netherlands Institute for Neuroscience, 1105 BA Amsterdam, the Netherlands
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26
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Chutipongtanate S, Cetinkaya H, Zhang X, Kuhnell D, Benefield D, Haffey W, Wyder M, Patel R, Conrey SC, Burrell AR, Langevin S, Nommsen-Rivers L, Newburg DS, Greis KD, Staat MA, Morrow AL. Prenatal SARS-CoV-2 infection alters postpartum human milk-derived extracellular vesicles. BIORXIV : THE PREPRINT SERVER FOR BIOLOGY 2023:2023.06.01.543234. [PMID: 37398231 PMCID: PMC10312504 DOI: 10.1101/2023.06.01.543234] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 07/04/2023]
Abstract
Human milk-derived extracellular vesicles (HMEVs) are crucial functional components in breast milk, contributing to infant health and development. Maternal conditions could affect HMEV cargos; however, the impact of SARS-CoV-2 infection on HMEVs remains unknown. This study evaluated the influence of SARS-CoV-2 infection during pregnancy on postpartum HMEV molecules. Milk samples (9 prenatal SARS-CoV-2 vs. 9 controls) were retrieved from the IMPRINT birth cohort. After defatting and casein micelle disaggregation, 1 mL milk was subjected to a sequential process of centrifugation, ultrafiltration, and qEV-size exclusion chromatography. Particle and protein characterizations were performed following the MISEV2018 guidelines. EV lysates were analyzed through proteomics and miRNA sequencing, while the intact EVs were biotinylated for surfaceomic analysis. Multi-Omics was employed to predict HMEV functions associated with prenatal SARS-CoV-2 infection. Demographic data between the prenatal SARS-CoV-2 and control groups were similar. The median duration from maternal SARS-CoV-2 test positivity to milk collection was 3 months (range: 1-6 months). Transmission electron microscopy showed the cup-shaped nanoparticles. Nanoparticle tracking analysis demonstrated particle diameters of <200 nm and yields of >1e11 particles from 1 mL milk. Western immunoblots detected ALIX, CD9 and HSP70, supporting the presence of HMEVs in the isolates. Thousands of HMEV cargos and hundreds of surface proteins were identified and compared. Multi-Omics predicted that mothers with prenatal SARS-CoV-2 infection produced HMEVs with enhanced functionalities involving metabolic reprogramming and mucosal tissue development, while mitigating inflammation and lower EV transmigration potential. Our findings suggest that SARS-CoV-2 infection during pregnancy boosts mucosal site-specific functions of HMEVs, potentially protecting infants against viral infections. Further prospective studies should be pursued to reevaluate the short- and long-term benefits of breastfeeding in the post-COVID era.
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27
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Yamasaki T, Tokutsu R, Sawa H, Razali NN, Hayashi M, Minagawa J. Small RNA-mediated silencing of phototropin suppresses the induction of photoprotection in the green alga Chlamydomonas reinhardtii. Proc Natl Acad Sci U S A 2023; 120:e2302185120. [PMID: 37098057 PMCID: PMC10160981 DOI: 10.1073/pnas.2302185120] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/13/2023] [Accepted: 03/30/2023] [Indexed: 04/26/2023] Open
Abstract
Small RNAs (sRNAs) form complexes with Argonaute proteins and bind to transcripts with complementary sequences to repress gene expression. sRNA-mediated regulation is conserved in a diverse range of eukaryotes and is involved in the control of various physiological functions. sRNAs are present in the unicellular green alga Chlamydomonas reinhardtii, and genetic analyses revealed that the core sRNA biogenesis and action mechanisms are conserved with those of multicellular organisms. However, the roles of sRNAs in this organism remain largely unknown. Here, we report that Chlamydomonas sRNAs contribute to the induction of photoprotection. In this alga, photoprotection is mediated by LIGHT HARVESTING COMPLEX STRESS-RELATED 3 (LHCSR3), whose expression is induced by light signals through the blue-light receptor phototropin (PHOT). We demonstrate here that sRNA-defective mutants showed increased PHOT abundance leading to greater LHCSR3 expression. Disruption of the precursor for two sRNAs predicted to bind to the PHOT transcript also increased PHOT accumulation and LHCSR3 expression. The induction of LHCSR3 in the mutants was enhanced by light containing blue wavelengths, but not by red light, indicating that the sRNAs regulate the degree of photoprotection via regulation of PHOT expression. Our results suggest that sRNAs are involved not only in the regulation of photoprotection but also in biological phenomena regulated by PHOT signaling.
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Affiliation(s)
- Tomohito Yamasaki
- Science and Technology Department, Natural Science Cluster, Kochi University, Kochi780-8520, Japan
| | - Ryutaro Tokutsu
- Division of Environmental Photobiology, National Institute for Basic Biology, Myodaiji, Okazaki444-8585, Japan
| | - Haruhi Sawa
- Department of Chemistry and Biotechnology, Faculty of Science and Technology, Kochi University, Kochi780-8520, Japan
| | - Nazifa Naziha Razali
- Department of Chemistry and Biotechnology, Faculty of Science and Technology, Kochi University, Kochi780-8520, Japan
| | - Momoka Hayashi
- Department of Chemistry and Biotechnology, Faculty of Science and Technology, Kochi University, Kochi780-8520, Japan
| | - Jun Minagawa
- Division of Environmental Photobiology, National Institute for Basic Biology, Myodaiji, Okazaki444-8585, Japan
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28
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Sun G, Qi M, Kim AS, Lizhar EM, Sun OW, Al-Abdullah IH, Riggs AD. Reassessing the Abundance of miRNAs in the Human Pancreas and Rodent Cell Lines and Its Implication. Noncoding RNA 2023; 9:ncrna9020020. [PMID: 36960965 PMCID: PMC10037588 DOI: 10.3390/ncrna9020020] [Citation(s) in RCA: 3] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/17/2023] [Revised: 02/24/2023] [Accepted: 03/13/2023] [Indexed: 03/22/2023] Open
Abstract
miRNAs are critical for pancreas development and function. However, we found that there are discrepancies regarding pancreatic miRNA abundance in published datasets. To obtain a more relevant profile that is closer to the true profile, we profiled small RNAs from human islets cells, acini, and four rodent pancreatic cell lines routinely used in diabetes and pancreatic research using a bias reduction protocol for small RNA sequencing. In contrast to the previous notion that miR-375-3p is the most abundant pancreatic miRNA, we found that miR-148a-3p and miR-7-5p were also abundant in islets. In silico studies using predicted and validated targets of these three miRNAs revealed that they may work cooperatively in endocrine and exocrine cells. Our results also suggest, compared to the most-studied miR-375, that both miR-148a-3p and miR-7-5p may play more critical roles in the human pancreas. Moreover, according to in silico-predicted targets, we found that miR-375-3p had a much broader target spectrum by targeting the coding sequence and the 5' untranslated region, rather than the conventional 3' untranslated region, suggesting additional unexplored roles of miR-375-3p beyond the pancreas. Our study provides a valuable new resource for studying miRNAs in pancreata.
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Affiliation(s)
- Guihua Sun
- Department of Diabetes Complications & Metabolism, Arthur Riggs Diabetes & Metabolism Research Institute, City of Hope, Duarte, CA 91010, USA
- Department of Neurodegenerative Diseases, Beckman Research Institute, City of Hope, Duarte, CA 91010, USA
| | - Meirigeng Qi
- Department of Translational Research & Cellular Therapeutics, Arthur Riggs Diabetes & Metabolism Research Institute, City of Hope, Duarte, CA 91010, USA
| | - Alexis S Kim
- Department of Diabetes Complications & Metabolism, Arthur Riggs Diabetes & Metabolism Research Institute, City of Hope, Duarte, CA 91010, USA
| | - Elizabeth M Lizhar
- Department of Diabetes Complications & Metabolism, Arthur Riggs Diabetes & Metabolism Research Institute, City of Hope, Duarte, CA 91010, USA
| | - Olivia W Sun
- Department of Diabetes & Cancer Metabolism, Arthur Riggs Diabetes & Metabolism Research Institute, City of Hope, Duarte, CA 91010, USA
| | - Ismail H Al-Abdullah
- Department of Translational Research & Cellular Therapeutics, Arthur Riggs Diabetes & Metabolism Research Institute, City of Hope, Duarte, CA 91010, USA
| | - Arthur D Riggs
- Department of Diabetes Complications & Metabolism, Arthur Riggs Diabetes & Metabolism Research Institute, City of Hope, Duarte, CA 91010, USA
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29
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Cheng J, Lin G, Wang T, Wang Y, Guo W, Liao J, Yang P, Chen J, Shao X, Lu X, Zhu L, Wang Y, Fan X. Massively Parallel CRISPR-Based Genetic Perturbation Screening at Single-Cell Resolution. ADVANCED SCIENCE (WEINHEIM, BADEN-WURTTEMBERG, GERMANY) 2023; 10:e2204484. [PMID: 36504444 PMCID: PMC9896079 DOI: 10.1002/advs.202204484] [Citation(s) in RCA: 7] [Impact Index Per Article: 3.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Figures] [Subscribe] [Scholar Register] [Received: 08/05/2022] [Revised: 11/09/2022] [Indexed: 06/17/2023]
Abstract
The clustered regularly interspaced short palindromic repeats (CRISPR)-based genetic screening has been demonstrated as a powerful approach for unbiased functional genomics research. Single-cell CRISPR screening (scCRISPR) techniques, which result from the combination of single-cell toolkits and CRISPR screening, allow dissecting regulatory networks in complex biological systems at unprecedented resolution. These methods allow cells to be perturbed en masse using a pooled CRISPR library, followed by high-content phenotyping. This is technically accomplished by annotating cells with sgRNA-specific barcodes or directly detectable sgRNAs. According to the integration of distinct single-cell technologies, these methods principally fall into four categories: scCRISPR with RNA-seq, scCRISPR with ATAC-seq, scCRISPR with proteome probing, and imaging-based scCRISPR. scCRISPR has deciphered genotype-phenotype relationships, genetic regulations, tumor biological issues, and neuropathological mechanisms. This review provides insight into the technical breakthrough of scCRISPR by systematically summarizing the advancements of various scCRISPR methodologies and analyzing their merits and limitations. In addition, an application-oriented approach guide is offered to meet researchers' individualized demands.
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Affiliation(s)
- Junyun Cheng
- Pharmaceutical Informatics InstituteCollege of Pharmaceutical SciencesZhejiang UniversityHangzhouZhejiang310058China
| | - Gaole Lin
- Pharmaceutical Informatics InstituteCollege of Pharmaceutical SciencesZhejiang UniversityHangzhouZhejiang310058China
| | - Tianhao Wang
- Pharmaceutical Informatics InstituteCollege of Pharmaceutical SciencesZhejiang UniversityHangzhouZhejiang310058China
| | - Yunzhu Wang
- Pharmaceutical Informatics InstituteCollege of Pharmaceutical SciencesZhejiang UniversityHangzhouZhejiang310058China
| | - Wenbo Guo
- Pharmaceutical Informatics InstituteCollege of Pharmaceutical SciencesZhejiang UniversityHangzhouZhejiang310058China
| | - Jie Liao
- Pharmaceutical Informatics InstituteCollege of Pharmaceutical SciencesZhejiang UniversityHangzhouZhejiang310058China
| | - Penghui Yang
- Pharmaceutical Informatics InstituteCollege of Pharmaceutical SciencesZhejiang UniversityHangzhouZhejiang310058China
| | - Jie Chen
- Pharmaceutical Informatics InstituteCollege of Pharmaceutical SciencesZhejiang UniversityHangzhouZhejiang310058China
| | - Xin Shao
- Pharmaceutical Informatics InstituteCollege of Pharmaceutical SciencesZhejiang UniversityHangzhouZhejiang310058China
| | - Xiaoyan Lu
- Pharmaceutical Informatics InstituteCollege of Pharmaceutical SciencesZhejiang UniversityHangzhouZhejiang310058China
- State Key Laboratory of Component‐Based Chinese MedicineInnovation Center in Zhejiang UniversityHangzhou310058China
- Jinhua Institute of Zhejiang UniversityJinhua321016China
| | - Ling Zhu
- The Save Sight InstituteFaculty of Medicine and Healththe University of SydneySydneyNSW2000Australia
| | - Yi Wang
- Pharmaceutical Informatics InstituteCollege of Pharmaceutical SciencesZhejiang UniversityHangzhouZhejiang310058China
- State Key Laboratory of Component‐Based Chinese MedicineInnovation Center in Zhejiang UniversityHangzhou310058China
- Future Health LaboratoryInnovation Center of Yangtze River DeltaZhejiang UniversityJiaxing314100China
| | - Xiaohui Fan
- Pharmaceutical Informatics InstituteCollege of Pharmaceutical SciencesZhejiang UniversityHangzhouZhejiang310058China
- State Key Laboratory of Component‐Based Chinese MedicineInnovation Center in Zhejiang UniversityHangzhou310058China
- Jinhua Institute of Zhejiang UniversityJinhua321016China
- The Save Sight InstituteFaculty of Medicine and Healththe University of SydneySydneyNSW2000Australia
- Future Health LaboratoryInnovation Center of Yangtze River DeltaZhejiang UniversityJiaxing314100China
- Westlake Laboratory of Life Sciences and BiomedicineHangzhou310024China
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30
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Ono H, Saito H. Sensing intracellular signatures with synthetic mRNAs. RNA Biol 2023; 20:588-602. [PMID: 37582192 PMCID: PMC10431736 DOI: 10.1080/15476286.2023.2244791] [Citation(s) in RCA: 4] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Revised: 06/30/2023] [Accepted: 07/31/2023] [Indexed: 08/17/2023] Open
Abstract
The bottom-up assembly of biological components in synthetic biology has contributed to a better understanding of natural phenomena and the development of new technologies for practical applications. Over the past few decades, basic RNA research has unveiled the regulatory roles of RNAs underlying gene regulatory networks; while advances in RNA biology, in turn, have highlighted the potential of a wide variety of RNA elements as building blocks to construct artificial systems. In particular, synthetic mRNA-based translational regulators, which respond to signals in cells and regulate the production of encoded output proteins, are gaining attention with the recent rise of mRNA therapeutics. In this Review, we discuss recent progress in RNA synthetic biology, mainly focusing on emerging technologies for sensing intracellular protein and RNA molecules and controlling translation.
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Affiliation(s)
- Hiroki Ono
- Department of Life Science Frontiers, Center for iPS Cell Research and Application, Kyoto University, Kyoto, Sakyo-Ku, Japan
| | - Hirohide Saito
- Department of Life Science Frontiers, Center for iPS Cell Research and Application, Kyoto University, Kyoto, Sakyo-Ku, Japan
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31
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Henriet E, Abdallah F, Laurent Y, Guimpied C, Clement E, Simon M, Pichon C, Baril P. Targeting TGF-β1/miR-21 pathway in keratinocytes reveals protective effects of silymarin on imiquimod-induced psoriasis mouse model. JID INNOVATIONS 2022; 3:100175. [PMID: 36968096 PMCID: PMC10034514 DOI: 10.1016/j.xjidi.2022.100175] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/14/2022] [Revised: 11/10/2022] [Accepted: 11/11/2022] [Indexed: 12/23/2022] Open
Abstract
Epidermal cells integrate multiple signals that activate the signaling pathways involved in skin homeostasis. TGF-β1 signaling pathway upregulates microRNA (miR)-21-5p in keratinocytes and is often deregulated in skin diseases. To identify the bioactive compounds that enable to modulate the TGF-β1/miR-21-5p signaling pathway, we screened a library of medicinal plant extracts using our miR-ON RILES luciferase reporter system placed under the control of the miR-21-5p in keratinocytes treated with TGF-β1. We identified silymarin, a mixture of flavonolignans extracted from Silybum marianum (L.) Gaertn., as the most potent regulator of miR-21-5p expression. Using Argonaute 2 immunoprecipitation and RT-qPCR, we showed that silymarin regulates the expression of miR-21-5p through a noncanonical TGF-β1 signaling pathway, whereas RNA-sequencing analysis revealed three unexpected transcriptomic signatures associated with keratinocyte differentiation, cell cycle, and lipid metabolism. Mechanistically, we demonstrated that SM blocks cell cycle progression, inhibits keratinocyte differentiation through repression of Notch3 expression, stimulates lipid synthesis via activation of PPARγ signaling and inhibits inflammatory responses by suppressing the transcriptional activity of NF-κB. We finally showed that topical application of silymarin alleviates the development of imiquimod-induced psoriasiform lesions in mice by abrogating the altered expression levels of markers involved in inflammation, proliferation, differentiation, and lipid metabolism.
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32
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Dou CX, Ying ZM, Tang LJ, Wang F, Jiang JH. Genetically Encoded Light-Up RNA Amplifier Dissecting MicroRNA Activity in Live Cells. Anal Chem 2022; 94:15481-15488. [DOI: 10.1021/acs.analchem.2c03643] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/28/2022]
Affiliation(s)
- Cai-Xia Dou
- State Key Laboratory of Chemo/Bio-Sensing and Chemometrics, College of Chemistry and Chemical Engineering, Hunan University, Changsha, Hunan 410082, China
| | - Zhan-Ming Ying
- State Key Laboratory of Chemo/Bio-Sensing and Chemometrics, College of Chemistry and Chemical Engineering, Hunan University, Changsha, Hunan 410082, China
| | - Li-Juan Tang
- State Key Laboratory of Chemo/Bio-Sensing and Chemometrics, College of Chemistry and Chemical Engineering, Hunan University, Changsha, Hunan 410082, China
| | - Fenglin Wang
- State Key Laboratory of Chemo/Bio-Sensing and Chemometrics, College of Chemistry and Chemical Engineering, Hunan University, Changsha, Hunan 410082, China
| | - Jian-Hui Jiang
- State Key Laboratory of Chemo/Bio-Sensing and Chemometrics, College of Chemistry and Chemical Engineering, Hunan University, Changsha, Hunan 410082, China
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33
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Dhar R, Mukherjee S, Mukerjee N, Mukherjee D, Devi A, Ashraf GM, Alserihi RF, Tayeb HH, Hashem AM, Alexiou A, Thorate N. Interrelation between extracellular vesicles miRNAs with chronic lung diseases. J Cell Physiol 2022; 237:4021-4036. [PMID: 36063496 DOI: 10.1002/jcp.30867] [Citation(s) in RCA: 14] [Impact Index Per Article: 4.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/29/2022] [Revised: 08/18/2022] [Accepted: 08/22/2022] [Indexed: 11/07/2022]
Abstract
Extracellular vehicles (EVs) are nanoscale lipid bilayer vesicles that carry biologically active biomolecule cargos like proteins, lipids, and nucleic acids (DNA, RNA) outside of the cell. Blood (serum/plasma), urine, and bronchoalveolar lavage fluid are all examples of biofluids from which they may be collected. EVs play a vital role in intracellular communication. The molecular signature of EVs largely depends on the parental cell's status. EVs are classified into two groups, (1) exosomes (originated by endogenous route) and (2) microvesicles (originated from the plasma membrane, also known as ectosomes). The quantity and types of EV cargo vary during normal conditions compared to pathological conditions (chronic inflammatory lung diseases or lung cancer). Consequently, EVs contain novel biomarkers that differ based on the cell type of origin and during lung diseases. Small RNAs (e.g., microRNAs) are transported by EVs, which is one of the most rapidly evolving research areas in the field of EVs biology. EV-mediated cargos transport small RNAs that can result in reprograming the target/recipient cells. Multiple chronic inflammatory lung illnesses, such as chronic obstructive pulmonary disease, asthma, pulmonary hypertension, pulmonary fibrosis, cystic fibrosis, acute lung injury, and lung cancer, have been demonstrated to be regulated by EV. In this review, we will consolidate the current knowledge and literature on the novel role of EVs and their small RNAs concerning chronic lung diseases (CLDs). Additionally, we will also provide better insight into the clinical and translational impact of mesenchymal stem cells-derived EVs as novel therapeutic agents in treating CLDs.
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Affiliation(s)
- Rajib Dhar
- Cancer and Stem Cell Biology Laboratory, Department of Genetic Engineering, SRM Institute of Science and Technology, Kattankulathur, Tamil Nadu, India
| | - Sayantanee Mukherjee
- Centre for Nanosciences and Molecular Medicine, Amrita Vishwa Vidyapeetham University, Kochi, Kerala, India
| | - Nobendu Mukerjee
- Department of Microbiology, Ramakrishna Mission Vivekananda Centenary College, Kolkata, West Bengal, India.,Department of Health Sciences, Novel Global Community Educational Foundation, Hebersham, Australia
| | | | - Arikketh Devi
- Cancer and Stem Cell Biology Laboratory, Department of Genetic Engineering, SRM Institute of Science and Technology, Kattankulathur, Tamil Nadu, India
| | - Ghulam Md Ashraf
- Pre-Clinical Research Unit, King Fahd Medical Research Center, King Abdulaziz University, Jeddah, Saudi Arabia.,Department of Medical Laboratory Sciences, Faculty of Applied Medical Sciences, King Abdulaziz University, Jeddah, Saudi Arabia
| | - Raed F Alserihi
- Department of Medical Laboratory Sciences, Faculty of Applied Medical Sciences, King Abdulaziz University, Jeddah, Saudi Arabia.,Hematology Research Unit, King Fahd Medical Research Center, King Abdulaziz University, Jeddah, Saudi Arabia.,Nanomedicine Unit, Center of Innovation in Personalized Medicine, King Abdulaziz University, Jeddah, Saudi Arabia
| | - Hossam H Tayeb
- Department of Medical Laboratory Sciences, Faculty of Applied Medical Sciences, King Abdulaziz University, Jeddah, Saudi Arabia.,Nanomedicine Unit, Center of Innovation in Personalized Medicine, King Abdulaziz University, Jeddah, Saudi Arabia.,King Fahd Medical Research Center, King Abdulaziz University, Jeddah, Saudi Arabia
| | - Anwar M Hashem
- Department of Medical Laboratory Sciences, Faculty of Applied Medical Sciences, King Abdulaziz University, Jeddah, Saudi Arabia.,Vaccines and Immunotherapy Unit, King Fahad Medical Research Center, Jeddah, Saudi Arabia
| | - Athanasios Alexiou
- Department of Science and Engineering, Novel Global Community Educational Foundation, Hebersham, Australia
| | - Nanasaheb Thorate
- Division of Medical Sciences, Nuffield Department of Women's & Reproductive Health, John Radcliffe Hospital, University of Oxford, Oxford, United Kingdom
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34
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Fiorillo C, Yen PS, Colantoni A, Mariconti M, Azevedo N, Lombardo F, Failloux AB, Arcà B. MicroRNAs and other small RNAs in Aedes aegypti saliva and salivary glands following chikungunya virus infection. Sci Rep 2022; 12:9536. [PMID: 35681077 PMCID: PMC9184468 DOI: 10.1038/s41598-022-13780-3] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/18/2022] [Accepted: 05/27/2022] [Indexed: 11/10/2022] Open
Abstract
Mosquito saliva facilitates blood feeding through the anti-haemostatic, anti-inflammatory and immunomodulatory properties of its proteins. However, the potential contribution of non-coding RNAs to host manipulation is still poorly understood. We analysed small RNAs from Aedes aegypti saliva and salivary glands and show here that chikungunya virus-infection triggers both the siRNA and piRNA antiviral pathways with limited effects on miRNA expression profiles. Saliva appears enriched in specific miRNA subsets and its miRNA content is well conserved among mosquitoes and ticks, clearly pointing to a non-random sorting and occurrence. Finally, we provide evidence that miRNAs from Ae. aegypti saliva may target human immune and inflammatory pathways, as indicated by prediction analysis and searching for experimentally validated targets of identical human miRNAs. Overall, we believe these observations convincingly support a scenario where both proteins and miRNAs from mosquito saliva are injected into vertebrates during blood feeding and contribute to the complex vector-host-pathogen interactions.
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Affiliation(s)
- Carmine Fiorillo
- Department of Public Health and Infectious Diseases - Division of Parasitology, "Sapienza" University, Piazzale Aldo Moro 5, 00185, Rome, Italy
| | - Pei-Shi Yen
- Arboviruses and Insect Vectors Unit, Institute Pasteur, 25 rue Dr. Roux, 75724, Paris Cedex 15, France
| | - Alessio Colantoni
- Department of Biology and Biotechnology, "Sapienza" University, Piazzale Aldo Moro 5, 00185, Rome, Italy
| | - Marina Mariconti
- Arboviruses and Insect Vectors Unit, Institute Pasteur, 25 rue Dr. Roux, 75724, Paris Cedex 15, France
| | - Nayara Azevedo
- Genomics Core Facility, European Molecular Biology Laboratory, Meyerhofstrasse 1, 69117, Heidelberg, Germany
| | - Fabrizio Lombardo
- Department of Public Health and Infectious Diseases - Division of Parasitology, "Sapienza" University, Piazzale Aldo Moro 5, 00185, Rome, Italy
| | - Anna-Bella Failloux
- Arboviruses and Insect Vectors Unit, Institute Pasteur, 25 rue Dr. Roux, 75724, Paris Cedex 15, France
| | - Bruno Arcà
- Department of Public Health and Infectious Diseases - Division of Parasitology, "Sapienza" University, Piazzale Aldo Moro 5, 00185, Rome, Italy.
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35
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Exosome Carrier Effects; Resistance to Digestion in Phagolysosomes May Assist Transfers to Targeted Cells; II Transfers of miRNAs Are Better Analyzed via Systems Approach as They Do Not Fit Conventional Reductionist Stoichiometric Concepts. Int J Mol Sci 2022; 23:ijms23116192. [PMID: 35682875 PMCID: PMC9181154 DOI: 10.3390/ijms23116192] [Citation(s) in RCA: 3] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/21/2022] [Revised: 04/18/2022] [Accepted: 04/26/2022] [Indexed: 01/27/2023] Open
Abstract
Carrier effects of extracellular vesicles (EV) like exosomes refer to properties of the vesicles that contribute to the transferred biologic effects of their contents to targeted cells. This can pertain to ingested small amounts of xenogeneic plant miRNAs and oral administration of immunosuppressive exosomes. The exosomes contribute carrier effects on transfers of miRNAs by contributing both to the delivery and the subsequent functional intracellular outcomes. This is in contrast to current quantitative canonical rules that dictate just the minimum copies of a miRNA for functional effects, and thus successful transfers, independent of the EV carrier effects. Thus, we argue here that transfers by non-canonical minute quantities of miRNAs must consider the EV carrier effects of functional low levels of exosome transferred miRNA that may not fit conventional reductionist stoichiometric concepts. Accordingly, we have examined traditional stoichiometry vs. systems biology that may be more appropriate for delivered exosome functional responses. Exosome carrier properties discussed include; their required surface activating interactions with targeted cells, potential alternate targets beyond mRNAs, like reaching a threshold, three dimensional aspects of the RNAs, added EV kinetic dynamic aspects making transfers four dimensional, and unique intracellular release from EV that resist intracellular digestion in phagolysosomes. Together these EV carrier considerations might allow systems analysis. This can then result in a more appropriate understanding of transferred exosome carrier-assisted functional transfers. A plea is made that the miRNA expert community, in collaboration with exosome experts, perform new experiments on molecular and quantitative miRNA functional effects in systems that include EVs, like variation in EV type and surface constituents, delivery, dose and time to hopefully create more appropriate and truly current canonical concepts of the consequent miRNA functional transfers by EVs like exosomes.
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36
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Abstract
MicroRNAs (miRNAs) belong to a class of endogenous small noncoding RNAs that regulate gene expression at the posttranscriptional level, through both translational repression and mRNA destabilization. They are key regulators of kidney morphogenesis, modulating diverse biological processes in different renal cell lineages. Dysregulation of miRNA expression disrupts early kidney development and has been implicated in the pathogenesis of developmental kidney diseases. In this Review, we summarize current knowledge of miRNA biogenesis and function and discuss in detail the role of miRNAs in kidney morphogenesis and developmental kidney diseases, including congenital anomalies of the kidney and urinary tract and Wilms tumor. We conclude by discussing the utility of miRNAs as potentially novel biomarkers and therapeutic agents.
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Affiliation(s)
- Débora Malta Cerqueira
- Division of Nephrology, Department of Pediatrics, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania, USA
- John G. Rangos Sr. Research Center, UPMC Children’s Hospital of Pittsburgh, Pittsburgh, Pennsylvania, USA
| | - Maliha Tayeb
- Division of Nephrology, Department of Pediatrics, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania, USA
- John G. Rangos Sr. Research Center, UPMC Children’s Hospital of Pittsburgh, Pittsburgh, Pennsylvania, USA
| | - Jacqueline Ho
- Division of Nephrology, Department of Pediatrics, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania, USA
- John G. Rangos Sr. Research Center, UPMC Children’s Hospital of Pittsburgh, Pittsburgh, Pennsylvania, USA
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37
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Mockly S, Houbron É, Seitz H. A rationalized definition of general tumor suppressor microRNAs excludes miR-34a. Nucleic Acids Res 2022; 50:4703-4712. [PMID: 35474387 PMCID: PMC9071449 DOI: 10.1093/nar/gkac277] [Citation(s) in RCA: 7] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/02/2021] [Revised: 03/18/2022] [Accepted: 04/08/2022] [Indexed: 12/24/2022] Open
Abstract
While several microRNAs (miRNAs) have been proposed to act as tumor suppressors, a consensual definition of tumor suppressing miRNAs is still missing. Similarly to coding genes, we propose that tumor suppressor miRNAs must show evidence of genetic or epigenetic inactivation in cancers, and exhibit an anti-tumorigenic (e.g., anti-proliferative) activity under endogenous expression levels. Here we observe that this definition excludes the most extensively studied tumor suppressor candidate miRNA, miR-34a. In analyzable cancer types, miR-34a does not appear to be down-regulated in primary tumors relatively to normal adjacent tissues. Deletion of miR-34a is occasionally found in human cancers, but it does not seem to be driven by an anti-tumorigenic activity of the miRNA, since it is not observed upon smaller, miR-34a-specific alterations. Its anti-proliferative action was observed upon large, supra-physiological transfection of synthetic miR-34a in cultured cells, and our data indicates that endogenous miR-34a levels do not have such an effect. Our results therefore argue against a general tumor suppressive function for miR-34a, providing an explanation to the lack of efficiency of synthetic miR-34a administration against solid tumors.
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Affiliation(s)
- Sophie Mockly
- Institut de Génétique Humaine, UMR 9002 CNRS and university of Montpellier, Montpellier, France
| | - Élisabeth Houbron
- Institut de Génétique Humaine, UMR 9002 CNRS and university of Montpellier, Montpellier, France
| | - Hervé Seitz
- Institut de Génétique Humaine, UMR 9002 CNRS and university of Montpellier, Montpellier, France
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38
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Moubarak RS, Koetz-Ploch L, Mullokandov G, Gaziel A, de Pablos-Aragoneses A, Argibay D, Kleffman K, Sokolova E, Berwick M, Thomas NE, Osman I, Brown BD, Hernando E. In Vivo miRNA Decoy Screen Reveals miR-124a as a Suppressor of Melanoma Metastasis. Front Oncol 2022; 12:852952. [PMID: 35480113 PMCID: PMC9036958 DOI: 10.3389/fonc.2022.852952] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/11/2022] [Accepted: 02/24/2022] [Indexed: 11/30/2022] Open
Abstract
Melanoma is a highly prevalent cancer with an increasing incidence worldwide and high metastatic potential. Brain metastasis is a major complication of the disease, as more than 50% of metastatic melanoma patients eventually develop intracranial disease. MicroRNAs (miRNAs) have been found to play an important role in the tumorigenicity of different cancers and have potential as markers of disease outcome. Identification of relevant miRNAs has generally stemmed from miRNA profiling studies of cells or tissues, but these approaches may have missed miRNAs with relevant functions that are expressed in subfractions of cancer cells. We performed an unbiased in vivo screen to identify miRNAs with potential functions as metastasis suppressors using a lentiviral library of miRNA decoys. Notably, we found that a significant fraction of melanomas that metastasized to the brain carried a decoy for miR-124a, a miRNA that is highly expressed in the brain/neurons. Additional loss- and gain-of-function in vivo validation studies confirmed miR-124a as a suppressor of melanoma metastasis and particularly of brain metastasis. miR-124a overexpression did not inhibit tumor growth in vivo, underscoring that miR-124a specifically controls processes required for melanoma metastatic growth, such as seeding and growth post-extravasation. Finally, we provide proof of principle of this miRNA as a promising therapeutic agent by showing its ability to impair metastatic growth of melanoma cells seeded in distal organs. Our efforts shed light on miR-124a as an antimetastatic agent, which could be leveraged therapeutically to impair metastatic growth and improve patient survival.
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Affiliation(s)
- Rana S. Moubarak
- Department of Pathology, New York University (NYU) School of Medicine, New York, NY, United States
- Interdisciplinary Melanoma Cooperative Group (IMCG), New York University (NYU) Cancer Institute, New York, NY, United States
- Laura and Isaac Perlmutter Cancer Center, New York University (NYU) Langone Health, New York, NY, United States
| | - Lisa Koetz-Ploch
- Department of Pathology, New York University (NYU) School of Medicine, New York, NY, United States
- Interdisciplinary Melanoma Cooperative Group (IMCG), New York University (NYU) Cancer Institute, New York, NY, United States
| | - Gavriel Mullokandov
- Department of Genetics and Genomic Sciences, Icahn School of Medicine at Mount Sinai, New York, NY, United States
| | - Avital Gaziel
- Department of Pathology, New York University (NYU) School of Medicine, New York, NY, United States
- Interdisciplinary Melanoma Cooperative Group (IMCG), New York University (NYU) Cancer Institute, New York, NY, United States
| | - Ana de Pablos-Aragoneses
- Department of Pathology, New York University (NYU) School of Medicine, New York, NY, United States
- Interdisciplinary Melanoma Cooperative Group (IMCG), New York University (NYU) Cancer Institute, New York, NY, United States
| | - Diana Argibay
- Department of Pathology, New York University (NYU) School of Medicine, New York, NY, United States
- Interdisciplinary Melanoma Cooperative Group (IMCG), New York University (NYU) Cancer Institute, New York, NY, United States
| | - Kevin Kleffman
- Department of Pathology, New York University (NYU) School of Medicine, New York, NY, United States
- Interdisciplinary Melanoma Cooperative Group (IMCG), New York University (NYU) Cancer Institute, New York, NY, United States
| | - Elena Sokolova
- Department of Pathology, New York University (NYU) School of Medicine, New York, NY, United States
- Interdisciplinary Melanoma Cooperative Group (IMCG), New York University (NYU) Cancer Institute, New York, NY, United States
| | - Marianne Berwick
- Division of Epidemiology, Biostatistics and Preventive Medicine, Department of Internal Medicine, University of New Mexico, Albuquerque, NM, United States
| | - Nancy E. Thomas
- Department of Dermatology, University of North Carolina, Chapel Hill, NC, United States
| | - Iman Osman
- Interdisciplinary Melanoma Cooperative Group (IMCG), New York University (NYU) Cancer Institute, New York, NY, United States
- Laura and Isaac Perlmutter Cancer Center, New York University (NYU) Langone Health, New York, NY, United States
- Ronald O. Perelman Department of Dermatology, New York University (NYU) School of Medicine, New York, NY, United States
| | - Brian D. Brown
- Department of Genetics and Genomic Sciences, Icahn School of Medicine at Mount Sinai, New York, NY, United States
| | - Eva Hernando
- Department of Pathology, New York University (NYU) School of Medicine, New York, NY, United States
- Interdisciplinary Melanoma Cooperative Group (IMCG), New York University (NYU) Cancer Institute, New York, NY, United States
- Laura and Isaac Perlmutter Cancer Center, New York University (NYU) Langone Health, New York, NY, United States
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Dhainaut M, Rose SA, Akturk G, Wroblewska A, Nielsen SR, Park ES, Buckup M, Roudko V, Pia L, Sweeney R, Le Berichel J, Wilk CM, Bektesevic A, Lee BH, Bhardwaj N, Rahman AH, Baccarini A, Gnjatic S, Pe'er D, Merad M, Brown BD. Spatial CRISPR genomics identifies regulators of the tumor microenvironment. Cell 2022; 185:1223-1239.e20. [PMID: 35290801 PMCID: PMC8992964 DOI: 10.1016/j.cell.2022.02.015] [Citation(s) in RCA: 141] [Impact Index Per Article: 47.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/05/2021] [Revised: 12/02/2021] [Accepted: 02/12/2022] [Indexed: 12/15/2022]
Abstract
While CRISPR screens are helping uncover genes regulating many cell-intrinsic processes, existing approaches are suboptimal for identifying extracellular gene functions, particularly in the tissue context. Here, we developed an approach for spatial functional genomics called Perturb-map. We applied Perturb-map to knock out dozens of genes in parallel in a mouse model of lung cancer and simultaneously assessed how each knockout influenced tumor growth, histopathology, and immune composition. Moreover, we paired Perturb-map and spatial transcriptomics for unbiased analysis of CRISPR-edited tumors. We found that in Tgfbr2 knockout tumors, the tumor microenvironment (TME) was converted to a fibro-mucinous state, and T cells excluded, concomitant with upregulated TGFβ and TGFβ-mediated fibroblast activation, indicating that TGFβ-receptor loss on cancer cells increased TGFβ bioavailability and its immunosuppressive effects on the TME. These studies establish Perturb-map for functional genomics within the tissue at single-cell resolution with spatial architecture preserved and provide insight into how TGFβ responsiveness of cancer cells can affect the TME.
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Affiliation(s)
- Maxime Dhainaut
- Precision Immunology Institute, Icahn School of Medicine at Mount Sinai, New York, NY, USA; Icahn Genomics Institute, Icahn School of Medicine at Mount Sinai, New York, NY, USA; Tisch Cancer Institute, Icahn School of Medicine at Mount Sinai, New York, NY, USA; Department of Genetics and Genomic Sciences, Icahn School of Medicine at Mount Sinai, New York, NY, USA
| | - Samuel A Rose
- Icahn Genomics Institute, Icahn School of Medicine at Mount Sinai, New York, NY, USA; Computational and Systems Biology Program, Memorial Sloan Kettering Cancer Center, New York, NY, USA
| | - Guray Akturk
- Precision Immunology Institute, Icahn School of Medicine at Mount Sinai, New York, NY, USA; Tisch Cancer Institute, Icahn School of Medicine at Mount Sinai, New York, NY, USA
| | - Aleksandra Wroblewska
- Precision Immunology Institute, Icahn School of Medicine at Mount Sinai, New York, NY, USA; Icahn Genomics Institute, Icahn School of Medicine at Mount Sinai, New York, NY, USA
| | - Sebastian R Nielsen
- Precision Immunology Institute, Icahn School of Medicine at Mount Sinai, New York, NY, USA; Icahn Genomics Institute, Icahn School of Medicine at Mount Sinai, New York, NY, USA
| | - Eun Sook Park
- Icahn Genomics Institute, Icahn School of Medicine at Mount Sinai, New York, NY, USA; Department of Genetics and Genomic Sciences, Icahn School of Medicine at Mount Sinai, New York, NY, USA
| | - Mark Buckup
- Precision Immunology Institute, Icahn School of Medicine at Mount Sinai, New York, NY, USA
| | - Vladimir Roudko
- Precision Immunology Institute, Icahn School of Medicine at Mount Sinai, New York, NY, USA; Tisch Cancer Institute, Icahn School of Medicine at Mount Sinai, New York, NY, USA
| | - Luisanna Pia
- Precision Immunology Institute, Icahn School of Medicine at Mount Sinai, New York, NY, USA; Icahn Genomics Institute, Icahn School of Medicine at Mount Sinai, New York, NY, USA
| | - Robert Sweeney
- Precision Immunology Institute, Icahn School of Medicine at Mount Sinai, New York, NY, USA
| | - Jessica Le Berichel
- Precision Immunology Institute, Icahn School of Medicine at Mount Sinai, New York, NY, USA
| | - C Matthias Wilk
- Precision Immunology Institute, Icahn School of Medicine at Mount Sinai, New York, NY, USA
| | - Anela Bektesevic
- Precision Immunology Institute, Icahn School of Medicine at Mount Sinai, New York, NY, USA; Icahn Genomics Institute, Icahn School of Medicine at Mount Sinai, New York, NY, USA
| | - Brian H Lee
- Precision Immunology Institute, Icahn School of Medicine at Mount Sinai, New York, NY, USA
| | - Nina Bhardwaj
- Precision Immunology Institute, Icahn School of Medicine at Mount Sinai, New York, NY, USA; Tisch Cancer Institute, Icahn School of Medicine at Mount Sinai, New York, NY, USA
| | - Adeeb H Rahman
- Precision Immunology Institute, Icahn School of Medicine at Mount Sinai, New York, NY, USA; Tisch Cancer Institute, Icahn School of Medicine at Mount Sinai, New York, NY, USA
| | - Alessia Baccarini
- Precision Immunology Institute, Icahn School of Medicine at Mount Sinai, New York, NY, USA; Icahn Genomics Institute, Icahn School of Medicine at Mount Sinai, New York, NY, USA
| | - Sacha Gnjatic
- Precision Immunology Institute, Icahn School of Medicine at Mount Sinai, New York, NY, USA; Tisch Cancer Institute, Icahn School of Medicine at Mount Sinai, New York, NY, USA
| | - Dana Pe'er
- Computational and Systems Biology Program, Memorial Sloan Kettering Cancer Center, New York, NY, USA
| | - Miriam Merad
- Precision Immunology Institute, Icahn School of Medicine at Mount Sinai, New York, NY, USA; Tisch Cancer Institute, Icahn School of Medicine at Mount Sinai, New York, NY, USA; Center of Excellence for Thoracic Oncology, Icahn School of Medicine at Mount Sinai, New York, NY, USA
| | - Brian D Brown
- Precision Immunology Institute, Icahn School of Medicine at Mount Sinai, New York, NY, USA; Icahn Genomics Institute, Icahn School of Medicine at Mount Sinai, New York, NY, USA; Tisch Cancer Institute, Icahn School of Medicine at Mount Sinai, New York, NY, USA; Department of Genetics and Genomic Sciences, Icahn School of Medicine at Mount Sinai, New York, NY, USA.
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40
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Simeone I, Rubolino C, Noviello T, Farinello D, Cerulo L, Marzi M, Nicassio F. Prediction and pan-cancer analysis of mammalian transcripts involved in target directed miRNA degradation. Nucleic Acids Res 2022; 50:2019-2035. [PMID: 35137158 PMCID: PMC8887481 DOI: 10.1093/nar/gkac057] [Citation(s) in RCA: 21] [Impact Index Per Article: 7.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/12/2021] [Revised: 01/14/2022] [Accepted: 01/25/2022] [Indexed: 12/04/2022] Open
Abstract
It is currently unknown how many RNA transcripts are able to induce degradation of microRNAs (miRNA) via the mechanism known as target-directed miRNA degradation (TDMD). We developed TDMDfinder, a computational pipeline that identifies 'high confidence' TDMD interactions in the Human and Mouse transcriptomes by combining sequence alignment and feature selection approaches. Our predictions suggested that TDMD is widespread, with potentially every miRNA controlled by endogenous targets. We experimentally tested 37 TDMDfinder predictions, of which 17 showed TDMD effects as measured by RT-qPCR and small RNA sequencing, linking the miR-17, miR-19, miR-30, miR-221, miR-26 and miR-23 families to novel endogenous TDMDs. In some cases, TDMD was found to affect different members of the same miRNA family selectively. Features like complementarity to the miRNA 3' region, bulge size and hybridization energy appeared to be the main factors determining sensitivity. Computational analyses performed using the multiomic TCGA platform substantiated the involvement of many TDMD transcripts in human cancer and highlighted 36 highly significant interactions, suggesting TDMD as a new potential oncogenic mechanism. In conclusion, TDMDfinder provides the first inventory of bona fide human and mouse TDMDs. Available as a free webtool, TDMDfinder allows users to search for any TDMD interaction of interest by customizing its selection criteria.
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Affiliation(s)
- Ines Simeone
- Center for Genomic Science of IIT@SEMM, Istituto Italiano di Tecnologia (IIT)—Via Adamello 16, 20139 Milan, Italy
| | - Carmela Rubolino
- Center for Genomic Science of IIT@SEMM, Istituto Italiano di Tecnologia (IIT)—Via Adamello 16, 20139 Milan, Italy
| | - Teresa Maria Rosaria Noviello
- Department of Electrical Engineering and Information Technology, University of Naples “Federico II”, Via Claudio 21, Naples 80128, Italy
- BioGeM, Institute of Genetic Research “Gaetano Salvatore”, Via Camporeale, Ariano Irpino (AV) 83031, Italy
| | - Diego Farinello
- Center for Genomic Science of IIT@SEMM, Istituto Italiano di Tecnologia (IIT)—Via Adamello 16, 20139 Milan, Italy
| | - Luigi Cerulo
- Dep. of Science and Technology, University of Sannio, via de Sanctis, 11, Benevento 82100, Italy
- BioGeM, Institute of Genetic Research “Gaetano Salvatore”, Via Camporeale, Ariano Irpino (AV) 83031, Italy
| | - Matteo Jacopo Marzi
- Center for Genomic Science of IIT@SEMM, Istituto Italiano di Tecnologia (IIT)—Via Adamello 16, 20139 Milan, Italy
| | - Francesco Nicassio
- Center for Genomic Science of IIT@SEMM, Istituto Italiano di Tecnologia (IIT)—Via Adamello 16, 20139 Milan, Italy
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41
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López de Las Hazas MC, Del Pozo-Acebo L, Dávalos A. Response to: Letter to the editor regarding "Dietary bovine milk miRNAs transported in extracellular vesicles are partially stable during GI digestion, are bioavailable and reach target tissues but need a minimum dose to impact on gene expression". Eur J Nutr 2022; 61:1697-1698. [PMID: 35192028 DOI: 10.1007/s00394-022-02816-z] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Accepted: 12/21/2021] [Indexed: 11/04/2022]
Affiliation(s)
- María-Carmen López de Las Hazas
- Laboratory of Epigenetics of Lipid Metabolism, Instituto Madrileño de Estudios Avanzados (IMDEA)-Alimentación, CEI UAM+CSIC, Ctra. De Cantoblanco 8, 28049, Madrid, Spain
| | - Lorena Del Pozo-Acebo
- Laboratory of Epigenetics of Lipid Metabolism, Instituto Madrileño de Estudios Avanzados (IMDEA)-Alimentación, CEI UAM+CSIC, Ctra. De Cantoblanco 8, 28049, Madrid, Spain
| | - Alberto Dávalos
- Laboratory of Epigenetics of Lipid Metabolism, Instituto Madrileño de Estudios Avanzados (IMDEA)-Alimentación, CEI UAM+CSIC, Ctra. De Cantoblanco 8, 28049, Madrid, Spain.
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42
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Høye E, Fromm B, Böttger PHM, Domanska D, Torgunrud A, Lund-Andersen C, Abrahamsen TW, Fretland Å, Dagenborg VJ, Lorenz S, Edwin B, Hovig E, Flatmark K. A comprehensive framework for analysis of microRNA sequencing data in metastatic colorectal cancer. NAR Cancer 2022; 4:zcab051. [PMID: 35047825 PMCID: PMC8759566 DOI: 10.1093/narcan/zcab051] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/18/2021] [Revised: 11/24/2021] [Accepted: 12/15/2021] [Indexed: 11/29/2022] Open
Abstract
Although microRNAs (miRNAs) contribute to all hallmarks of cancer, miRNA dysregulation in metastasis remains poorly understood. The aim of this work was to reliably identify miRNAs associated with metastatic progression of colorectal cancer (CRC) using novel and previously published next-generation sequencing (NGS) datasets generated from 268 samples of primary (pCRC) and metastatic CRC (mCRC; liver, lung and peritoneal metastases) and tumor adjacent tissues. Differential expression analysis was performed using a meticulous bioinformatics pipeline, including only bona fide miRNAs, and utilizing miRNA-tailored quality control and processing. Five miRNAs were identified as up-regulated at multiple metastatic sites Mir-210_3p, Mir-191_5p, Mir-8-P1b_3p [mir-141–3p], Mir-1307_5p and Mir-155_5p. Several have previously been implicated in metastasis through involvement in epithelial-to-mesenchymal transition and hypoxia, while other identified miRNAs represent novel findings. The use of a publicly available pipeline facilitates reproducibility and allows new datasets to be added as they become available. The set of miRNAs identified here provides a reliable starting-point for further research into the role of miRNAs in metastatic progression.
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Affiliation(s)
| | | | | | | | | | | | | | | | | | | | | | | | - Kjersti Flatmark
- To whom correspondence should be addressed. Tel: +47 22 78 18 63;
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Peng J, Liang Q, Xu Z, Cai Y, Peng B, Li J, Zhang W, Kang F, Hong Q, Yan Y, Zhang M. Current Understanding of Exosomal MicroRNAs in Glioma Immune Regulation and Therapeutic Responses. Front Immunol 2022; 12:813747. [PMID: 35095909 PMCID: PMC8796999 DOI: 10.3389/fimmu.2021.813747] [Citation(s) in RCA: 21] [Impact Index Per Article: 7.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/12/2021] [Accepted: 12/27/2021] [Indexed: 02/05/2023] Open
Abstract
Exosomes, the small extracellular vesicles, are released by multiple cell types, including tumor cells, and represent a novel avenue for intercellular communication via transferring diverse biomolecules. Recently, microRNAs (miRNAs) were demonstrated to be enclosed in exosomes and therefore was protected from degradation. Such exosomal miRNAs can be transmitted to recipient cells where they could regulate multiple cancer-associated biological processes. Accumulative evidence suggests that exosomal miRNAs serve essential roles in modifying the glioma immune microenvironment and potentially affecting the malignant behaviors and therapeutic responses. As exosomal miRNAs are detectable in almost all kinds of biofluids and correlated with clinicopathological characteristics of glioma, they might be served as promising biomarkers for gliomas. We reviewed the novel findings regarding the biological functions of exosomal miRNAs during glioma pathogenesis and immune regulation. Furthermore, we elaborated on their potential clinical applications as biomarkers in glioma diagnosis, prognosis and treatment response prediction. Finally, we summarized the accessible databases that can be employed for exosome-associated miRNAs identification and functional exploration of cancers, including glioma.
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Affiliation(s)
- Jinwu Peng
- Department of Pathology, Xiangya Hospital, Central South University, Changsha, China
- Department of Pathology, Xiangya Changde Hospital, Changde, China
| | - Qiuju Liang
- Department of Pharmacy, Xiangya Hospital, Central South University, Changsha, China
| | - Zhijie Xu
- Department of Pathology, Xiangya Hospital, Central South University, Changsha, China
- Department of Pathology, Xiangya Changde Hospital, Changde, China
- National Clinical Research Center for Geriatric Disorders, Xiangya Hospital, Central South University, Changsha, China
| | - Yuan Cai
- Department of Pathology, Xiangya Hospital, Central South University, Changsha, China
| | - Bi Peng
- Department of Pathology, Xiangya Hospital, Central South University, Changsha, China
| | - Jianbo Li
- Department of Pathology, Xiangya Changde Hospital, Changde, China
| | - Wenqin Zhang
- Department of Pathology, Xiangya Changde Hospital, Changde, China
| | - Fanhua Kang
- Department of Pathology, Xiangya Changde Hospital, Changde, China
| | - Qianhui Hong
- Department of Pathology, Xiangya Changde Hospital, Changde, China
| | - Yuanliang Yan
- Department of Pharmacy, Xiangya Hospital, Central South University, Changsha, China
- National Clinical Research Center for Geriatric Disorders, Xiangya Hospital, Central South University, Changsha, China
| | - Mingyu Zhang
- Department of Neurosurgery, Xiangya Hospital, Central South University, Changsha, China
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44
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Westphal MS, Lee E, Schadt EE, Sholler GS, Zhu J. Identification of Let-7 miRNA Activity as a Prognostic Biomarker of SHH Medulloblastoma. Cancers (Basel) 2021; 14:cancers14010139. [PMID: 35008302 PMCID: PMC8750188 DOI: 10.3390/cancers14010139] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/03/2021] [Revised: 12/22/2021] [Accepted: 12/23/2021] [Indexed: 11/16/2022] Open
Abstract
Medulloblastoma (MB) is the most common pediatric embryonal brain tumor. The current consensus classifies MB into four molecular subgroups: sonic hedgehog-activated (SHH), wingless-activated (WNT), Group 3, and Group 4. MYCN and let-7 play a critical role in MB. Thus, we inferred the activity of miRNAs in MB by using the ActMiR procedure. SHH-MB has higher MYCN expression than the other subgroups. We showed that high MYCN expression with high let-7 activity is significantly associated with worse overall survival, and this association was validated in an independent MB dataset. Altogether, our results suggest that let-7 activity and MYCN can further categorize heterogeneous SHH tumors into more and less-favorable prognostic subtypes, which provide critical information for personalizing treatment options for SHH-MB. Comparing the expression differences between the two SHH-MB prognostic subtypes with compound perturbation profiles, we identified FGFR inhibitors as one potential treatment option for SHH-MB patients with the less-favorable prognostic subtype.
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Affiliation(s)
| | - Eunjee Lee
- Sema4, 333 Ludlow St., Stamford, CT 06902, USA; (M.S.W.); (E.L.); (E.E.S.)
- Department of Genetics and Genomic Sciences, Icahn School of Medicine at Mount Sinai, 1 Gustave L. Levy Pl, New York, NY 10029, USA
| | - Eric E. Schadt
- Sema4, 333 Ludlow St., Stamford, CT 06902, USA; (M.S.W.); (E.L.); (E.E.S.)
- Department of Genetics and Genomic Sciences, Icahn School of Medicine at Mount Sinai, 1 Gustave L. Levy Pl, New York, NY 10029, USA
| | - Giselle S. Sholler
- Helen DeVos Children’s Hospital, Grand Rapids, MI 49503, USA;
- College of Human Medicine, Michigan State University, Grand Rapids, MI 49503, USA
| | - Jun Zhu
- Sema4, 333 Ludlow St., Stamford, CT 06902, USA; (M.S.W.); (E.L.); (E.E.S.)
- Department of Genetics and Genomic Sciences, Icahn School of Medicine at Mount Sinai, 1 Gustave L. Levy Pl, New York, NY 10029, USA
- The Tisch Cancer Institute, Icahn School of Medicine at Mount Sinai, 1 Gustave L. Levy Pl, New York, NY 10029, USA
- Correspondence:
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45
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Tomé-Carneiro J, de Las Hazas MCL, Boughanem H, Böttcher Y, Cayir A, Macias González M, Dávalos A. Up-to-date on the evidence linking miRNA-related epitranscriptomic modifications and disease settings. Can these modifications affect cross-kingdom regulation? RNA Biol 2021; 18:586-599. [PMID: 34843412 DOI: 10.1080/15476286.2021.2002003] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 10/19/2022] Open
Abstract
The field of epitranscriptomics is rapidly developing. Several modifications (e.g. methylations) have been identified for different RNA types. Current evidence shows that chemical RNA modifications can influence the whole molecule's secondary structure, translatability, functionality, stability, and degradation, and some are dynamically and reversibly modulated. miRNAs, in particular, are not only post-transcriptional modulators of gene expression but are themselves submitted to regulatory mechanisms. Understanding how these modifications are regulated and the resulting pathological consequences when dysregulation occurs is essential for the development of new therapeutic targets. In humans and other mammals, dietary components have been shown to affect miRNA expression and may also induce chemical modifications in miRNAs. The identification of chemical modifications in miRNAs (endogenous and exogenous) that can impact host gene expression opens up an alternative way to select new specific therapeutic targets.Hence, the aim of this review is to briefly address how RNA epitranscriptomic modifications can affect miRNA biogenesis and to summarize the existing evidence showing the connection between the (de)regulation of these processes and disease settings. In addition, we hypothesize on the potential effect certain chemical modifications could have on the potential cross-kingdom journey of dietary plant miRNAs.
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Affiliation(s)
- João Tomé-Carneiro
- Laboratory of Functional Foods, Madrid Institute for Advanced Studies (IMDEA)-food, CEI UAM + CSIM, Spain
| | | | - Hatim Boughanem
- Instituto de Investigación Biomédica de Málaga (Ibima), Unidad de Gestión Clínica de Endocrinología Y Nutrición Del Hospital Virgen de La Victoria, Málaga, Spain.,Instituto de Salud Carlos Iii (Isciii), Consorcio Ciber, M.p. Fisiopatología de La Obesidad Y Nutrición (Ciberobn), Madrid, Spain.,Vocational Health College, Canakkale Onsekiz Mart University, Canakkale, Turkey
| | - Yvonne Böttcher
- Institute of Clinical Medicine, Department of Clinical Molecular Biology (EpiGen), University of Oslo, Oslo, Norway.,Department of Medical Services and Techniques (EpiGen), Akershus Universitetssykehus, Lørenskog, Norway
| | - Akin Cayir
- Institute of Clinical Medicine, Department of Clinical Molecular Biology (EpiGen), University of Oslo, Oslo, Norway.,Vocational Health College, Canakkale Onsekiz Mart University, Canakkale, Turkey
| | - Manuel Macias González
- Instituto de Investigación Biomédica de Málaga (Ibima), Unidad de Gestión Clínica de Endocrinología Y Nutrición Del Hospital Virgen de La Victoria, Málaga, Spain.,Instituto de Salud Carlos Iii (Isciii), Consorcio Ciber, M.p. Fisiopatología de La Obesidad Y Nutrición (Ciberobn), Madrid, Spain
| | - Alberto Dávalos
- Laboratory of Epigenetics of Lipid Metabolism, Madrid Institute for Advanced Studies (IMDEA)-food, CEI UAM + CSIC, Spain
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46
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Solvin ÅØ, Chawla K, Olsen LC, Hegre SA, Danielsen K, Jenssen M, Furberg AS, Saunes M, Hveem K, Saetrom P, Løset M. MicroRNA profiling of psoriatic skin identifies 11 miRNAs associated with disease severity. Exp Dermatol 2021; 31:535-547. [PMID: 34748247 DOI: 10.1111/exd.14497] [Citation(s) in RCA: 14] [Impact Index Per Article: 3.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/22/2021] [Revised: 11/01/2021] [Accepted: 11/03/2021] [Indexed: 12/18/2022]
Abstract
MicroRNAs (miRNAs) are small non-coding RNAs that have emerged as central regulators of gene expression and powerful biomarkers of disease. Much is yet unknown about their role in psoriasis pathology. To globally characterize the miRNAome of psoriatic skin, skin biopsies were collected from psoriatic cases (n = 75) and non-psoriatic controls (n = 46) and RNA sequenced. Count data were meta-analysed with a previously published dataset (cases, n = 24, controls, n = 20), increasing the number of psoriatic cases fourfold from previously published studies. Differential gene expression analyses were performed comparing lesional psoriatic (PP), non-lesional psoriatic (PN) and control (NN) skin. Further, functional enrichment and cell-specific analyses were performed. Across all contrasts, we identified 439 significantly differentially expressed miRNAs (DEMs), of which 85 were novel for psoriasis and 11 were related to disease severity. Meta-analyses identified 20 DEMs between PN and NN, suggesting an inherent change in the constitution of all skin in psoriasis. By integrating the miRNA transcriptome with mRNA target interactions, we identified several functionally enriched terms, including "thyroid hormone signalling," "insulin resistance" and various infectious diseases. Cell-specific expression analyses revealed that the upregulated DEMs were enriched in epithelial and immune cells. This study provides the most comprehensive overview of the miRNAome in psoriatic skin to date and identifies a miRNA signature related to psoriasis severity. Our results may represent molecular links between psoriasis and related comorbidities and have outlined potential directions for future functional studies to identify biomarkers and treatment targets.
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Affiliation(s)
- Åshild Ø Solvin
- K.G. Jebsen Center for Genetic Epidemiology, Department of Public Health and Nursing, Norwegian University of Science and Technology (NTNU), Trondheim, Norway
| | - Konika Chawla
- K.G. Jebsen Center for Genetic Epidemiology, Department of Public Health and Nursing, Norwegian University of Science and Technology (NTNU), Trondheim, Norway.,Department of Clinical and Molecular Medicine, Norwegian University of Science and Technology (NTNU), Trondheim, Norway.,Bioinformatics Core Facility, Norwegian University of Science and Technology (NTNU), Trondheim, Norway
| | - Lene C Olsen
- K.G. Jebsen Center for Genetic Epidemiology, Department of Public Health and Nursing, Norwegian University of Science and Technology (NTNU), Trondheim, Norway.,Department of Clinical and Molecular Medicine, Norwegian University of Science and Technology (NTNU), Trondheim, Norway.,Bioinformatics Core Facility, Norwegian University of Science and Technology (NTNU), Trondheim, Norway
| | - Siv Anita Hegre
- K.G. Jebsen Center for Genetic Epidemiology, Department of Public Health and Nursing, Norwegian University of Science and Technology (NTNU), Trondheim, Norway.,Department of Clinical and Molecular Medicine, Norwegian University of Science and Technology (NTNU), Trondheim, Norway.,Bioinformatics Core Facility, Norwegian University of Science and Technology (NTNU), Trondheim, Norway
| | - Kjersti Danielsen
- Department of Dermatology, University Hospital of North Norway, Tromsø, Norway
| | - Marita Jenssen
- Department of Dermatology, University Hospital of North Norway, Tromsø, Norway
| | - Anne-Sofie Furberg
- Faculty of Health Sciences and Social Care, Molde University College, Molde, Norway.,Department of Microbiology and Infection Control, University Hospital of North Norway, Tromsø, Norway
| | - Marit Saunes
- Department of Dermatology, Clinic of Orthopaedy, Rheumatology and Dermatology, St. Olavs Hospital Trondheim University Hospital, Trondheim, Norway
| | - Kristian Hveem
- K.G. Jebsen Center for Genetic Epidemiology, Department of Public Health and Nursing, Norwegian University of Science and Technology (NTNU), Trondheim, Norway.,HUNT Research Centre, Department of Public Health and Nursing, Norwegian University of Science and Technology (NTNU), Trondheim, Norway
| | - Pål Saetrom
- K.G. Jebsen Center for Genetic Epidemiology, Department of Public Health and Nursing, Norwegian University of Science and Technology (NTNU), Trondheim, Norway.,Department of Clinical and Molecular Medicine, Norwegian University of Science and Technology (NTNU), Trondheim, Norway.,Bioinformatics Core Facility, Norwegian University of Science and Technology (NTNU), Trondheim, Norway.,Department of Computer Science, Norwegian University of Science and Technology (NTNU), Trondheim, Norway
| | - Mari Løset
- K.G. Jebsen Center for Genetic Epidemiology, Department of Public Health and Nursing, Norwegian University of Science and Technology (NTNU), Trondheim, Norway.,Department of Dermatology, Clinic of Orthopaedy, Rheumatology and Dermatology, St. Olavs Hospital Trondheim University Hospital, Trondheim, Norway
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47
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Askenase PW. Exosomes provide unappreciated carrier effects that assist transfers of their miRNAs to targeted cells; I. They are 'The Elephant in the Room'. RNA Biol 2021; 18:2038-2053. [PMID: 33944671 PMCID: PMC8582996 DOI: 10.1080/15476286.2021.1885189] [Citation(s) in RCA: 6] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/07/2020] [Revised: 01/23/2021] [Accepted: 01/30/2021] [Indexed: 12/19/2022] Open
Abstract
Extracellular vesicles (EV), such as exosomes, are emerging biologic entities that mediate important newly recognized functional effects. Exosomes are intracellular endosome-originating, cell-secreted, small nano-size EV. They can transfer cargo molecules like miRNAs to act intracellularly in targeted acceptor cells, to then mediate epigenetic functional alterations. Exosomes among EV, are universal nanoparticles of life that are present across all species. Some critics mistakenly hold exosomes to concepts and standards of cells, whereas they are subcellular nanospheres that are a million times smaller, have neither nuclei nor mitochondria, are far less complex and currently cannot be studied deeply and elegantly by many and diverse technologies developed for cells over many years. There are important concerns about the seeming impossibility of biologically significant exosome transfers of very small amounts of miRNAs resulting in altered targeted cell functions. These hesitations are based on current canonical concepts developed for non-physiological application of miRNAs alone, or artificial non-quantitative genetic expression. Not considered is that the natural physiologic intercellular transit via exosomes can contribute numerous augmenting carrier effects to functional miRNA transfers. Some of these are particularly stimulated complex extracellular and intracellular physiologic processes activated in the exosome acceptor cells that can crucially influence the intracellular effects of the transferred miRNAs. These can lead to molecular chemical changes altering DNA expression for mediating functional changes of the targeted cells. Such exosome mediated molecular transfers of epigenetic functional alterations, are the most exciting and life-altering property that these nano EV bring to virtually all of biology and medicine. .Abbreviations: Ab, Antibody Ag Antigen; APC, Antigen presenting cells; CS, contact sensitivity; DC, Dendritic cells; DTH, Delayed-type hypersensitivity; EV, extracellular vesicles; EV, Extracellular vesicle; FLC, Free light chains of antibodies; GI, gastrointestinal; IP, Intraperitoneal administration; IV, intravenous administration; OMV, Outer membrane vesicles released by bacteria; PE, Phos-phatidylethanolamine; PO, oral administration.
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Affiliation(s)
- Philip W. Askenase
- Section of Rheumatology, Allergy and Clinical Immunology Department of Internal Medicine, Yale University School of Medicine, New Haven, CT, USA
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48
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López de Las Hazas MC, Del Pozo-Acebo L, Hansen MS, Gil-Zamorano J, Mantilla-Escalante DC, Gómez-Coronado D, Marín F, Garcia-Ruiz A, Rasmussen JT, Dávalos A. Dietary bovine milk miRNAs transported in extracellular vesicles are partially stable during GI digestion, are bioavailable and reach target tissues but need a minimum dose to impact on gene expression. Eur J Nutr 2021; 61:1043-1056. [PMID: 34716465 DOI: 10.1007/s00394-021-02720-y] [Citation(s) in RCA: 50] [Impact Index Per Article: 12.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/17/2021] [Accepted: 10/18/2021] [Indexed: 12/17/2022]
Abstract
PURPOSE Extracellular RNAs are unstable and rapidly degraded unless protected. Bovine-milk extracellular vesicles (EVs) confer protection to dietary miRNAs, although it remains unclear whether this importantly improves their chances of reaching host target cells to exert biological effects. METHODS Caco-2, HT-29, Hep-G2 and FHs-74 cell lines were exposed to natural/labelled milk EVs to evaluate cellular uptake. Five frequently reported human milk miRNAs (miR-146b-5p, miR-148a-3p, miR-30a-5p, miR-26a-5p, and miR-22-3p) were loaded into EVs. The intracellular concentration of each miRNA in cells was determined. In addition, an animal study giving an oral dose of loaded EVs in C57BL6/ mice were performed. Gene expression regulation was assessed by microarray analysis. RESULTS Digestive stability analysis showed high overall degradation of exogenous miRNAs, although EV-protected miRNAs better resisted gastrointestinal digestion compared to free miRNAs (tenfold higher levels). Importantly, orally delivered EV-loaded miRNAs reached host organs, including brain, in mice. However, no biological effect has been identified. CONCLUSION Milk EVs protect miRNAs from degradation and facilitate cellular uptake. miRNA concentration in EVs from bovine milk might be insufficient to produce gene modulation. Nevertheless, sizable amounts of exogenous miRNAs may be loaded into EVs, and orally delivered EV-loaded miRNAs can reach tissues in vivo, increasing the possibility of exerting biological effects. Further investigation is justified as this could have an impact in the field of nutrition and health (i.e., infant formulas elaboration).
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Affiliation(s)
- María-Carmen López de Las Hazas
- Laboratory of Epigenetics of Lipid Metabolism, IMDEA Food Institute, CEI UAM+CSIC, Ctra. De Cantoblanco 8, 28049, Madrid, Spain
| | - Lorena Del Pozo-Acebo
- Laboratory of Epigenetics of Lipid Metabolism, IMDEA Food Institute, CEI UAM+CSIC, Ctra. De Cantoblanco 8, 28049, Madrid, Spain
| | - Maria S Hansen
- Department of Molecular Biology and Genetics, Aarhus University, 8000, Aarhus, Denmark
| | - Judit Gil-Zamorano
- Laboratory of Epigenetics of Lipid Metabolism, IMDEA Food Institute, CEI UAM+CSIC, Ctra. De Cantoblanco 8, 28049, Madrid, Spain
| | - Diana C Mantilla-Escalante
- Laboratory of Epigenetics of Lipid Metabolism, IMDEA Food Institute, CEI UAM+CSIC, Ctra. De Cantoblanco 8, 28049, Madrid, Spain
| | - Diego Gómez-Coronado
- Department of Biochemistry-Research, Hospital Universitario Ramón y Cajal, IRYCIS, 28034, Madrid, Spain
- CIBER de Fisiopatología de la Obesidad y Nutrición (CIBEROBN), Instituto de Salud Carlos III (ISCIII), 28034 Madrid, Spain
| | - Francisco Marín
- Department of Applied Chemistry-Physics, Faculty of Science, University Autónoma of Madrid, 28049, Madrid, Spain
| | - Almudena Garcia-Ruiz
- Laboratory of Epigenetics of Lipid Metabolism, IMDEA Food Institute, CEI UAM+CSIC, Ctra. De Cantoblanco 8, 28049, Madrid, Spain
| | - Jan T Rasmussen
- Department of Molecular Biology and Genetics, Aarhus University, 8000, Aarhus, Denmark
| | - Alberto Dávalos
- Laboratory of Epigenetics of Lipid Metabolism, IMDEA Food Institute, CEI UAM+CSIC, Ctra. De Cantoblanco 8, 28049, Madrid, Spain.
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49
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Zhang J, Li D, Zhang R, Peng R, Li J. Delivery of microRNA-21-sponge and pre-microRNA-122 by MS2 virus-like particles to therapeutically target hepatocellular carcinoma cells. Exp Biol Med (Maywood) 2021; 246:2463-2472. [PMID: 34644206 DOI: 10.1177/15353702211035689] [Citation(s) in RCA: 14] [Impact Index Per Article: 3.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/20/2022] Open
Abstract
MicroRNAs are related to the development of hepatocellular carcinoma and can serve as potential therapeutic targets. Therapeutic strategies increasing tumor-suppressive microRNAs and reducing oncogenic microRNAs have been developed. Herein, the effects of simultaneously altering two microRNAs using MS2 virus-like particles were studied. The sequences of microRNA-21-sponge and pre-microRNA-122 were connected and cloned into a virus-like particle expression vector. Virus-like particles containing microRNA-21-sponge and pre-microRNA-122 sequences were prepared and crosslinked with a cell-specific peptide targeting hepatocellular carcinoma cells. Delivery effects were studied using RT-qPCR and functional assays to investigate the level of target mRNAs, cell toxicity, and the effects of proliferation, invasion, and migration. Virus-like particles delivered miR-21-sponge into cells, with the Ct value reaching 10 at most. The linked pre-miR-122 was processed into mature miR-122. The mRNA targets of miR-21 were derepressed as predicted and upregulated 1.2-2.8-fold, and the expression of proteins was elevated correspondingly. Proliferation, migration, and invasion of HCC cells were inhibited by miR-21-sponge. Simultaneous delivery of miR-21-sponge and miR-122 further decreased proliferation, migration, and invasion by up to 34%, 63%, and 65%, respectively. And the combination promoted the apoptosis of HCC cells. In conclusion, delivering miR-21-sponge and miR-122 using virus-like particles modified by cell-specific peptides is an effective and convenient strategy to correct microRNA dysregulation in hepatocellular carcinoma cells and is a promising therapeutic strategy for hepatocellular carcinoma.
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Affiliation(s)
- Jiawei Zhang
- National Center for Clinical Laboratories, Beijing Hospital, National Center of Gerontology; Institute of Geriatric Medicine, Chinese Academy of Medical Sciences, Beijing 100730, P.R. China.,Graduate School of Peking Union Medical College, Chinese Academy of Medical Sciences, Beijing 100730, P.R. China.,Beijing Engineering Research Center of Laboratory Medicine, Beijing Hospital, Beijing 100730, P.R. China
| | - Dandan Li
- National Center for Clinical Laboratories, Beijing Hospital, National Center of Gerontology; Institute of Geriatric Medicine, Chinese Academy of Medical Sciences, Beijing 100730, P.R. China.,Graduate School of Peking Union Medical College, Chinese Academy of Medical Sciences, Beijing 100730, P.R. China.,Beijing Engineering Research Center of Laboratory Medicine, Beijing Hospital, Beijing 100730, P.R. China
| | - Rui Zhang
- National Center for Clinical Laboratories, Beijing Hospital, National Center of Gerontology; Institute of Geriatric Medicine, Chinese Academy of Medical Sciences, Beijing 100730, P.R. China.,Beijing Engineering Research Center of Laboratory Medicine, Beijing Hospital, Beijing 100730, P.R. China
| | - Rongxue Peng
- National Center for Clinical Laboratories, Beijing Hospital, National Center of Gerontology; Institute of Geriatric Medicine, Chinese Academy of Medical Sciences, Beijing 100730, P.R. China.,Beijing Engineering Research Center of Laboratory Medicine, Beijing Hospital, Beijing 100730, P.R. China
| | - Jinming Li
- National Center for Clinical Laboratories, Beijing Hospital, National Center of Gerontology; Institute of Geriatric Medicine, Chinese Academy of Medical Sciences, Beijing 100730, P.R. China.,Graduate School of Peking Union Medical College, Chinese Academy of Medical Sciences, Beijing 100730, P.R. China.,Beijing Engineering Research Center of Laboratory Medicine, Beijing Hospital, Beijing 100730, P.R. China
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50
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Massart J, Sjögren RJO, Egan B, Garde C, Lindgren M, Gu W, Ferreira DMS, Katayama M, Ruas JL, Barrès R, O'Gorman DJ, Zierath JR, Krook A. Endurance exercise training-responsive miR-19b-3p improves skeletal muscle glucose metabolism. Nat Commun 2021; 12:5948. [PMID: 34642330 PMCID: PMC8511155 DOI: 10.1038/s41467-021-26095-0] [Citation(s) in RCA: 21] [Impact Index Per Article: 5.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/31/2019] [Accepted: 09/09/2021] [Indexed: 12/13/2022] Open
Abstract
Skeletal muscle is a highly adaptable tissue and remodels in response to exercise training. Using short RNA sequencing, we determine the miRNA profile of skeletal muscle from healthy male volunteers before and after a 14-day aerobic exercise training regime. Among the exercise training-responsive miRNAs identified, miR-19b-3p was selected for further validation. Overexpression of miR-19b-3p in human skeletal muscle cells increases insulin signaling, glucose uptake, and maximal oxygen consumption, recapitulating the adaptive response to aerobic exercise training. Overexpression of miR-19b-3p in mouse flexor digitorum brevis muscle enhances contraction-induced glucose uptake, indicating that miR-19b-3p exerts control on exercise training-induced adaptations in skeletal muscle. Potential targets of miR-19b-3p that are reduced after aerobic exercise training include KIF13A, MAPK6, RNF11, and VPS37A. Amongst these, RNF11 silencing potentiates glucose uptake in human skeletal muscle cells. Collectively, we identify miR-19b-3p as an aerobic exercise training-induced miRNA that regulates skeletal muscle glucose metabolism.
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Affiliation(s)
- Julie Massart
- Department of Molecular Medicine and Surgery, Section of Integrative Physiology, Karolinska Institutet, Stockholm, Sweden
| | - Rasmus J O Sjögren
- Department of Molecular Medicine and Surgery, Section of Integrative Physiology, Karolinska Institutet, Stockholm, Sweden
| | - Brendan Egan
- Department of Physiology and Pharmacology, Section of Integrative Physiology, Karolinska Institutet, Stockholm, Sweden
- Department of Health & Human Performance, Dublin City University, Dublin, Ireland
| | - Christian Garde
- Novo Nordisk Foundation Center for Basic Metabolic Research, University of Copenhagen, Copenhagen, Denmark
| | - Magnus Lindgren
- Department of Molecular Medicine and Surgery, Section of Integrative Physiology, Karolinska Institutet, Stockholm, Sweden
| | - Weifeng Gu
- University of Massachusetts Medical School, Worchester, MA, USA
- Department of Cell Biology and Neuroscience, University of California at Riverside, Riverside, CA, USA
| | - Duarte M S Ferreira
- Department of Physiology and Pharmacology, Section of Molecular and Cellular Exercise Physiology, Karolinska Institutet, Stockholm, Sweden
| | - Mutsumi Katayama
- Department of Physiology and Pharmacology, Section of Integrative Physiology, Karolinska Institutet, Stockholm, Sweden
| | - Jorge L Ruas
- Department of Physiology and Pharmacology, Section of Molecular and Cellular Exercise Physiology, Karolinska Institutet, Stockholm, Sweden
| | - Romain Barrès
- Novo Nordisk Foundation Center for Basic Metabolic Research, University of Copenhagen, Copenhagen, Denmark
| | - Donal J O'Gorman
- Department of Health & Human Performance, Dublin City University, Dublin, Ireland
| | - Juleen R Zierath
- Department of Molecular Medicine and Surgery, Section of Integrative Physiology, Karolinska Institutet, Stockholm, Sweden
- Department of Physiology and Pharmacology, Section of Integrative Physiology, Karolinska Institutet, Stockholm, Sweden
- Novo Nordisk Foundation Center for Basic Metabolic Research, University of Copenhagen, Copenhagen, Denmark
| | - Anna Krook
- Department of Physiology and Pharmacology, Section of Integrative Physiology, Karolinska Institutet, Stockholm, Sweden.
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