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Abou Daya F, Mandigo T, Patel D, Math S, Ober L, Maher M, Melkani G, Walker J, Saxena R. Drosophila Modeling Identifies Increased Sleep as a Link Between Insomnia and Cardiovascular Disease. BIORXIV : THE PREPRINT SERVER FOR BIOLOGY 2025:2025.04.07.647668. [PMID: 40291700 PMCID: PMC12026989 DOI: 10.1101/2025.04.07.647668] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 04/30/2025]
Abstract
Insomnia is a common sleep disorder associated with negative long-term health outcomes. Mendelian randomization studies have found that insomnia significantly increases the risk of cardiovascular disease (CVD). To better understand the link between sleep and heart health, we identify genes associated with both insomnia and CVD. We model the disruption of the Drosophila melanogaster orthologs in neurons and cardiac tissue to characterize their cell-autonomous and non-cell-autonomous role in regulating sleep and cardiac physiology. We identify three genes that function in neurons and the heart to cell-autonomously regulate the function of each tissue. We find that the disruption of insomnia- and CVD-associated Drosophila orthologs in the heart most often lead to increased nighttime sleep. Inversely, disruptions in neurons that lead to increased sleep most often result in an elevated heart rate. To confirm the association between increased sleep and cardiac function, we performed a genetic correlation analysis from human data between long sleep-related traits and adverse cardiac outcomes. Significant correlations were found between most long sleep traits and heart failure, coronary artery disease, or myocardial infarction, reinforcing our findings in the fly linking increased or excessive sleep and altered cardiac health.
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Puig-Barbe A, Dettmann S, Nirello VD, Moor H, Azami S, Edgar BA, Varga-Weisz P, Korzelius J, de Navascués J. A bHLH interaction code controls bipotential differentiation and self-renewal in the Drosophila gut. Cell Rep 2025; 44:115398. [PMID: 40089983 DOI: 10.1016/j.celrep.2025.115398] [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: 09/19/2023] [Revised: 02/04/2025] [Accepted: 02/14/2025] [Indexed: 03/18/2025] Open
Abstract
Multipotent adult stem cells balance self-renewal with differentiation into various cell types. How this balance is regulated at the transcriptional level is poorly understood. Here, we show that a network of basic helix-loop-helix (bHLH) transcription factors controls both stemness and bipotential differentiation in the Drosophila adult intestine. We find that homodimers of Daughterless (Da), a homolog of mammalian E proteins, maintain self-renewal of intestinal stem cells (ISCs), antagonizing the enteroendocrine fate promoted by heterodimers of Da and Scute (Sc; homolog of ASCL). The HLH factor Extramacrochaetae (Emc; homologous to Id proteins) promotes absorptive differentiation by titrating Da and Sc. Emc prevents the committed absorptive progenitor from dedifferentiating, underscoring the plasticity of these cells. Switching physical interaction partners in this way enables the active maintenance of stemness while priming stem cells for differentiation along two alternative fates. Such regulatory logic is likely operative in other bipotent stem cell systems.
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Affiliation(s)
- Aleix Puig-Barbe
- School of Biosciences, Cardiff University, The Sir Martin Evans Building, Museum Avenue, Cardiff CF10 3AX, UK
| | - Svenja Dettmann
- DKFZ/ZMBH Alliance, University of Heidelberg, Deutsches Krebsforschungszentrum, Im Neuenheimer Feld 280, 69120 Heidelberg, Germany; AbbVie Germany GmbH & Co. KG, 81 Mainzer Str., 65189 Wiesbaden, Frankfurt, Germany
| | - Vinícius Dias Nirello
- International Laboratory for Microbiome Host Epigenetics, Department of Genetics, Evolution, Microbiology, and Immunology, Institute of Biology, University of Campinas, Campinas, SP 13083-862, Brazil
| | - Helen Moor
- School of Life Sciences, University of Essex, Wivenhoe Park, Colchester CO4 3SQ, UK
| | - Sina Azami
- Max Planck Institute for Biology of Ageing, Joseph-Stelzmann-Straße 9B, 50931 Köln, Germany
| | - Bruce A Edgar
- DKFZ/ZMBH Alliance, University of Heidelberg, Deutsches Krebsforschungszentrum, Im Neuenheimer Feld 280, 69120 Heidelberg, Germany; Huntsman Cancer Institute & Department of Oncological Sciences, University of Utah, Salt Lake City, UT 84112, USA
| | - Patrick Varga-Weisz
- International Laboratory for Microbiome Host Epigenetics, Department of Genetics, Evolution, Microbiology, and Immunology, Institute of Biology, University of Campinas, Campinas, SP 13083-862, Brazil; School of Life Sciences, University of Essex, Wivenhoe Park, Colchester CO4 3SQ, UK
| | - Jerome Korzelius
- DKFZ/ZMBH Alliance, University of Heidelberg, Deutsches Krebsforschungszentrum, Im Neuenheimer Feld 280, 69120 Heidelberg, Germany; Max Planck Institute for Biology of Ageing, Joseph-Stelzmann-Straße 9B, 50931 Köln, Germany; School of Biosciences, University of Kent, Canterbury CT2 7NZ, UK
| | - Joaquín de Navascués
- School of Biosciences, Cardiff University, The Sir Martin Evans Building, Museum Avenue, Cardiff CF10 3AX, UK; School of Life Sciences, University of Essex, Wivenhoe Park, Colchester CO4 3SQ, UK.
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Nandagopal S, Terrio A, Vicente FZ, Megason SG, Jambhekar A, Lahav G. Activation-derepression synergy enables a bHLH network to coordinate a signal-specific fate response. Cell Rep 2024; 43:115077. [PMID: 39671287 PMCID: PMC11774475 DOI: 10.1016/j.celrep.2024.115077] [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: 05/09/2024] [Revised: 09/27/2024] [Accepted: 11/25/2024] [Indexed: 12/15/2024] Open
Abstract
Stem cells integrate multiple environmental signals to activate appropriate fate programs. To ensure coherent responses, alternative fates must be concomitantly inactivated. However, mechanisms that coordinate fates in a signal-specific manner are not fully understood. Here, we investigate the role of a network of basic-helix-loop-helix (bHLH) transcription factors in neural stem cells, which integrate leukemia inhibitory factor (LIF) and bone morphogenetic protein (BMP) signaling to synergistically induce glial fibrillary acidic protein (GFAP), a key astrocyte-fate determinant. Using quantitative RNA-fluorescence in situ hybridization (FISH) and ectopic expression, we find that multiple bHLHs that promote alternative fates also repress GFAP but are all suppressed by BMP and, to a lesser extent, LIF. Mathematical modeling shows that synergy arises from this coordinated derepression of GFAP combined with its activation by LIF signaling. Finally, we determine how coordinated and tunable derepression results from extensive cross-regulation among bHLHs. Activation-derepression synergy could be broadly utilized to couple signaling and fate, particularly across the numerous developmental systems regulated by bHLH factors.
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Affiliation(s)
- Sandy Nandagopal
- Department of Systems Biology, Blavatnik Institute at Harvard Medical School, Boston, MA 02115, USA.
| | - Alexsandra Terrio
- Department of Systems Biology, Blavatnik Institute at Harvard Medical School, Boston, MA 02115, USA
| | - Fernando Z Vicente
- Department of Systems Biology, Blavatnik Institute at Harvard Medical School, Boston, MA 02115, USA
| | - Sean G Megason
- Department of Systems Biology, Blavatnik Institute at Harvard Medical School, Boston, MA 02115, USA
| | - Ashwini Jambhekar
- Department of Systems Biology, Blavatnik Institute at Harvard Medical School, Boston, MA 02115, USA
| | - Galit Lahav
- Department of Systems Biology, Blavatnik Institute at Harvard Medical School, Boston, MA 02115, USA.
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Nair S, Baker NE. Extramacrochaetae regulates Notch signaling in the Drosophila eye through non-apoptotic caspase activity. eLife 2024; 12:RP91988. [PMID: 39564985 PMCID: PMC11578588 DOI: 10.7554/elife.91988] [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] [Indexed: 11/21/2024] Open
Abstract
Many cell fate decisions are determined transcriptionally. Accordingly, some fate specification is prevented by Inhibitor of DNA-binding (Id) proteins that interfere with DNA binding by master regulatory transcription factors. We show that the Drosophila Id protein Extra macrochaetae (Emc) also affects developmental decisions by regulating caspase activity. Emc, which prevents proneural bHLH transcription factors from specifying neural cell fate, also prevents homodimerization of another bHLH protein, Daughterless (Da), and thereby maintains expression of the Death-Associated Inhibitor of Apoptosis (diap1) gene. Accordingly, we found that multiple effects of emc mutations on cell growth and on eye development were all caused by activation of caspases. These effects included acceleration of the morphogenetic furrow, failure of R7 photoreceptor cell specification, and delayed differentiation of non-neuronal cone cells. Within emc mutant clones, Notch signaling was elevated in the morphogenetic furrow, increasing morphogenetic furrow speed. This was associated with caspase-dependent increase in levels of Delta protein, the transmembrane ligand for Notch. Posterior to the morphogenetic furrow, elevated Delta cis-inhibited Notch signaling that was required for R7 specification and cone cell differentiation. Growth inhibition of emc mutant clones in wing imaginal discs also depended on caspases. Thus, emc mutations reveal the importance of restraining caspase activity even in non-apoptotic cells to prevent abnormal development, in the Drosophila eye through effects on Notch signaling.
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Affiliation(s)
- Sudershana Nair
- Department of Genetics, Albert Einstein College of MedicineBronxUnited States
| | - Nicholas E Baker
- Department of Genetics, Albert Einstein College of MedicineBronxUnited States
- Department of Developmental and Molecular Biology, Albert Einstein College of MedicineBronxUnited States
- Department of Ophthalmology and Visual Sciences, Albert Einstein College of MedicineBronxUnited States
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Nair S, Baker NE. Extramacrochaetae regulates Notch signaling in the Drosophila eye through non-apoptotic caspase activity. BIORXIV : THE PREPRINT SERVER FOR BIOLOGY 2024:2023.10.04.560841. [PMID: 39131389 PMCID: PMC11312471 DOI: 10.1101/2023.10.04.560841] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 08/13/2024]
Abstract
Many cell fate decisions are determined transcriptionally. Accordingly, some fate specification is prevented by Inhibitor of DNA binding (Id) proteins that interfere with DNA binding by master regulatory transcription factors. We show that the Drosophila Id protein Extra macrochaetae (Emc) also affects developmental decisions by regulating caspase activity. Emc, which prevents proneural bHLH transcription factors from specifying neural cell fate, also prevents homodimerization of another bHLH protein, Daughterless (Da), and thereby maintains expression of the Death-Associated Inhibitor of Apoptosis (diap1) gene. Accordingly, we found that multiple effects of emc mutations on cell growth and on eye development were all caused by activation of caspases. These effects included acceleration of the morphogenetic furrow, failure of R7 photoreceptor cell specification, and delayed differentiation of non-neuronal cone cells. Within emc mutant clones, Notch signaling was elevated in the morphogenetic furrow, increasing morphogenetic furrow speed. This was associated with caspase-dependent increase in levels of Delta protein, the transmembrane ligand for Notch. Posterior to the morphogenetic furrow, elevated Delta cis-inhibited Notch signaling that was required for R7 specification and cone cell differentiation. Growth inhibition of emc mutant clones in wing imaginal discs also depended on caspases. Thus, emc mutations reveal the importance of restraining caspase activity even in non-apoptotic cells to prevent abnormal development, in the Drosophila eye through effects on Notch signaling.
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Affiliation(s)
- Sudershana Nair
- Department of Genetics, Albert Einstein College of Medicine, 1300 Morris Park Avenue, Bronx, NY 10461
- Present address: Department of and Physiology, NYU School of Medicine, 435 East 30 St, New York, NY
| | - Nicholas E Baker
- Department of Genetics, Albert Einstein College of Medicine, 1300 Morris Park Avenue, Bronx, NY 10461
- Department of Developmental and Molecular Biology, Albert Einstein College of Medicine, 1300 Morris Park Avenue, Bronx, NY 10461
- Department of Ophthalmology and Visual Sciences, Albert Einstein College of Medicine, 1300 Morris Park Avenue, Bronx, NY 10461
- Present address: Department of Microbiology and Molecular Genetics, University of California, Irvine, 2011 Biological Sciences 3, Irvine, CA 92697-2300
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Reddy Onteddu V, Bhattacharya A, Baker NE. The Id protein Extramacrochaetae restrains the E protein Daughterless to regulate Notch, Rap1, and Sevenless within the R7 equivalence group of the Drosophila eye. Biol Open 2024; 13:bio060124. [PMID: 39041866 PMCID: PMC11360143 DOI: 10.1242/bio.060124] [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: 08/16/2023] [Accepted: 07/15/2024] [Indexed: 07/24/2024] Open
Abstract
The Drosophila Id gene extramacrochaetae (emc) is required during Drosophila eye development for proper cell fate specification within the R7 equivalence group. Without emc, R7 cells develop like R1/6 cells, and there are delays and deficits in differentiation of non-neuronal cone cells. Although emc encodes an Inhibitor of DNA-binding (Id) protein that is known to antagonize proneural bHLH protein function, no proneural gene is known for R7 or cone cell fates. These fates are also independent of daughterless (da), which encodes the ubiquitous E protein heterodimer partner of proneural bHLH proteins. We report here that the effects of emc mutations disappear in the absence of da, and are partially mimicked by forced expression of Da dimers, indicating that emc normally restrains da from interfering with R7 and cone cell specification, as occurs in emc mutants. emc, and da, regulate three known contributors to R7 fate, which are Notch signaling, Rap1, and Sevenless. R7 specification is partially restored to emc mutant cells by mutation of RapGap1, confirming that Rap1 activity, in addition to Notch activity, is a critical target of emc. These findings exemplify how mutations of an Id protein gene can affect processes that do not require any bHLH protein, by restraining Da activity within physiological bounds.
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Affiliation(s)
- Venkateswara Reddy Onteddu
- Department of Genetics, Albert Einstein College of Medicine, 1300 Morris Park Avenue, Bronx, NY 10461,USA
| | - Abhishek Bhattacharya
- Department of Genetics, Albert Einstein College of Medicine, 1300 Morris Park Avenue, Bronx, NY 10461,USA
| | - Nicholas E. Baker
- Department of Genetics, Albert Einstein College of Medicine, 1300 Morris Park Avenue, Bronx, NY 10461,USA
- Department of Ophthalmology and Visual Sciences, Albert Einstein College of Medicine, 1300 Morris Park Avenue, Bronx, NY 10461,USA
- Department of Developmental and Molecular Biology, Albert Einstein College of Medicine, 1300 Morris Park Avenue, Bronx, NY 10461,USA
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Klumpe HE, Garcia-Ojalvo J, Elowitz MB, Antebi YE. The computational capabilities of many-to-many protein interaction networks. Cell Syst 2023; 14:430-446. [PMID: 37348461 PMCID: PMC10318606 DOI: 10.1016/j.cels.2023.05.001] [Citation(s) in RCA: 12] [Impact Index Per Article: 6.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/28/2023] [Revised: 04/14/2023] [Accepted: 05/11/2023] [Indexed: 06/24/2023]
Abstract
Many biological circuits comprise sets of protein variants that interact with one another in a many-to-many, or promiscuous, fashion. These architectures can provide powerful computational capabilities that are especially critical in multicellular organisms. Understanding the principles of biochemical computations in these circuits could allow more precise control of cellular behaviors. However, these systems are inherently difficult to analyze, due to their large number of interacting molecular components, partial redundancies, and cell context dependence. Here, we discuss recent experimental and theoretical advances that are beginning to reveal how promiscuous circuits compute, what roles those computations play in natural biological contexts, and how promiscuous architectures can be applied for the design of synthetic multicellular behaviors.
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Affiliation(s)
- Heidi E Klumpe
- Department of Biomedical Engineering, Boston University, Boston, MA 02215, USA; Biological Design Center, Boston University, Boston, MA 02215, USA
| | - Jordi Garcia-Ojalvo
- Department of Medicine and Life Sciences, Pompeu Fabra University, 08003 Barcelona, Spain.
| | - Michael B Elowitz
- Division of Biology and Biological Engineering, California Institute of Technology, Pasadena, CA 91125, USA; Howard Hughes Medical Institute, Chevy Chase, MD 20815, USA.
| | - Yaron E Antebi
- Department of Molecular Genetics, Weizmann Institute of Science 76100, Rehovot, Israel.
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Cecchini K, Biasini A, Yu T, Säflund M, Mou H, Arif A, Eghbali A, Colpan C, Gainetdinov I, de Rooij DG, Weng Z, Zamore PD, Özata DM. The transcription factor TCFL5 responds to A-MYB to elaborate the male meiotic program in mice. Reproduction 2023; 165:183-196. [PMID: 36395073 PMCID: PMC9812935 DOI: 10.1530/rep-22-0355] [Citation(s) in RCA: 10] [Impact Index Per Article: 5.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/03/2022] [Accepted: 11/17/2022] [Indexed: 11/18/2022]
Abstract
In brief The testis-specific transcription factor, TCFL5, expressed in pachytene spermatocytes regulates the meiotic gene expression program in collaboration with the transcription factor A-MYB. Abstract In male mice, the transcription factors STRA8 and MEISON initiate meiosis I. We report that STRA8/MEISON activates the transcription factors A-MYB and TCFL5, which together reprogram gene expression after spermatogonia enter into meiosis. TCFL5 promotes the transcription of genes required for meiosis, mRNA turnover, miR-34/449 production, meiotic exit, and spermiogenesis. This transcriptional architecture is conserved in rhesus macaque, suggesting TCFL5 plays a central role in meiosis and spermiogenesis in placental mammals. Tcfl5em1/em1 mutants are sterile, and spermatogenesis arrests at the mid- or late-pachytene stage of meiosis. Moreover, Tcfl5+/em1 mutants produce fewer motile sperm.
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Affiliation(s)
- Katharine Cecchini
- RNA Therapeutics Institute and Howard Hughes Medical Institute, University of Massachusetts Medical School, 368 Plantation Street, Worcester, MA 01605, USA
| | - Adriano Biasini
- RNA Therapeutics Institute and Howard Hughes Medical Institute, University of Massachusetts Medical School, 368 Plantation Street, Worcester, MA 01605, USA
| | - Tianxiong Yu
- Program in Bioinformatics and Integrative Biology, University of Massachusetts Medical School, Worcester, MA 01605, USA
| | - Martin Säflund
- Department of Molecular Biosciences, The Wenner-Gren Institute, Stockholm University, S-106 91 Stockholm, Sweden
| | - Haiwei Mou
- Cold Spring Harbor Laboratory, Cold Spring Harbor, NY 11724, USA
| | - Amena Arif
- RNA Therapeutics Institute and Howard Hughes Medical Institute, University of Massachusetts Medical School, 368 Plantation Street, Worcester, MA 01605, USA
- Present address: Beam Therapeutics, 238 Main St, Cambridge, MA 02142, USA
| | - Atiyeh Eghbali
- Department of Molecular Biosciences, The Wenner-Gren Institute, Stockholm University, S-106 91 Stockholm, Sweden
| | - Cansu Colpan
- RNA Therapeutics Institute and Howard Hughes Medical Institute, University of Massachusetts Medical School, 368 Plantation Street, Worcester, MA 01605, USA
- Present address: Voyager Therapeutics, 75 Sidney St, Cambridge, MA 02139, USA
| | - Ildar Gainetdinov
- RNA Therapeutics Institute and Howard Hughes Medical Institute, University of Massachusetts Medical School, 368 Plantation Street, Worcester, MA 01605, USA
| | - Dirk G. de Rooij
- Reproductive Biology Group, Division of Developmental Biology, Department of Biology, Faculty of Science, Utrecht University, Utrecht 3584, the Netherlands
| | - Zhiping Weng
- Program in Bioinformatics and Integrative Biology, University of Massachusetts Medical School, Worcester, MA 01605, USA
| | - Phillip D. Zamore
- RNA Therapeutics Institute and Howard Hughes Medical Institute, University of Massachusetts Medical School, 368 Plantation Street, Worcester, MA 01605, USA
| | - Deniz M. Özata
- Department of Molecular Biosciences, The Wenner-Gren Institute, Stockholm University, S-106 91 Stockholm, Sweden
- Lead contact
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Brown RE, Jacobse J, Anant SA, Blunt KM, Chen B, Vega PN, Jones CT, Pilat JM, Revetta F, Gorby AH, Stengel KR, Choksi YA, Palin K, Piazuelo MB, Washington MK, Lau KS, Goettel JA, Hiebert SW, Short SP, Williams CS. MTG16 regulates colonic epithelial differentiation, colitis, and tumorigenesis by repressing E protein transcription factors. JCI Insight 2022; 7:e153045. [PMID: 35503250 PMCID: PMC9220854 DOI: 10.1172/jci.insight.153045] [Citation(s) in RCA: 11] [Impact Index Per Article: 3.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/15/2021] [Accepted: 04/13/2022] [Indexed: 12/04/2022] Open
Abstract
Aberrant epithelial differentiation and regeneration contribute to colon pathologies, including inflammatory bowel disease (IBD) and colitis-associated cancer (CAC). Myeloid translocation gene 16 (MTG16, also known as CBFA2T3) is a transcriptional corepressor expressed in the colonic epithelium. MTG16 deficiency in mice exacerbates colitis and increases tumor burden in CAC, though the underlying mechanisms remain unclear. Here, we identified MTG16 as a central mediator of epithelial differentiation, promoting goblet and restraining enteroendocrine cell development in homeostasis and enabling regeneration following dextran sulfate sodium-induced (DSS-induced) colitis. Transcriptomic analyses implicated increased Ephrussi box-binding transcription factor (E protein) activity in MTG16-deficient colon crypts. Using a mouse model with a point mutation that attenuates MTG16:E protein interactions (Mtg16P209T), we showed that MTG16 exerts control over colonic epithelial differentiation and regeneration by repressing E protein-mediated transcription. Mimicking murine colitis, MTG16 expression was increased in biopsies from patients with active IBD compared with unaffected controls. Finally, uncoupling MTG16:E protein interactions partially phenocopied the enhanced tumorigenicity of Mtg16-/- colon in the azoxymethane/DSS-induced model of CAC, indicating that MTG16 protects from tumorigenesis through additional mechanisms. Collectively, our results demonstrate that MTG16, via its repression of E protein targets, is a key regulator of cell fate decisions during colon homeostasis, colitis, and cancer.
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Affiliation(s)
- Rachel E. Brown
- Program in Cancer Biology and
- Medical Scientist Training Program, Vanderbilt University School of Medicine, Nashville, Tennessee, USA
| | - Justin Jacobse
- Department of Pathology, Microbiology and Immunology, Vanderbilt University, Nashville, Tennessee, USA
- Willem Alexander Children’s Hospital, Leiden University Medical Center, Leiden, Netherlands
- Department of Medicine, Division of Gastroenterology, Hepatology, and Nutrition, Vanderbilt University Medical Center, Nashville, Tennessee, USA
| | | | - Koral M. Blunt
- Department of Medicine, Division of Gastroenterology, Hepatology, and Nutrition, Vanderbilt University Medical Center, Nashville, Tennessee, USA
| | - Bob Chen
- Program in Chemical and Physical Biology, Vanderbilt University School of Medicine, Nashville, Tennessee, USA
- Epithelial Biology Center, Vanderbilt University Medical Center, Nashville, Tennessee, USA
| | - Paige N. Vega
- Epithelial Biology Center, Vanderbilt University Medical Center, Nashville, Tennessee, USA
- Department of Cell and Developmental Biology and
| | - Chase T. Jones
- Department of Medicine, Division of Gastroenterology, Hepatology, and Nutrition, Vanderbilt University Medical Center, Nashville, Tennessee, USA
| | | | - Frank Revetta
- Department of Pathology, Microbiology and Immunology, Vanderbilt University, Nashville, Tennessee, USA
| | - Aidan H. Gorby
- Department of Medicine, Division of Gastroenterology, Hepatology, and Nutrition, Vanderbilt University Medical Center, Nashville, Tennessee, USA
| | - Kristy R. Stengel
- Department of Biochemistry, Vanderbilt University School of Medicine, Nashville, Tennessee, USA
| | - Yash A. Choksi
- Program in Cancer Biology and
- Department of Medicine, Division of Gastroenterology, Hepatology, and Nutrition, Vanderbilt University Medical Center, Nashville, Tennessee, USA
- Veterans Affairs Tennessee Valley Health Care System, Nashville, Tennessee, USA
- Center for Mucosal Inflammation and Cancer, Vanderbilt University Medical Center, Nashville, Tennessee, USA
| | - Kimmo Palin
- Department of Medical and Clinical Genetics
- Applied Tumor Genomics Research Program, Research Programs Unit, and
- iCAN Digital Precision Cancer Medicine Flagship, University of Helsinki, Helsinki, Finland
| | - M. Blanca Piazuelo
- Department of Medicine, Division of Gastroenterology, Hepatology, and Nutrition, Vanderbilt University Medical Center, Nashville, Tennessee, USA
| | - Mary Kay Washington
- Department of Pathology, Microbiology and Immunology, Vanderbilt University, Nashville, Tennessee, USA
| | - Ken S. Lau
- Epithelial Biology Center, Vanderbilt University Medical Center, Nashville, Tennessee, USA
- Department of Cell and Developmental Biology and
- Vanderbilt-Ingram Cancer Center, Vanderbilt University Medical Center, Nashville, Tennessee, USA
| | - Jeremy A. Goettel
- Program in Cancer Biology and
- Department of Pathology, Microbiology and Immunology, Vanderbilt University, Nashville, Tennessee, USA
- Department of Medicine, Division of Gastroenterology, Hepatology, and Nutrition, Vanderbilt University Medical Center, Nashville, Tennessee, USA
- Center for Mucosal Inflammation and Cancer, Vanderbilt University Medical Center, Nashville, Tennessee, USA
| | - Scott W. Hiebert
- Department of Biochemistry, Vanderbilt University School of Medicine, Nashville, Tennessee, USA
- Vanderbilt-Ingram Cancer Center, Vanderbilt University Medical Center, Nashville, Tennessee, USA
| | - Sarah P. Short
- Program in Cancer Biology and
- Department of Medicine, Division of Gastroenterology, Hepatology, and Nutrition, Vanderbilt University Medical Center, Nashville, Tennessee, USA
- Center for Mucosal Inflammation and Cancer, Vanderbilt University Medical Center, Nashville, Tennessee, USA
| | - Christopher S. Williams
- Program in Cancer Biology and
- Medical Scientist Training Program, Vanderbilt University School of Medicine, Nashville, Tennessee, USA
- Department of Medicine, Division of Gastroenterology, Hepatology, and Nutrition, Vanderbilt University Medical Center, Nashville, Tennessee, USA
- Veterans Affairs Tennessee Valley Health Care System, Nashville, Tennessee, USA
- Center for Mucosal Inflammation and Cancer, Vanderbilt University Medical Center, Nashville, Tennessee, USA
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10
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Zhu R, Del Rio-Salgado JM, Garcia-Ojalvo J, Elowitz MB. Synthetic multistability in mammalian cells. Science 2022; 375:eabg9765. [PMID: 35050677 DOI: 10.1126/science.abg9765] [Citation(s) in RCA: 61] [Impact Index Per Article: 20.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/21/2022]
Abstract
In multicellular organisms, gene regulatory circuits generate thousands of molecularly distinct, mitotically heritable states through the property of multistability. Designing synthetic multistable circuits would provide insight into natural cell fate control circuit architectures and would allow engineering of multicellular programs that require interactions among distinct cell types. We created MultiFate, a naturally inspired, synthetic circuit that supports long-term, controllable, and expandable multistability in mammalian cells. MultiFate uses engineered zinc finger transcription factors that transcriptionally self-activate as homodimers and mutually inhibit one another through heterodimerization. Using a model-based design, we engineered MultiFate circuits that generate as many as seven states, each stable for at least 18 days. MultiFate permits controlled state switching and modulation of state stability through external inputs and can be expanded with additional transcription factors. These results provide a foundation for engineering multicellular behaviors in mammalian cells.
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Affiliation(s)
- Ronghui Zhu
- Division of Biology and Biological Engineering, California Institute of Technology, Pasadena, CA 91125, USA
| | - Jesus M Del Rio-Salgado
- Division of Biology and Biological Engineering, California Institute of Technology, Pasadena, CA 91125, USA
| | - Jordi Garcia-Ojalvo
- Department of Experimental and Health Sciences, Universitat Pompeu Fabra, 08003 Barcelona, Spain
| | - Michael B Elowitz
- Division of Biology and Biological Engineering, California Institute of Technology, Pasadena, CA 91125, USA.,Howard Hughes Medical Institute, California Institute of Technology, Pasadena, CA 91125, USA
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11
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Xu J, Wang JJ, Xue HW, Zhang GH. Leaf direction: Lamina joint development and environmental responses. PLANT, CELL & ENVIRONMENT 2021; 44:2441-2454. [PMID: 33866581 DOI: 10.1111/pce.14065] [Citation(s) in RCA: 16] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 01/19/2021] [Accepted: 04/07/2021] [Indexed: 06/12/2023]
Abstract
Plant architecture plays a major role in canopy photosynthesis and biomass production, and plants adjust their growth (and thus architecture) in response to changing environments. Leaf angle is one of the most important traits in rice (Oryza sativa L.) plant architecture, because leaf angle strongly affects leaf direction and rice production, with more-erect leaves being advantageous for high-density plantings. The degree of leaf bending depends on the morphology of the lamina joint, which connects the leaf and the sheath. In this review, we discuss cell morphology in different lamina joint tissues and describe the underlying genetic network that governs this morphology and thus regulates leaf direction. Furthermore, we focus on the mechanism by how environmental factors influence rice leaf angle. Our review provides a theoretical framework for the future genetic improvement of rice leaf orientation and plant architecture.
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Affiliation(s)
- Jing Xu
- State Key Laboratory of Rice Biology, China National Rice Research Institute, Hangzhou, China
| | - Jia-Jia Wang
- State Key Laboratory of Rice Biology, China National Rice Research Institute, Hangzhou, China
| | - Hong-Wei Xue
- Joint Center for Single Cell Biology, School of Agriculture and Biology, Shanghai Jiao Tong University, Shanghai, China
| | - Guang-Heng Zhang
- State Key Laboratory of Rice Biology, China National Rice Research Institute, Hangzhou, China
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12
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Johnson RI. Hexagonal patterning of the Drosophila eye. Dev Biol 2021; 478:173-182. [PMID: 34245727 DOI: 10.1016/j.ydbio.2021.07.004] [Citation(s) in RCA: 22] [Impact Index Per Article: 5.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/07/2021] [Revised: 07/04/2021] [Accepted: 07/06/2021] [Indexed: 10/24/2022]
Abstract
A complex network of transcription factor interactions propagates across the larval eye disc to establish columns of evenly-spaced R8 precursor cells, the founding cells of Drosophila ommatidia. After the recruitment of additional photoreceptors to each ommatidium, the surrounding cells are organized into their stereotypical pattern during pupal development. These support cells - comprised of pigment and cone cells - are patterned to encapsulate the photoreceptors and separate ommatidia with an hexagonal honeycomb lattice. Since the proteins and processes essential for correct eye patterning are conserved, elucidating how these function and change during Drosophila eye patterning can substantially advance our understanding of transcription factor and signaling networks, cytoskeletal structures, adhesion complexes, and the biophysical properties of complex tissues during their morphogenesis. Our understanding of many of these aspects of Drosophila eye patterning is largely descriptive. Many important questions, especially relating to the regulation and integration of cellular events, remain.
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Affiliation(s)
- Ruth I Johnson
- Biology Department, Wesleyan University, 52 Lawn Avenue, Middletown, CT, USA.
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13
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Yoshikawa G, Miyazaki K, Ogata H, Miyazaki M. The Evolution of Rag Gene Enhancers and Transcription Factor E and Id Proteins in the Adaptive Immune System. Int J Mol Sci 2021; 22:ijms22115888. [PMID: 34072618 PMCID: PMC8199221 DOI: 10.3390/ijms22115888] [Citation(s) in RCA: 8] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/01/2021] [Revised: 05/23/2021] [Accepted: 05/24/2021] [Indexed: 11/17/2022] Open
Abstract
Adaptive immunity relies on the V(D)J DNA recombination of immunoglobulin (Ig) and T cell receptor (TCR) genes, which enables the recognition of highly diverse antigens and the elicitation of antigen-specific immune responses. This process is mediated by recombination-activating gene (Rag) 1 and Rag2 (Rag1/2), whose expression is strictly controlled in a cell type-specific manner; the expression of Rag1/2 genes represents a hallmark of lymphoid lineage commitment. Although Rag genes are known to be evolutionally conserved among jawed vertebrates, how Rag genes are regulated by lineage-specific transcription factors (TFs) and how their regulatory system evolved among vertebrates have not been fully elucidated. Here, we reviewed the current body of knowledge concerning the cis-regulatory elements (CREs) of Rag genes and the evolution of the basic helix-loop-helix TF E protein regulating Rag gene CREs, as well as the evolution of the antagonist of this protein, the Id protein. This may help to understand how the adaptive immune system develops along with the evolution of responsible TFs and enhancers.
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Affiliation(s)
- Genki Yoshikawa
- Bioinformatics Center, Institute for Chemical Research, Kyoto University, Uji 611-0011, Japan;
| | - Kazuko Miyazaki
- Laboratory of Immunology, Institute for Frontier Life and Medical Sciences, Kyoto University, Kyoto 606-8507, Japan;
| | - Hiroyuki Ogata
- Bioinformatics Center, Institute for Chemical Research, Kyoto University, Uji 611-0011, Japan;
- Correspondence: (H.O.); (M.M.)
| | - Masaki Miyazaki
- Laboratory of Immunology, Institute for Frontier Life and Medical Sciences, Kyoto University, Kyoto 606-8507, Japan;
- Correspondence: (H.O.); (M.M.)
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14
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Wojnarowicz PM, Escolano MG, Huang YH, Desai B, Chin Y, Shah R, Xu S, Yadav S, Yaklichkin S, Ouerfelli O, Soni RK, Philip J, Montrose DC, Healey JH, Rajasekhar VK, Garland WA, Ratiu J, Zhuang Y, Norton L, Rosen N, Hendrickson RC, Zhou XK, Iavarone A, Massague J, Dannenberg AJ, Lasorella A, Benezra R. Anti-tumor effects of an ID antagonist with no observed acquired resistance. NPJ Breast Cancer 2021; 7:58. [PMID: 34031428 PMCID: PMC8144414 DOI: 10.1038/s41523-021-00266-0] [Citation(s) in RCA: 12] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/03/2020] [Accepted: 04/15/2021] [Indexed: 12/19/2022] Open
Abstract
ID proteins are helix-loop-helix (HLH) transcriptional regulators frequently overexpressed in cancer. ID proteins inhibit basic-HLH transcription factors often blocking differentiation and sustaining proliferation. A small-molecule, AGX51, targets ID proteins for degradation and impairs ocular neovascularization in mouse models. Here we show that AGX51 treatment of cancer cell lines impairs cell growth and viability that results from an increase in reactive oxygen species (ROS) production upon ID degradation. In mouse models, AGX51 treatment suppresses breast cancer colonization in the lung, regresses the growth of paclitaxel-resistant breast tumors when combined with paclitaxel and reduces tumor burden in sporadic colorectal neoplasia. Furthermore, in cells and mice, we fail to observe acquired resistance to AGX51 likely the result of the inability to mutate the binding pocket without loss of ID function and efficient degradation of the ID proteins. Thus, AGX51 is a first-in-class compound that antagonizes ID proteins, shows strong anti-tumor effects and may be further developed for the management of multiple cancers.
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Affiliation(s)
- Paulina M Wojnarowicz
- Cancer Biology and Genetics Program, Memorial Sloan Kettering Cancer Center, New York, NY, USA
| | - Marta Garcia Escolano
- Cancer Biology and Genetics Program, Memorial Sloan Kettering Cancer Center, New York, NY, USA
| | - Yun-Han Huang
- Cancer Biology and Genetics Program, Memorial Sloan Kettering Cancer Center, New York, NY, USA
- Weill Cornell/Sloan Kettering/Rockefeller Tri-Institutional MD-PhD Program, New York, NY, 10065, USA
- Gerstner Sloan Kettering Graduate School of Biomedical Sciences, New York, NY, 10065, USA
| | - Bina Desai
- Cancer Biology and Genetics Program, Memorial Sloan Kettering Cancer Center, New York, NY, USA
| | - Yvette Chin
- Cancer Biology and Genetics Program, Memorial Sloan Kettering Cancer Center, New York, NY, USA
| | - Riddhi Shah
- Cancer Biology and Genetics Program, Memorial Sloan Kettering Cancer Center, New York, NY, USA
| | - Sijia Xu
- Cancer Biology and Genetics Program, Memorial Sloan Kettering Cancer Center, New York, NY, USA
| | - Saurabh Yadav
- Cancer Biology and Genetics Program, Memorial Sloan Kettering Cancer Center, New York, NY, USA
| | - Sergey Yaklichkin
- Cancer Biology and Genetics Program, Memorial Sloan Kettering Cancer Center, New York, NY, USA
| | - Ouathek Ouerfelli
- Organic Synthesis Core Facility, Memorial Sloan Kettering Cancer Center, New York, NY, USA
| | - Rajesh Kumar Soni
- Proteomics & Microchemistry Core Facility, Memorial Sloan Kettering Cancer Center, New York, NY, USA
| | - John Philip
- Proteomics & Microchemistry Core Facility, Memorial Sloan Kettering Cancer Center, New York, NY, USA
| | - David C Montrose
- Department of Medicine, Weill Cornell Medical College, New York, NY, USA
- Department of Pathology, Renaissance School of Medicine, Stony Brook University, Stony Brook, NY, USA
| | - John H Healey
- Orthopedics Service, Memorial Sloan Kettering Cancer Center, New York, NY, USA
| | | | | | - Jeremy Ratiu
- Department of Immunology, Duke University, Durham, NC, USA
| | - Yuan Zhuang
- Department of Immunology, Duke University, Durham, NC, USA
| | - Larry Norton
- Evelyn H. Lauder Breast Center, Memorial Sloan Kettering Cancer Center, New York, NY, USA
| | - Neal Rosen
- Molecular Pharmacology Program, Memorial Sloan Kettering Cancer Center, New York, NY, USA
| | - Ronald C Hendrickson
- Proteomics & Microchemistry Core Facility, Memorial Sloan Kettering Cancer Center, New York, NY, USA
| | - Xi Kathy Zhou
- Department of Healthcare Policy and Research Weill Cornell Medical College, New York, NY, USA
| | - Antonio Iavarone
- Department of Neurology, Department of Pathology, Institute for Cancer Genetics, Columbia University Medical Center, New York, NY, USA
| | - Joan Massague
- Cancer Biology and Genetics Program, Memorial Sloan Kettering Cancer Center, New York, NY, USA
| | | | - Anna Lasorella
- Department of Pediatrics, Department of Pathology, Institute for Cancer Genetics, Columbia University Medical Center, New York, NY, USA
| | - Robert Benezra
- Cancer Biology and Genetics Program, Memorial Sloan Kettering Cancer Center, New York, NY, USA.
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15
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Teixeira JR, Szeto RA, Carvalho VMA, Muotri AR, Papes F. Transcription factor 4 and its association with psychiatric disorders. Transl Psychiatry 2021; 11:19. [PMID: 33414364 PMCID: PMC7791034 DOI: 10.1038/s41398-020-01138-0] [Citation(s) in RCA: 43] [Impact Index Per Article: 10.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 07/15/2020] [Revised: 11/28/2020] [Accepted: 12/03/2020] [Indexed: 02/06/2023] Open
Abstract
The human transcription factor 4 gene (TCF4) encodes a helix-loop-helix transcription factor widely expressed throughout the body and during neural development. Mutations in TCF4 cause a devastating autism spectrum disorder known as Pitt-Hopkins syndrome, characterized by a range of aberrant phenotypes including severe intellectual disability, absence of speech, delayed cognitive and motor development, and dysmorphic features. Moreover, polymorphisms in TCF4 have been associated with schizophrenia and other psychiatric and neurological conditions. Details about how TCF4 genetic variants are linked to these diseases and the role of TCF4 during neural development are only now beginning to emerge. Here, we provide a comprehensive review of the functions of TCF4 and its protein products at both the cellular and organismic levels, as well as a description of pathophysiological mechanisms associated with this gene.
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Affiliation(s)
- José R. Teixeira
- grid.411087.b0000 0001 0723 2494Department of Genetics, Evolution, Microbiology and Immunology, Institute of Biology, University of Campinas, Campinas, São Paulo Brazil
| | - Ryan A. Szeto
- grid.266100.30000 0001 2107 4242Department of Pediatrics/Rady Children’s Hospital, School of Medicine, University of California San Diego, La Jolla, CA USA
| | - Vinicius M. A. Carvalho
- grid.411087.b0000 0001 0723 2494Department of Genetics, Evolution, Microbiology and Immunology, Institute of Biology, University of Campinas, Campinas, São Paulo Brazil ,grid.266100.30000 0001 2107 4242Department of Pediatrics/Rady Children’s Hospital, School of Medicine, University of California San Diego, La Jolla, CA USA
| | - Alysson R. Muotri
- grid.266100.30000 0001 2107 4242Department of Pediatrics/Rady Children’s Hospital, School of Medicine, University of California San Diego, La Jolla, CA USA ,grid.266100.30000 0001 2107 4242Department of Cellular & Molecular Medicine, School of Medicine, University of California San Diego, La Jolla, CA USA ,grid.266100.30000 0001 2107 4242Kavli Institute for Brain and Mind, University of California San Diego, La Jolla, CA USA ,grid.266100.30000 0001 2107 4242Center for Academic Research and Training in Anthropogeny (CARTA), University of California San Diego, La Jolla, CA USA
| | - Fabio Papes
- Department of Genetics, Evolution, Microbiology and Immunology, Institute of Biology, University of Campinas, Campinas, São Paulo, Brazil. .,Department of Pediatrics/Rady Children's Hospital, School of Medicine, University of California San Diego, La Jolla, CA, USA.
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16
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Tamberg L, Jaago M, Säälik K, Sirp A, Tuvikene J, Shubina A, Kiir CS, Nurm K, Sepp M, Timmusk T, Palgi M. Daughterless, the Drosophila orthologue of TCF4, is required for associative learning and maintenance of the synaptic proteome. Dis Model Mech 2020; 13:dmm042747. [PMID: 32641419 PMCID: PMC7406316 DOI: 10.1242/dmm.042747] [Citation(s) in RCA: 9] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/03/2019] [Accepted: 06/24/2020] [Indexed: 12/11/2022] Open
Abstract
Mammalian transcription factor 4 (TCF4) has been linked to schizophrenia and intellectual disabilities, such as Pitt-Hopkins syndrome (PTHS). Here, we show that similarly to mammalian TCF4, fruit fly orthologue Daughterless (Da) is expressed widely in the Drosophila brain. Furthermore, silencing of da, using several central nervous system-specific Gal4 driver lines, impairs appetitive associative learning of the larvae and leads to decreased levels of the synaptic proteins Synapsin (Syn) and Discs large 1 (Dlg1), suggesting the involvement of Da in memory formation. Here, we demonstrate that Syn and dlg1 are direct target genes of Da in adult Drosophila heads, as Da binds to the regulatory regions of these genes and the modulation of Da levels alter the levels of Syn and dlg1 mRNA. Silencing of da also affects negative geotaxis of the adult flies, suggesting the impairment of locomotor function. Overall, our findings suggest that Da regulates Drosophila larval memory and adult negative geotaxis, possibly via its synaptic target genes Syn and dlg1 These behavioural phenotypes can be further used as a PTHS model to screen for therapeutics.This article has an associated First Person interview with the first author of the paper.
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Affiliation(s)
- Laura Tamberg
- Department of Chemistry and Biotechnology, Tallinn University of Technology, Akadeemia tee 15, Tallinn 12618, Estonia
| | - Mariliis Jaago
- Department of Chemistry and Biotechnology, Tallinn University of Technology, Akadeemia tee 15, Tallinn 12618, Estonia
- Protobios LLC, Mäealuse 4, Tallinn 12618, Estonia
| | - Kristi Säälik
- Department of Chemistry and Biotechnology, Tallinn University of Technology, Akadeemia tee 15, Tallinn 12618, Estonia
| | - Alex Sirp
- Department of Chemistry and Biotechnology, Tallinn University of Technology, Akadeemia tee 15, Tallinn 12618, Estonia
| | - Jürgen Tuvikene
- Department of Chemistry and Biotechnology, Tallinn University of Technology, Akadeemia tee 15, Tallinn 12618, Estonia
- Protobios LLC, Mäealuse 4, Tallinn 12618, Estonia
| | - Anastassia Shubina
- Department of Chemistry and Biotechnology, Tallinn University of Technology, Akadeemia tee 15, Tallinn 12618, Estonia
| | - Carl Sander Kiir
- Department of Chemistry and Biotechnology, Tallinn University of Technology, Akadeemia tee 15, Tallinn 12618, Estonia
| | - Kaja Nurm
- Department of Chemistry and Biotechnology, Tallinn University of Technology, Akadeemia tee 15, Tallinn 12618, Estonia
| | - Mari Sepp
- Department of Chemistry and Biotechnology, Tallinn University of Technology, Akadeemia tee 15, Tallinn 12618, Estonia
| | - Tõnis Timmusk
- Department of Chemistry and Biotechnology, Tallinn University of Technology, Akadeemia tee 15, Tallinn 12618, Estonia
- Protobios LLC, Mäealuse 4, Tallinn 12618, Estonia
| | - Mari Palgi
- Department of Chemistry and Biotechnology, Tallinn University of Technology, Akadeemia tee 15, Tallinn 12618, Estonia
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17
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Rao C, Malaguti M, Mason JO, Lowell S. The transcription factor E2A drives neural differentiation in pluripotent cells. Development 2020; 147:dev184093. [PMID: 32487737 PMCID: PMC7328008 DOI: 10.1242/dev.184093] [Citation(s) in RCA: 11] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/21/2019] [Accepted: 05/26/2020] [Indexed: 12/21/2022]
Abstract
The intrinsic mechanisms that link extracellular signalling to the onset of neural differentiation are not well understood. In pluripotent mouse cells, BMP blocks entry into the neural lineage via transcriptional upregulation of inhibitor of differentiation (Id) factors. We have previously identified the major binding partner of Id proteins in pluripotent cells as the basic helix-loop-helix (bHLH) transcription factor (TF) E2A. Id1 can prevent E2A from forming heterodimers with bHLH TFs or from forming homodimers. Here, we show that overexpression of a forced E2A homodimer is sufficient to drive robust neural commitment in pluripotent cells, even under non-permissive conditions. Conversely, we find that E2A null cells display a defect in their neural differentiation capacity. E2A acts as an upstream activator of neural lineage genes, including Sox1 and Foxd4, and as a repressor of Nodal signalling. Our results suggest a crucial role for E2A in establishing neural lineage commitment in pluripotent cells.
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Affiliation(s)
- Chandrika Rao
- MRC Centre for Regenerative Medicine, Institute for Stem Cell Research, School of Biological Sciences, University of Edinburgh, 5 Little France Drive, Edinburgh EH16 4UU, UK
| | - Mattias Malaguti
- MRC Centre for Regenerative Medicine, Institute for Stem Cell Research, School of Biological Sciences, University of Edinburgh, 5 Little France Drive, Edinburgh EH16 4UU, UK
| | - John O Mason
- Centre for Discovery Brain Sciences, University of Edinburgh, 15 George Square, Edinburgh EH8 9XD, UK
- Simons Initiative for the Developing Brain, Hugh Robson Building, George Square, Edinburgh EH8 9XD, UK
| | - Sally Lowell
- MRC Centre for Regenerative Medicine, Institute for Stem Cell Research, School of Biological Sciences, University of Edinburgh, 5 Little France Drive, Edinburgh EH16 4UU, UK
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18
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Sharma S, Plotkin M. Id1 expression in kidney endothelial cells protects against diabetes-induced microvascular injury. FEBS Open Bio 2020; 10:1447-1462. [PMID: 31957231 PMCID: PMC7396439 DOI: 10.1002/2211-5463.12793] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/24/2019] [Revised: 11/25/2019] [Accepted: 01/16/2020] [Indexed: 11/08/2022] Open
Abstract
The inhibitor of differentiation (Id) transcription regulators, which are induced in response to oxidative stress, promote cell proliferation and inhibit senescence. Inhibitor of differentiation 1 (Id1) expression is limited to endothelial cells (EC) in the normal mouse kidney and is required for a normal response to injury. Endothelial dysfunction leads to the development of diabetic nephropathy, and so, we hypothesized that endothelial Id1 may help protect against hyperglycemia-induced microvascular injury and nephropathy. Here, we tested this hypothesis by using streptozotocin to induce diabetes in Id1 knockout (KO) mice and WT B6;129 littermates and examining the mice at 3 months. Expression of Id1 was observed to be increased 15-fold in WT kidney EC, and Id1 KO mice exhibited increased mesangial and myofibroblast proliferation, matrix deposition, and albuminuria compared with WT mice. Electron microscopy demonstrated peritubular capillary EC injury and lumen narrowing, and fluorescence microangiography showed a 45% reduction in capillary perfusion area with no reduction in CD31-stained areas in Id1 KO mice. Microarray analysis of EC isolated from WT and KO control and diabetic mice demonstrated activation of senescence pathways in KO cells. Kidneys from KO diabetic mice showed increased histological expression of senescence markers. In addition, premature senescence in cultured KO EC was also seen in response to oxidative stress. In conclusion, endothelial Id1 upregulation with hyperglycemia protects against microvascular injury and senescence and subsequent nephropathy.
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Affiliation(s)
| | - Matthew Plotkin
- Department of Nephrology, John L. McClellan VA Hospital, University of Arkansas for Medical Sciences, Little Rock, AR, USA
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19
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Wojnarowicz PM, Lima E Silva R, Ohnaka M, Lee SB, Chin Y, Kulukian A, Chang SH, Desai B, Garcia Escolano M, Shah R, Garcia-Cao M, Xu S, Kadam R, Goldgur Y, Miller MA, Ouerfelli O, Yang G, Arakawa T, Albanese SK, Garland WA, Stoller G, Chaudhary J, Norton L, Soni RK, Philip J, Hendrickson RC, Iavarone A, Dannenberg AJ, Chodera JD, Pavletich N, Lasorella A, Campochiaro PA, Benezra R. A Small-Molecule Pan-Id Antagonist Inhibits Pathologic Ocular Neovascularization. Cell Rep 2019; 29:62-75.e7. [PMID: 31577956 PMCID: PMC6896334 DOI: 10.1016/j.celrep.2019.08.073] [Citation(s) in RCA: 31] [Impact Index Per Article: 5.2] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/14/2018] [Revised: 08/09/2019] [Accepted: 08/23/2019] [Indexed: 02/01/2023] Open
Abstract
Id helix-loop-helix (HLH) proteins (Id1-4) bind E protein bHLH transcription factors, preventing them from forming active transcription complexes that drive changes in cell states. Id proteins are primarily expressed during development to inhibit differentiation, but they become re-expressed in adult tissues in diseases of the vasculature and cancer. We show that the genetic loss of Id1/Id3 reduces ocular neovascularization in mouse models of wet age-related macular degeneration (AMD) and retinopathy of prematurity (ROP). An in silico screen identifies AGX51, a small-molecule Id antagonist. AGX51 inhibits the Id1-E47 interaction, leading to ubiquitin-mediated degradation of Ids, cell growth arrest, and reduced viability. AGX51 is well-tolerated in mice and phenocopies the genetic loss of Id expression in AMD and ROP models by inhibiting retinal neovascularization. Thus, AGX51 is a first-in-class compound that antagonizes an interaction formerly considered undruggable and that may have utility in the management of multiple diseases.
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Affiliation(s)
- Paulina M Wojnarowicz
- Cancer Biology and Genetics Program, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA
| | - Raquel Lima E Silva
- Departments of Ophthalmology and Neuroscience, The Johns Hopkins University School of Medicine, Baltimore, MD 21287, USA
| | - Masayuki Ohnaka
- Departments of Ophthalmology and Neuroscience, The Johns Hopkins University School of Medicine, Baltimore, MD 21287, USA
| | - Sang Bae Lee
- Institute for Cancer Genetics, Columbia University Medical Center, New York, NY 10032, USA
| | - Yvette Chin
- Cancer Biology and Genetics Program, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA
| | - Anita Kulukian
- Cancer Biology and Genetics Program, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA
| | - Sung-Hee Chang
- Department of Medicine, Weill Cornell Medical College, New York, NY 10065, USA
| | - Bina Desai
- Cancer Biology and Genetics Program, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA
| | - Marta Garcia Escolano
- Cancer Biology and Genetics Program, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA
| | - Riddhi Shah
- Cancer Biology and Genetics Program, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA
| | - Marta Garcia-Cao
- Cancer Biology and Genetics Program, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA
| | - Sijia Xu
- Cancer Biology and Genetics Program, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA
| | - Rashmi Kadam
- Cancer Biology and Genetics Program, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA
| | - Yehuda Goldgur
- Structural Biology Program, Sloan Kettering Institute, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA
| | - Meredith A Miller
- Structural Biology Program, Sloan Kettering Institute, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA
| | - Ouathek Ouerfelli
- Organic Synthesis Core Facility, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA
| | - Guangli Yang
- Organic Synthesis Core Facility, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA
| | - Tsutomu Arakawa
- Alliance Protein Laboratories, a Division of KBI Biopharma, San Diego, CA 92121, USA
| | - Steven K Albanese
- Computational Biology Program, Sloan Kettering Institute, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA
| | | | - Glenn Stoller
- Ophthalmic Consultants of Long Island, Lynbrook, NY 11563, USA
| | - Jaideep Chaudhary
- Center for Cancer Research and Therapeutic Development, Clark Atlanta University, Atlanta, GA 30314, USA
| | - Larry Norton
- Evelyn H. Lauder Breast Center, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA
| | - Rajesh Kumar Soni
- Proteomics & Microchemistry Core Facility, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA
| | - John Philip
- Proteomics & Microchemistry Core Facility, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA
| | - Ronald C Hendrickson
- Proteomics & Microchemistry Core Facility, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA
| | - Antonio Iavarone
- Department of Neurology, Department of Pathology, Institute for Cancer Genetics, Columbia University Medical Center, New York, NY 10032, USA
| | - Andrew J Dannenberg
- Department of Medicine, Weill Cornell Medical College, New York, NY 10065, USA
| | - John D Chodera
- Computational Biology Program, Sloan Kettering Institute, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA
| | - Nikola Pavletich
- Structural Biology Program, Sloan Kettering Institute, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA
| | - Anna Lasorella
- Department of Pediatrics, Department of Pathology, Institute for Cancer Genetics, Columbia University Medical Center, New York, NY 10032, USA
| | - Peter A Campochiaro
- Departments of Ophthalmology and Neuroscience, The Johns Hopkins University School of Medicine, Baltimore, MD 21287, USA
| | - Robert Benezra
- Cancer Biology and Genetics Program, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA.
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20
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Schneeberger PE, Bierhals T, Neu A, Hempel M, Kutsche K. de novo MEPCE nonsense variant associated with a neurodevelopmental disorder causes disintegration of 7SK snRNP and enhanced RNA polymerase II activation. Sci Rep 2019; 9:12516. [PMID: 31467394 PMCID: PMC6715695 DOI: 10.1038/s41598-019-49032-0] [Citation(s) in RCA: 11] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/08/2019] [Accepted: 08/19/2019] [Indexed: 02/06/2023] Open
Abstract
In eukaryotes, the elongation phase of transcription by RNA polymerase II (RNAP II) is regulated by the transcription elongation factor b (P-TEFb), composed of Cyclin-T1 and cyclin-dependent kinase 9. The release of RNAP II is mediated by phosphorylation through P-TEFb that in turn is under control by the inhibitory 7SK small nuclear ribonucleoprotein (snRNP) complex. The 7SK snRNP consists of the 7SK non-coding RNA and the proteins MEPCE, LARP7, and HEXIM1/2. Biallelic LARP7 loss-of-function variants underlie Alazami syndrome characterized by growth retardation and intellectual disability. We report a boy with global developmental delay and seizures carrying the de novo MEPCE nonsense variant c.1552 C > T/p.(Arg518*). mRNA and protein analyses identified nonsense-mediated mRNA decay to underlie the decreased amount of MEPCE in patient fibroblasts followed by LARP7 and 7SK snRNA downregulation and HEXIM1 upregulation. Reduced binding of HEXIM1 to Cyclin-T1, hyperphosphorylation of the RNAP II C-terminal domain, and upregulated expression of ID2, ID3, MRPL11 and snRNAs U1, U2 and U4 in patient cells are suggestive of enhanced activation of P-TEFb. Flavopiridol treatment and ectopic MEPCE protein expression in patient fibroblasts rescued increased expression of six RNAP II-sensitive genes and suggested a possible repressive effect of MEPCE on P-TEFb-dependent transcription of specific genes.
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Affiliation(s)
- Pauline E Schneeberger
- Institute of Human Genetics, University Medical Center Hamburg-Eppendorf, Hamburg, Germany
| | - Tatjana Bierhals
- Institute of Human Genetics, University Medical Center Hamburg-Eppendorf, Hamburg, Germany
| | - Axel Neu
- Childrens Hospital, University Medical Center Hamburg-Eppendorf, Hamburg, Germany
| | - Maja Hempel
- Institute of Human Genetics, University Medical Center Hamburg-Eppendorf, Hamburg, Germany
| | - Kerstin Kutsche
- Institute of Human Genetics, University Medical Center Hamburg-Eppendorf, Hamburg, Germany.
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21
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CCN2-MAPK-Id-1 loop feedback amplification is involved in maintaining stemness in oxaliplatin-resistant hepatocellular carcinoma. Hepatol Int 2019; 13:440-453. [PMID: 31250351 PMCID: PMC6661033 DOI: 10.1007/s12072-019-09960-5] [Citation(s) in RCA: 12] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 02/21/2019] [Accepted: 06/11/2019] [Indexed: 12/24/2022]
Abstract
Background Hepatocellular carcinoma (HCC) is the second leading cause of cancer death worldwide. Chemotherapy is an alternative treatment for advanced HCCs, but chemo-resistance prevents cancer therapies from achieving stable and complete responses. Understanding the underlying mechanisms in chemo-resistance is critical to improve the efficacy of HCC. Methods The expression levels of Id-1 and CCN2 were detected in large cohorts of HCCs, and functional analyses of Id-1 and CCN2 were performed both in vitro and in vivo. cDNA microarrays were performed to evaluate the alterations of expression profiling of HCC cells with overexpression of CCN2. Finally, the role of downstream signaling of MAPK/Id-1 signaling pathway in oxaliplatin resistance were also explored. Results The increased expression of Id-1 and CCN2 were closely related to oxaliplatin resistance in HCC. Upregulation of CCN2 and Id-1 was independently associated with shorter survival and increased recurrence in HCC patients, and significantly enhanced oxaliplatin resistance and promoted lung metastasis in vivo, whereas knock-down of their expression significantly reversed the chemo-resistance and inhibited HCC cell stemness. cDNA microarrays and PCR revealed that Id-1 and MAPK pathway were the downstream signaling of CCN2. CCN2 significantly enhanced oxaliplatin resistance by activating the MAPK/Id-1 signaling pathway, and Id-1 could upregulate CCN2 in a positive feedback manner. Conclusions CCN2/MAPK/Id-1 loop feedback amplification is involved in oxaliplatin resistance, and the combination of oxaliplatin with inhibitor of CCN2 or MAPK signaling could provide a promising approach to ameliorating oxaliplatin resistance in HCC. Electronic supplementary material The online version of this article (10.1007/s12072-019-09960-5) contains supplementary material, which is available to authorized users.
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22
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On the traces of tcf12: Investigation of the gene expression pattern during development and cranial suture patterning in zebrafish (Danio rerio). PLoS One 2019; 14:e0218286. [PMID: 31188878 PMCID: PMC6561585 DOI: 10.1371/journal.pone.0218286] [Citation(s) in RCA: 6] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/22/2019] [Accepted: 05/29/2019] [Indexed: 12/11/2022] Open
Abstract
The transcription factor 12 (tcf12) is a basic Helix-Loop-Helix protein (bHLH) of the E-protein family, proven to play an important role in developmental processes like neurogenesis, mesoderm formation, and cranial vault development. In humans, mutations in TCF12 lead to craniosynostosis, a congenital birth disorder characterized by the premature fusion of one or several of the cranial sutures. Current research has been primarily focused on functional studies of TCF12, hence the cellular expression profile of this gene during embryonic development and early stages of ossification remains poorly understood. Here we present the establishment and detailed analysis of two transgenic tcf12:EGFP fluorescent zebrafish (Danio rerio) reporter lines. Using these transgenic lines, we analyzed the general spatiotemporal expression pattern of tcf12 during different developmental stages and put emphasis on skeletal development and cranial suture patterning. We identified robust tcf12 promoter-driven EGFP expression in the central nervous system (CNS), the heart, the pronephros, and the somites of zebrafish embryos. Additionally, expression was observed inside the muscles and bones of the viscerocranium in juvenile and adult fish. During cranial vault development, the transgenic fish show a high amount of tcf12 expressing cells at the growth fronts of the ossifying frontal and parietal bones and inside the emerging cranial sutures. Subsequently, we tested the transcriptional activity of three evolutionary conserved non-coding elements (CNEs) located in the tcf12 locus by transient transgenic assays and compared their in vivo activity to the expression pattern determined in the transgenic tcf12:EGFP lines. We could validate two of them as tcf12 enhancer elements driving specific gene expression in the CNS during embryogenesis. Our newly established transgenic lines enhance the understanding of tcf12 gene regulation and open up the possibilities for further functional investigation of these novel tcf12 enhancer elements in zebrafish.
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Li K, Baker NE. Transcriptional and post-transcriptional regulation of extra macrochaetae during Drosophila adult peripheral neurogenesis. Dev Biol 2019; 449:41-51. [PMID: 30771303 DOI: 10.1016/j.ydbio.2019.02.003] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/18/2018] [Revised: 02/11/2019] [Accepted: 02/11/2019] [Indexed: 11/18/2022]
Abstract
Regulation of the Drosophila ID protein Extra macrochaetae (Emc) is important because reduced Emc levels have been proposed to favor proneural gene activity and thereby define a prepattern for neurogenesis. Recent studies suggest a major role for post-translational control of Emc levels. To further define the mechanisms of Emc regulation, we identified two redundant cis-regulatory regions by germline transformation-rescue experiments that make use of new molecularly-defined emc mutants. We distinguished the mechanisms by which Daughterless (Da) regulated Emc expression, finding post-translational regulation in most tissues, and additional transcriptional regulation in the eye imaginal disc posterior to the morphogenetic furrow. Dpp and Hh signaling pathways repressed Emc transcriptionally and post-translationally within the morphogenetic furrow of the eye disc, whereas Wg signaling repressed Emc expression at the anterior margin of the wing imaginal disc. Although the emc 3' UTR is potentially regulatory, no effect of miRNA pathways on Emc protein levels was discernible. Our work supports recent evidence that post-transcriptional mechanisms contribute more to regulation of Emc protein levels than transcriptional mechanisms do.
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Affiliation(s)
- Ke Li
- Department of Genetics, Albert Einstein College of Medicine, 1300 Morris Park Avenue, Bronx NY 10461, USA
| | - Nicholas E Baker
- Department of Genetics, Albert Einstein College of Medicine, 1300 Morris Park Avenue, Bronx NY 10461, USA; Department of Developmental and Molecular Biology, Albert Einstein College of Medicine, 1300 Morris Park Avenue, Bronx NY 10461, USA; Department of Ophthalmology and Visual Sciences, Albert Einstein College of Medicine, 1300 Morris Park Avenue, Bronx NY 10461, USA.
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24
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Ishibashi T, Hatori R, Maeda R, Nakamura M, Taguchi T, Matsuyama Y, Matsuno K. E and ID proteins regulate cell chirality and left-right asymmetric development in Drosophila. Genes Cells 2019; 24:214-230. [PMID: 30624823 DOI: 10.1111/gtc.12669] [Citation(s) in RCA: 10] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/01/2018] [Revised: 12/13/2018] [Accepted: 12/28/2018] [Indexed: 02/07/2023]
Abstract
How left-right (LR) asymmetric forms in the animal body is a fundamental problem in Developmental Biology. Although the mechanisms for LR asymmetry are well studied in some species, they are still poorly understood in invertebrates. We previously showed that the intrinsic LR asymmetry of cells (designated as cell chirality) drives LR asymmetric development in the Drosophila embryonic hindgut, although the machinery of the cell chirality formation remains elusive. Here, we found that the Drosophila homologue of the Id gene, extra macrochaetae (emc), is required for the normal LR asymmetric morphogenesis of this organ. Id proteins, including Emc, are known to interact with and inhibit E-box-binding proteins (E proteins), such as Drosophila Daughterless (Da). We found that the suppression of da by wild-type emc was essential for cell chirality formation and for normal LR asymmetric development of the embryonic hindgut. Myosin ID (MyoID), which encodes the Drosophila Myosin ID protein, is known to regulate cell chirality. We further showed that Emc-Da regulates cell chirality formation, in which Emc functions upstream of or parallel to MyoID. Abnormal Id-E protein regulation is involved in various human diseases. Our results suggest that defects in cell shape may contribute to the pathogenesis of such diseases.
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Affiliation(s)
- Tomoki Ishibashi
- Department of Biological Sciences, Osaka University, Toyonaka, Osaka, Japan
| | - Ryo Hatori
- Department of Biological Sciences, Osaka University, Toyonaka, Osaka, Japan
| | - Reo Maeda
- Department of Biological Science and Technology, Tokyo University of Science, Noda, Chiba, Japan
| | | | - Tomohiro Taguchi
- Department of Biological Science and Technology, Tokyo University of Science, Noda, Chiba, Japan
| | - Yoko Matsuyama
- Department of Biological Sciences, Osaka University, Toyonaka, Osaka, Japan
| | - Kenji Matsuno
- Department of Biological Sciences, Osaka University, Toyonaka, Osaka, Japan
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25
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Different roles of E proteins in t(8;21) leukemia: E2-2 compromises the function of AETFC and negatively regulates leukemogenesis. Proc Natl Acad Sci U S A 2018; 116:890-899. [PMID: 30593567 DOI: 10.1073/pnas.1809327116] [Citation(s) in RCA: 14] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/11/2022] Open
Abstract
The AML1-ETO fusion protein, generated by the t(8;21) chromosomal translocation, is causally involved in nearly 20% of acute myeloid leukemia (AML) cases. In leukemic cells, AML1-ETO resides in and functions through a stable protein complex, AML1-ETO-containing transcription factor complex (AETFC), that contains multiple transcription (co)factors. Among these AETFC components, HEB and E2A, two members of the ubiquitously expressed E proteins, directly interact with AML1-ETO, confer new DNA-binding capacity to AETFC, and are essential for leukemogenesis. However, the third E protein, E2-2, is specifically silenced in AML1-ETO-expressing leukemic cells, suggesting E2-2 as a negative factor of leukemogenesis. Indeed, ectopic expression of E2-2 selectively inhibits the growth of AML1-ETO-expressing leukemic cells, and this inhibition requires the bHLH DNA-binding domain. RNA-seq and ChIP-seq analyses reveal that, despite some overlap, the three E proteins differentially regulate many target genes. In particular, studies show that E2-2 both redistributes AETFC to, and activates, some genes associated with dendritic cell differentiation and represses MYC target genes. In AML patients, the expression of E2-2 is relatively lower in the t(8;21) subtype, and an E2-2 target gene, THPO, is identified as a potential predictor of relapse. In a mouse model of human t(8;21) leukemia, E2-2 suppression accelerates leukemogenesis. Taken together, these results reveal that, in contrast to HEB and E2A, which facilitate AML1-ETO-mediated leukemogenesis, E2-2 compromises the function of AETFC and negatively regulates leukemogenesis. The three E proteins thus define a heterogeneity of AETFC, which improves our understanding of the precise mechanism of leukemogenesis and assists development of diagnostic/therapeutic strategies.
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26
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Spatial regulation of expanded transcription in the Drosophila wing imaginal disc. PLoS One 2018; 13:e0201317. [PMID: 30063727 PMCID: PMC6067730 DOI: 10.1371/journal.pone.0201317] [Citation(s) in RCA: 6] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/09/2018] [Accepted: 07/12/2018] [Indexed: 12/13/2022] Open
Abstract
Growth and patterning are coordinated during development to define organ size and shape. The growth, proliferation and differentiation of Drosophila wings are regulated by several conserved signaling pathways. Here, we show that the Salvador-Warts-Hippo (SWH) and Notch pathways converge on an enhancer in the expanded (ex) gene, which also responds to levels of the bHLH transcription factor Daughterless (Da). Separate cis-regulatory elements respond to Salvador-Warts-Hippo (SWH) and Notch pathways, to bHLH proteins, and to unidentified factors that repress ex transcription in the wing pouch and in the proneural region at the anterior wing margin. Senseless, a zinc-finger transcription factor acting in proneural regions, had a negative impact on ex transcription in the proneural region, but the transcriptional repressor Hairy had no effect. Our study suggests that a complex pattern of ex transcription results from integration of a uniform SWH signal with multiple other inputs, rather than from a pattern of SWH signaling.
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27
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Li K, Baker NE. Regulation of the Drosophila ID protein Extra macrochaetae by proneural dimerization partners. eLife 2018; 7:33967. [PMID: 29687780 PMCID: PMC5915177 DOI: 10.7554/elife.33967] [Citation(s) in RCA: 13] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/30/2017] [Accepted: 03/25/2018] [Indexed: 12/13/2022] Open
Abstract
Proneural bHLH proteins are transcriptional regulators of neural fate specification. Extra macrochaetae (Emc) forms inactive heterodimers with both proneural bHLH proteins and their bHLH partners (represented in Drosophila by Daughterless). It is generally thought that varying levels of Emc define a prepattern that determines where proneural bHLH genes can be effective. We report that instead it is the bHLH proteins that determine the pattern of Emc levels. Daughterless level sets Emc protein levels in most cells, apparently by stabilizing Emc in heterodimers. Emc is destabilized in proneural regions by local competition for heterodimer formation by proneural bHLH proteins including Atonal or AS-C proteins. Reflecting this post-translational control through protein stability, uniform emc transcription is sufficient for almost normal patterns of neurogenesis. Protein stability regulated by exchanges between bHLH protein dimers could be a feature of bHLH-mediated developmental events.
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Affiliation(s)
- Ke Li
- Department of Genetics, Albert Einstein College of Medicine, Bronx, United States
| | - Nicholas E Baker
- Department of Genetics, Albert Einstein College of Medicine, Bronx, United States.,Department of Developmental and Molecular Biology, Albert Einstein College of Medicine, Bronx, United States.,Department of Ophthalmology and Visual Sciences, Albert Einstein College of Medicine, Bronx, United States
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28
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Lan Q, Cao M, Kollipara RK, Rosa JB, Kittler R, Jiang H. FoxA transcription factor Fork head maintains the intestinal stem/progenitor cell identities in Drosophila. Dev Biol 2017; 433:324-343. [PMID: 29108672 DOI: 10.1016/j.ydbio.2017.09.002] [Citation(s) in RCA: 14] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/13/2017] [Revised: 08/21/2017] [Accepted: 09/03/2017] [Indexed: 01/13/2023]
Abstract
Understanding how somatic stem cells respond to tissue needs is important, since aberrant somatic stem cell behaviors may lead to tissue degeneration or tumorigenesis. Here, from an in vivo RNAi screen targeting transcription factors that regulate intestinal regeneration, we uncovered a requirement for the Drosophila FoxA transcription factor Fork head (Fkh) in the maintenance of intestinal stem/progenitor cell identities. FoxA/Fkh maintains the expressions of stem/progenitor cell markers and is required for stem cell proliferation during intestinal homeostasis and regeneration. Furthermore, FoxA/Fkh prevents the intestinal stem/progenitor cells from precocious differentiation into the Enterocyte lineage, likely in cooperation with the transcription factor bHLH/Daughterless (Da). In addition, loss of FoxA/Fkh suppresses the intestinal tumorigenesis caused by Notch pathway inactivation. To reveal the gene program underlying stem/progenitor cell identities, we profiled the genome-wide chromatin binding sites of transcription factors Fkh and Da, and interestingly, around half of Fkh binding regions are shared by Da, further suggesting their collaborative roles. Finally, we identified the genes associated with their shared binding regions. This comprehensive gene list may contain stem/progenitor maintenance factors functioning downstream of Fkh and Da, and would be helpful for future gene discoveries in the Drosophila intestinal stem cell lineage.
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Affiliation(s)
- Qing Lan
- Department of Molecular Biology, University of Texas Southwestern Medical Center, Dallas, TX 75390, United States.
| | - Min Cao
- Department of Molecular Biology, University of Texas Southwestern Medical Center, Dallas, TX 75390, United States
| | - Rahul K Kollipara
- Eugene McDermott Center for Human Growth and Development, University of Texas Southwestern Medical Center, Dallas, TX 75390, United States
| | - Jeffrey B Rosa
- Department of Molecular Biology, University of Texas Southwestern Medical Center, Dallas, TX 75390, United States
| | - Ralf Kittler
- Eugene McDermott Center for Human Growth and Development, University of Texas Southwestern Medical Center, Dallas, TX 75390, United States
| | - Huaqi Jiang
- Department of Molecular Biology, University of Texas Southwestern Medical Center, Dallas, TX 75390, United States.
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29
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Bhattacharya A, Li K, Quiquand M, Rimesso G, Baker NE. The Notch pathway regulates the Second Mitotic Wave cell cycle independently of bHLH proteins. Dev Biol 2017; 431:309-320. [PMID: 28919436 DOI: 10.1016/j.ydbio.2017.08.035] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/23/2016] [Revised: 02/08/2017] [Accepted: 08/22/2017] [Indexed: 12/31/2022]
Abstract
Notch regulates both neurogenesis and cell cycle activity to coordinate precursor cell generation in the differentiating Drosophila eye. Mosaic analysis with mitotic clones mutant for Notch components was used to identify the pathway of Notch signaling that regulates the cell cycle in the Second Mitotic Wave. Although S phase entry depends on Notch signaling and on the transcription factor Su(H), the transcriptional co-activator Mam and the bHLH repressor genes of the E(spl)-Complex were not essential, although these are Su(H) coactivators and targets during the regulation of neurogenesis. The Second Mitotic Wave showed little dependence on ubiquitin ligases neuralized or mindbomb, and although the ligand Delta is required non-autonomously, partial cell cycle activity occurred in the absence of known Notch ligands. We found that myc was not essential for the Second Mitotic Wave. The Second Mitotic Wave did not require the HLH protein Extra macrochaetae, and the bHLH protein Daughterless was required only cell-nonautonomously. Similar cell cycle phenotypes for Daughterless and Atonal were consistent with requirement for neuronal differentiation to stimulate Delta expression, affecting Notch activity in the Second Mitotic Wave indirectly. Therefore Notch signaling acts to regulate the Second Mitotic Wave without activating bHLH gene targets.
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Affiliation(s)
- Abhishek Bhattacharya
- Department of Genetics, Albert Einstein College of Medicine, 1300 Morris Park Avenue, Bronx, NY 10461, USA
| | - Ke Li
- Department of Genetics, Albert Einstein College of Medicine, 1300 Morris Park Avenue, Bronx, NY 10461, USA
| | - Manon Quiquand
- Department of Genetics, Albert Einstein College of Medicine, 1300 Morris Park Avenue, Bronx, NY 10461, USA
| | - Gerard Rimesso
- Department of Genetics, Albert Einstein College of Medicine, 1300 Morris Park Avenue, Bronx, NY 10461, USA
| | - Nicholas E Baker
- Department of Genetics, Albert Einstein College of Medicine, 1300 Morris Park Avenue, Bronx, NY 10461, USA; Department of Developmental and Molecular Biology, Albert Einstein College of Medicine, 1300 Morris Park Avenue, Bronx, NY 10461, USA; Department of Ophthalmology and Visual Sciences, Albert Einstein College of Medicine, 1300 Morris Park Avenue, Bronx, NY 10461, USA.
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30
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Neto M, Naval-Sánchez M, Potier D, Pereira PS, Geerts D, Aerts S, Casares F. Nuclear receptors connect progenitor transcription factors to cell cycle control. Sci Rep 2017; 7:4845. [PMID: 28687780 PMCID: PMC5501803 DOI: 10.1038/s41598-017-04936-7] [Citation(s) in RCA: 13] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/18/2016] [Accepted: 05/23/2017] [Indexed: 01/31/2023] Open
Abstract
The specification and growth of organs is controlled simultaneously by networks of transcription factors. While the connection between these transcription factors with fate determinants is increasingly clear, how they establish the link with the cell cycle is far less understood. Here we investigate this link in the developing Drosophila eye, where two transcription factors, the MEIS1 homologue hth and the Zn-finger tsh, synergize to stimulate the proliferation of naïve eye progenitors. Experiments combining transcriptomics, open-chromatin profiling, motif analysis and functional assays indicate that these progenitor transcription factors exert a global regulation of the proliferation program. Rather than directly regulating cell cycle genes, they control proliferation through an intermediary layer of nuclear receptors of the ecdysone/estrogen-signaling pathway. This regulatory subnetwork between hth, tsh and nuclear receptors might be conserved from Drosophila to mammals, as we find a significant co-overexpression of their human homologues in specific cancer types.
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Affiliation(s)
- Marta Neto
- CABD, Andalusian Centre for Developmental Biology, CSIC-UPO-JA, 41013, Seville, Spain.,i3S - Instituto de Investigação e Inovação em Saúde, Universidade do Porto, Rua Alfredo Allen, 208, 4200-135, Porto, Portugal.,IBMC - Instituto de Biologia Molecular e Celular, Universidade do Porto, Rua Alfredo Allen, 208, 4200-135, Porto, Portugal
| | | | - Delphine Potier
- School of Medicine, University of Leuven, box 602 3000, Leuven, Belgium
| | - Paulo S Pereira
- i3S - Instituto de Investigação e Inovação em Saúde, Universidade do Porto, Rua Alfredo Allen, 208, 4200-135, Porto, Portugal.,IBMC - Instituto de Biologia Molecular e Celular, Universidade do Porto, Rua Alfredo Allen, 208, 4200-135, Porto, Portugal
| | - Dirk Geerts
- Department of Medical Biology L2-109, Academic Medical Center, University of Amsterdam, Amsterdam, The Netherlands
| | - Stein Aerts
- School of Medicine, University of Leuven, box 602 3000, Leuven, Belgium.
| | - Fernando Casares
- CABD, Andalusian Centre for Developmental Biology, CSIC-UPO-JA, 41013, Seville, Spain.
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31
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Corson F, Couturier L, Rouault H, Mazouni K, Schweisguth F. Self-organized Notch dynamics generate stereotyped sensory organ patterns in Drosophila. Science 2017; 356:science.aai7407. [PMID: 28386027 DOI: 10.1126/science.aai7407] [Citation(s) in RCA: 85] [Impact Index Per Article: 10.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/06/2016] [Accepted: 03/20/2017] [Indexed: 12/26/2022]
Abstract
The emergence of spatial patterns in developing multicellular organisms relies on positional cues and cell-cell communication. Drosophila sensory organs have informed a paradigm in which these operate in two distinct steps: Prepattern factors drive localized proneural activity, then Notch-mediated lateral inhibition singles out neural precursors. Here we show that self-organization through Notch signaling also establishes the proneural stripes that resolve into rows of sensory bristles on the fly thorax. Patterning, initiated by a gradient of Delta ligand expression, progresses through inhibitory signaling between and within stripes. Thus, Notch signaling can support self-organized tissue patterning as a prepattern is transduced by cell-cell interactions into a refined arrangement of cellular fates.
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Affiliation(s)
- Francis Corson
- Laboratoire de Physique Statistique, Ecole Normale Supérieure, CNRS, Université Pierre et Marie Curie, Université Paris Diderot, 75005 Paris, France.
| | - Lydie Couturier
- Department of Developmental and Stem Cell Biology, Institut Pasteur, 75015 Paris, France.,CNRS, UMR3738, 75015 Paris, France
| | - Hervé Rouault
- Department of Developmental and Stem Cell Biology, Institut Pasteur, 75015 Paris, France.,CNRS, UMR3738, 75015 Paris, France
| | - Khalil Mazouni
- Department of Developmental and Stem Cell Biology, Institut Pasteur, 75015 Paris, France.,CNRS, UMR3738, 75015 Paris, France
| | - François Schweisguth
- Department of Developmental and Stem Cell Biology, Institut Pasteur, 75015 Paris, France. .,CNRS, UMR3738, 75015 Paris, France
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32
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Jang S, An G, Li HY. Rice Leaf Angle and Grain Size Are Affected by the OsBUL1 Transcriptional Activator Complex. PLANT PHYSIOLOGY 2017; 173:688-702. [PMID: 27879391 PMCID: PMC5210763 DOI: 10.1104/pp.16.01653] [Citation(s) in RCA: 82] [Impact Index Per Article: 10.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 10/25/2016] [Accepted: 11/18/2016] [Indexed: 05/04/2023]
Abstract
Rice atypical HLH protein Oryza sativa BRASSINOSTEROID UPREGULATED 1-LIKE1 (OsBUL1) is preferentially expressed in the lamina joint where it controls cell elongation and positively affects leaf angles. OsBUL1 knockout mutant (osbul1) and transgenic rice for double-stranded RNA interference (dsRNAi) of OsBUL1 produced erect leaves with smaller grains, whereas OsBUL1 overexpressors and an activation tagging line of OsBUL1 exhibited increased lamina inclination and grain size. Moreover, OsBUL1 expression was induced by brassinolide (BL) and osbul1 did not respond to BL treatment. To understand the molecular network of OsBUL1 function in rice, we isolated a novel OsBUL1-interacting protein, LO9-177, an uncharacterized protein containing a KxDL motif, and functionally studied it with respect to the lamina inclination and grain size of rice. OsBUL1 COMPLEX1 (OsBC1) is a basic helix-loop-helix (bHLH) transcriptional activator that interacts with OsBUL1 only in the presence of LO9-177 forming a possible trimeric complex for cell elongation in the lamina joint of rice. Expression of OsBC1 is also upregulated by BL and has a similar pattern to that of OsBUL1 Transgenic rice plants expressing OsBC1 under the control of OsBUL1 promoter showed increased grain size as well as leaf bending, while transgenic lines for dsRNAi and/or expressing a dominant repressor form of OsBC1 displayed reduced plant height and grain size. Together, these results demonstrated that a novel protein complex consisting of OsBUL1, LO9-177, and OsBC1 is associated with the HLH-bHLH system, providing new insight into the molecular functional network based on HLH-bHLH proteins for cell elongation.
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Affiliation(s)
- Seonghoe Jang
- Biotechnology Center in Southern Taiwan, Tainan 74145/Agricultural Biotechnology Research Center, Academia Sinica, Nankang, Taipei 11529, Taiwan (S.J., H.-Y.L.);
- Institute of Tropical Plant Science, National Cheng Kung University, Tainan 70101, Taiwan (S.J.); and
- Crop Biotech Institute, Kyung Hee University, Yongin 17102, Korea (G.A.)
| | - Gynheung An
- Biotechnology Center in Southern Taiwan, Tainan 74145/Agricultural Biotechnology Research Center, Academia Sinica, Nankang, Taipei 11529, Taiwan (S.J., H.-Y.L.)
- Institute of Tropical Plant Science, National Cheng Kung University, Tainan 70101, Taiwan (S.J.); and
- Crop Biotech Institute, Kyung Hee University, Yongin 17102, Korea (G.A.)
| | - Hsing-Yi Li
- Biotechnology Center in Southern Taiwan, Tainan 74145/Agricultural Biotechnology Research Center, Academia Sinica, Nankang, Taipei 11529, Taiwan (S.J., H.-Y.L.)
- Institute of Tropical Plant Science, National Cheng Kung University, Tainan 70101, Taiwan (S.J.); and
- Crop Biotech Institute, Kyung Hee University, Yongin 17102, Korea (G.A.)
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33
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D'Rozario M, Zhang T, Waddell EA, Zhang Y, Sahin C, Sharoni M, Hu T, Nayal M, Kutty K, Liebl F, Hu W, Marenda DR. Type I bHLH Proteins Daughterless and Tcf4 Restrict Neurite Branching and Synapse Formation by Repressing Neurexin in Postmitotic Neurons. Cell Rep 2016; 15:386-97. [PMID: 27050508 DOI: 10.1016/j.celrep.2016.03.034] [Citation(s) in RCA: 25] [Impact Index Per Article: 2.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/21/2015] [Revised: 01/22/2016] [Accepted: 03/09/2016] [Indexed: 11/17/2022] Open
Abstract
Proneural proteins of the class I/II basic-helix-loop-helix (bHLH) family are highly conserved transcription factors. Class I bHLH proteins are expressed in a broad number of tissues during development, whereas class II bHLH protein expression is more tissue restricted. Our understanding of the function of class I/II bHLH transcription factors in both invertebrate and vertebrate neurobiology is largely focused on their function as regulators of neurogenesis. Here, we show that the class I bHLH proteins Daughterless and Tcf4 are expressed in postmitotic neurons in Drosophila melanogaster and mice, respectively, where they function to restrict neurite branching and synapse formation. Our data indicate that Daughterless performs this function in part by restricting the expression of the cell adhesion molecule Neurexin. This suggests a role for these proteins outside of their established roles in neurogenesis.
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Affiliation(s)
| | - Ting Zhang
- Department of Neuroscience, Temple University School of Medicine, Philadelphia, PA 19140, USA
| | - Edward A Waddell
- Department of Biology, Drexel University, Philadelphia, PA 19104, USA
| | - Yonggang Zhang
- Department of Neuroscience, Temple University School of Medicine, Philadelphia, PA 19140, USA
| | - Cem Sahin
- Department of Electrical and Computer Engineering, Drexel University, Philadelphia, PA 19104, USA
| | - Michal Sharoni
- Department of Biology, Drexel University, Philadelphia, PA 19104, USA
| | - Tina Hu
- Department of Biology, Drexel University, Philadelphia, PA 19104, USA
| | - Mohammad Nayal
- Department of Biology, Drexel University, Philadelphia, PA 19104, USA
| | - Kaveesh Kutty
- Department of Biology, Drexel University, Philadelphia, PA 19104, USA
| | - Faith Liebl
- Department of Biological Sciences, Southern Illinois University Edwardsville, Edwardsville, IL 62026, USA
| | - Wenhui Hu
- Department of Neuroscience, Temple University School of Medicine, Philadelphia, PA 19140, USA.
| | - Daniel R Marenda
- Department of Biology, Drexel University, Philadelphia, PA 19104, USA; Department of Neurobiology and Anatomy, Drexel University College of Medicine, Philadelphia, PA 19104, USA.
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Wang LH, Baker NE. E Proteins and ID Proteins: Helix-Loop-Helix Partners in Development and Disease. Dev Cell 2016; 35:269-80. [PMID: 26555048 DOI: 10.1016/j.devcel.2015.10.019] [Citation(s) in RCA: 124] [Impact Index Per Article: 13.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/08/2015] [Revised: 10/18/2015] [Accepted: 10/23/2015] [Indexed: 01/12/2023]
Abstract
The basic Helix-Loop-Helix (bHLH) proteins represent a well-known class of transcriptional regulators. Many bHLH proteins act as heterodimers with members of a class of ubiquitous partners, the E proteins. A widely expressed class of inhibitory heterodimer partners-the Inhibitor of DNA-binding (ID) proteins-also exists. Genetic and molecular analyses in humans and in knockout mice implicate E proteins and ID proteins in a wide variety of diseases, belying the notion that they are non-specific partner proteins. Here, we explore relationships of E proteins and ID proteins to a variety of disease processes and highlight gaps in knowledge of disease mechanisms.
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Affiliation(s)
- Lan-Hsin Wang
- Department of Genetics, Albert Einstein College of Medicine, 1300 Morris Park Avenue, Bronx, NY 10461, USA
| | - Nicholas E Baker
- Department of Genetics, Albert Einstein College of Medicine, 1300 Morris Park Avenue, Bronx, NY 10461, USA; Department of Developmental and Molecular Biology, Albert Einstein College of Medicine, 1300 Morris Park Avenue, Bronx, NY 10461, USA; Department of Ophthalmology and Visual Sciences, Albert Einstein College of Medicine, 1300 Morris Park Avenue, Bronx, NY 10461, USA.
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35
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Tamberg L, Sepp M, Timmusk T, Palgi M. Introducing Pitt-Hopkins syndrome-associated mutations of TCF4 to Drosophila daughterless. Biol Open 2015; 4:1762-71. [PMID: 26621827 PMCID: PMC4736037 DOI: 10.1242/bio.014696] [Citation(s) in RCA: 16] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/23/2022] Open
Abstract
Pitt-Hopkins syndrome (PTHS) is caused by haploinsufficiency of Transcription factor 4 (TCF4), one of the three human class I basic helix-loop-helix transcription factors called E-proteins. Drosophila has a single E-protein, Daughterless (Da), homologous to all three mammalian counterparts. Here we show that human TCF4 can rescue Da deficiency during fruit fly nervous system development. Overexpression of Da or TCF4 specifically in adult flies significantly decreases their survival rates, indicating that these factors are crucial even after development has been completed. We generated da transgenic fruit fly strains with corresponding missense mutations R578H, R580W, R582P and A614V found in TCF4 of PTHS patients and studied the impact of these mutations in vivo. Overexpression of wild type Da as well as human TCF4 in progenitor tissues induced ectopic sensory bristles and the rough eye phenotype. By contrast, overexpression of DaR580W and DaR582P that disrupt DNA binding reduced the number of bristles and induced the rough eye phenotype with partial lack of pigmentation, indicating that these act dominant negatively. Compared to the wild type, DaR578H and DaA614V were less potent in induction of ectopic bristles and the rough eye phenotype, respectively, suggesting that these are hypomorphic. All studied PTHS-associated mutations that we introduced into Da led to similar effects in vivo as the same mutations in TCF4 in vitro. Consequently, our Drosophila models of PTHS are applicable for further studies aiming to unravel the molecular mechanisms of this disorder. Summary: Introducing mutations of the TCF4 gene found in human patients into its fly orthologue daughterless allows the generation of Drosophila models for research into Pitt-Hopkins syndrome.
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Affiliation(s)
- Laura Tamberg
- Laboratory of Molecular Neurobiology, Department of Gene Technology, Tallinn University of Technology, Akadeemia Rd.15, Tallinn 12618, Estonia
| | - Mari Sepp
- Laboratory of Molecular Neurobiology, Department of Gene Technology, Tallinn University of Technology, Akadeemia Rd.15, Tallinn 12618, Estonia
| | - Tõnis Timmusk
- Laboratory of Molecular Neurobiology, Department of Gene Technology, Tallinn University of Technology, Akadeemia Rd.15, Tallinn 12618, Estonia
| | - Mari Palgi
- Laboratory of Molecular Neurobiology, Department of Gene Technology, Tallinn University of Technology, Akadeemia Rd.15, Tallinn 12618, Estonia
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36
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The elimination of an adult segment by the Hox gene Abdominal-B. Mech Dev 2015; 138 Pt 2:210-217. [DOI: 10.1016/j.mod.2015.08.002] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/28/2015] [Revised: 07/31/2015] [Accepted: 08/01/2015] [Indexed: 11/19/2022]
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37
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Zhang T, Du W. Groucho restricts rhomboid expression and couples EGFR activation with R8 selection during Drosophila photoreceptor differentiation. Dev Biol 2015; 407:246-55. [PMID: 26417727 DOI: 10.1016/j.ydbio.2015.09.011] [Citation(s) in RCA: 9] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/05/2015] [Revised: 07/18/2015] [Accepted: 09/24/2015] [Indexed: 11/17/2022]
Abstract
Notch and EGFR signaling pathways play important roles in photoreceptor differentiation during Drosophila eye development. Notch signaling induces Enhancer of Split (E(spl)) proteins to repress atonal (ato) expression and restrict R8 photoreceptor cell fate. The R8 precursors express rhomboid (rho), which is required for the release of active EGFR ligand to activate EGFR signaling in surrounding cells for the subsequent stepwise recruitment. However, it is not clear about the mechanisms of transcriptional regulation of rho and how the lateral inhibition of Notch signaling and rho expression are coordinated. In this study, we show that inactivation of Groucho (Gro), an evolutionally conserved transcriptional corepressor, inhibits Ato upregulation, delays R8 determination, and promotes differentiation of R2-5 type of neurons. We demonstrate that these phenotypes are caused by a combination of the loss of Notch-mediated lateral inhibition and the precocious activation of EGFR signaling due to deregulated rho expression. Blocking EGFR signaling by Pnt-RNAi in conjunction with Gro-inactivation leads to lateral inhibition defects with deregulated Ato expression and R8 differentiation. We further show that inactivation of E(spl), which are the Gro binding transcription factors, causes deregulated rho expression and extra R8 cells within and posterior to the morphogenetic furrow (MF), and that E(spl) mediates the binding of Gro to the regulatory regions of both rho and ato genes in eye disc cells. Our results suggest that Gro inhibits rho expression in undifferentiated cells and represses the expression of both ato and rho in non-R8 precursors during initiation of photoreceptor differentiation in an E(spl)-dependent manner. The latter function of Gro provides novel insights into the mechanism that coordinates R8 specification with the restriction of initial rho expression to developing R8 cells.
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Affiliation(s)
- Tianyi Zhang
- Ben May Department for Cancer Research, The University of Chicago, 929 E. 57th Street, Chicago, IL 60637, USA
| | - Wei Du
- Ben May Department for Cancer Research, The University of Chicago, 929 E. 57th Street, Chicago, IL 60637, USA.
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38
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Hoffmann A, Spengler D. Role of ZAC1 in transient neonatal diabetes mellitus and glucose metabolism. World J Biol Chem 2015; 6:95-109. [PMID: 26322169 PMCID: PMC4549774 DOI: 10.4331/wjbc.v6.i3.95] [Citation(s) in RCA: 11] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 04/20/2015] [Revised: 06/19/2015] [Accepted: 07/11/2015] [Indexed: 02/05/2023] Open
Abstract
Transient neonatal diabetes mellitus 1 (TNDM1) is a rare genetic disorder representing with severe neonatal hyperglycaemia followed by remission within one and a half year and adolescent relapse with type 2 diabetes in half of the patients. Genetic defects in TNDM1 comprise uniparental isodisomy of chromosome 6, duplication of the minimal TNDM1 locus at 6q24, or relaxation of genomically imprinted ZAC1/HYMAI. Whereas the function of HYMAI, a non-coding mRNA, is still unidentified, biochemical and molecular studies show that zinc finger protein 1 regulating apoptosis and cell cycle arrest (ZAC1) behaves as a factor with versatile transcriptional functions dependent on binding to specific GC-rich DNA motives and interconnected regulation of recruited coactivator activities. Genome-wide expression profiling enabled the isolation of a number of Zac1 target genes known to regulate different aspects of β-cell function and peripheral insulin sensitivity. Among these, upregulation of Pparγ and Tcf4 impairs insulin-secretion and β-cell proliferation. Similarly, Zac1-mediated upregulation of Socs3 may attenuate β-cell proliferation and survival by inhibition of growth factor signalling. Additionally, Zac1 directly represses Pac1 and Rasgrf1 with roles in insulin secretion and β-cell proliferation. Collectively, concerted dysregulation of these target genes could contribute to the onset and course of TNDM1. Interestingly, Zac1 overexpression in β-cells spares the effects of stimulatory G-protein signaling on insulin secretion and raises the prospect for tailored treatments in relapsed TNDM1 patients. Overall, these results suggest that progress on the molecular and cellular foundations of monogenetic forms of diabetes can advance personalized therapy in addition to deepening the understanding of insulin and glucose metabolism in general.
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39
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Jahan I, Pan N, Elliott KL, Fritzsch B. The quest for restoring hearing: Understanding ear development more completely. Bioessays 2015. [PMID: 26208302 DOI: 10.1002/bies.201500044] [Citation(s) in RCA: 36] [Impact Index Per Article: 3.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/20/2022]
Abstract
Neurosensory hearing loss is a growing problem of super-aged societies. Cochlear implants can restore some hearing, but rebuilding a lost hearing organ would be superior. Research has discovered many cellular and molecular steps to develop a hearing organ but translating those insights into hearing organ restoration remains unclear. For example, we cannot make various hair cell types and arrange them into their specific patterns surrounded by the right type of supporting cells in the right numbers. Our overview of the topologically highly organized and functionally diversified cellular mosaic of the mammalian hearing organ highlights what is known and unknown about its development. Following this analysis, we suggest critical steps to guide future attempts toward restoration of a functional organ of Corti. We argue that generating mutant mouse lines that mimic human pathology to fine-tune attempts toward long-term functional restoration are needed to go beyond the hope generated by restoring single hair cells in postnatal sensory epithelia.
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Affiliation(s)
- Israt Jahan
- Department of Biology, CLAS, University of Iowa, Iowa City, IA, USA
| | - Ning Pan
- Department of Biology, CLAS, University of Iowa, Iowa City, IA, USA
| | - Karen L Elliott
- Department of Biology, CLAS, University of Iowa, Iowa City, IA, USA
| | - Bernd Fritzsch
- Department of Biology, CLAS, University of Iowa, Iowa City, IA, USA
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40
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Hoffmann A, Daniel G, Schmidt-Edelkraut U, Spengler D. Roles of imprinted genes in neural stem cells. Epigenomics 2015; 6:515-32. [PMID: 25431944 DOI: 10.2217/epi.14.42] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/17/2022] Open
Abstract
Imprinted genes and neural stem cells (NSC) play an important role in the developing and mature brain. A central theme of imprinted gene function in NSCs is cell survival and G1 arrest to control cell division, cell-cycle exit, migration and differentiation. Moreover, genomic imprinting can be epigenetically switched off at some genes to ensure stem cell quiescence and differentiation. At the genome scale, imprinted genes are organized in dynamic networks formed by interchromosomal interactions and transcriptional coregulation of imprinted and nonimprinted genes. Such multilayered networks may synchronize NSC activity with the demand from the niche resembling their roles in adjusting fetal size.
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Affiliation(s)
- Anke Hoffmann
- Max Planck Institute of Psychiatry, Translational Research, Kraepelinstrasse 2-10, 80804 Munich, Germany
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41
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Analysis of tandem E-box motifs within human Complement receptor 2 (CR2/CD21) promoter reveals cell specific roles for RP58, E2A, USF and localized chromatin accessibility. Int J Biochem Cell Biol 2015; 64:107-19. [DOI: 10.1016/j.biocel.2015.03.016] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/28/2015] [Revised: 03/03/2015] [Accepted: 03/18/2015] [Indexed: 02/06/2023]
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42
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Spratford CM, Kumar JP. Inhibition of Daughterless by Extramacrochaetae mediates Notch-induced cell proliferation. Development 2015; 142:2058-68. [PMID: 25977368 DOI: 10.1242/dev.121855] [Citation(s) in RCA: 8] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/07/2015] [Accepted: 04/16/2015] [Indexed: 12/30/2022]
Abstract
During development, the rate of cell proliferation must be constantly monitored so that an individual tissue achieves its correct size. Mutations in genes that normally promote tissue growth often result in undersized, disorganized and non-functional organs. However, mutations in genes that encode growth inhibitors can trigger the onset of tumorigenesis and cancer. The developing eye of the fruit fly, Drosophila melanogaster, has become a premier model system for studies that are focused on identifying the molecular mechanisms that underpin growth control. Here, we examine the mechanism by which the Notch pathway, a major contributor to growth, promotes cell proliferation in the developing eye. Current models propose that the Notch pathway directly influences cell proliferation by regulating growth-promoting genes such as four-jointed, cyclin D1 and E2f1. Here, we show that, in addition to these mechanisms, some Notch signaling is devoted to blocking the growth-suppressing activity of the bHLH DNA-binding protein Daughterless (Da). We demonstrate that Notch signaling activates the expression of extramacrochaetae (emc), which encodes a helix-loop-helix (HLH) transcription factor. Emc, in turn, then forms a biochemical complex with Da. As Emc lacks a basic DNA-binding domain, the Emc-Da heterodimer cannot bind to and regulate genomic targets. One effect of Da sequestration is to relieve the repression on growth. Here, we present data supporting our model that Notch-induced cell proliferation in the developing eye is mediated in part by the activity of Emc.
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Affiliation(s)
- Carrie M Spratford
- Department of Biology, Indiana University, Bloomington, IN 47405, USA Department of Molecular, Cell and Developmental Biology, University of California, Los Angeles, CA 90095, USA
| | - Justin P Kumar
- Department of Biology, Indiana University, Bloomington, IN 47405, USA
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43
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Abstract
Balancing cell growth with differentiation is essential for tissue integrity. In this issue of Developmental Cell, Wang and Baker (2015) demonstrate unsuspected cross-talk between bHLH transcription factors, important regulators of organogenesis, with the Hippo tumor suppressor pathway to ensure that inappropriately differentiating cells are eliminated during development.
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Affiliation(s)
- Tiffany A Cook
- Division of Developmental Biology, Cincinnati Children's Hospital Medical Center, Cincinnati, OH 45229, USA.
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44
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Daniel G, Schmidt-Edelkraut U, Spengler D, Hoffmann A. Imprinted Zac1 in neural stem cells. World J Stem Cells 2015; 7:300-314. [PMID: 25815116 PMCID: PMC4369488 DOI: 10.4252/wjsc.v7.i2.300] [Citation(s) in RCA: 6] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 07/25/2014] [Revised: 09/24/2014] [Accepted: 11/19/2014] [Indexed: 02/06/2023] Open
Abstract
Neural stem cells (NSCs) and imprinted genes play an important role in brain development. On historical grounds, these two determinants have been largely studied independently of each other. Recent evidence suggests, however, that NSCs can reset select genomic imprints to prevent precocious depletion of the stem cell reservoir. Moreover, imprinted genes like the transcriptional regulator Zac1 can fine tune neuronal vs astroglial differentiation of NSCs. Zac1 binds in a sequence-specific manner to pro-neuronal and imprinted genes to confer transcriptional regulation and furthermore coregulates members of the p53-family in NSCs. At the genome scale, Zac1 is a central hub of an imprinted gene network comprising genes with an important role for NSC quiescence, proliferation and differentiation. Overall, transcriptional, epigenomic, and genomic mechanisms seem to coordinate the functional relationships of NSCs and imprinted genes from development to maturation, and possibly aging.
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45
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Kiparaki M, Zarifi I, Delidakis C. bHLH proteins involved in Drosophila neurogenesis are mutually regulated at the level of stability. Nucleic Acids Res 2015; 43:2543-59. [PMID: 25694512 PMCID: PMC4357701 DOI: 10.1093/nar/gkv083] [Citation(s) in RCA: 16] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/13/2022] Open
Abstract
Proneural bHLH activators are expressed in all neuroectodermal regions prefiguring events of central and peripheral neurogenesis. Drosophila Sc is a prototypical proneural activator that heterodimerizes with the E-protein Daughterless (Da) and is antagonized by, among others, the E(spl) repressors. We determined parameters that regulate Sc stability in Drosophila S2 cells. We found that Sc is a very labile phosphoprotein and its turnover takes place via at least three proteasome-dependent mechanisms. (i) When Sc is in excess of Da, its degradation is promoted via its transactivation domain (TAD). (ii) In a DNA-bound Da/Sc heterodimer, Sc degradation is promoted via an SPTSS phosphorylation motif and the AD1 TAD of Da; Da is spared in the process. (iii) When E(spl)m7 is expressed, it complexes with Sc or Da/Sc and promotes their degradation in a manner that requires the corepressor Groucho and the Sc SPTSS motif. Da/Sc reciprocally promotes E(spl)m7 degradation. Since E(spl)m7 is a direct target of Notch, the mutual destabilization of Sc and E(spl) may contribute in part to the highly conserved anti-neural activity of Notch. Sc variants lacking the SPTSS motif are dramatically stabilized and are hyperactive in transgenic flies. Our results propose a novel mechanism of regulation of neurogenesis, involving the stability of key players in the process.
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Affiliation(s)
- Marianthi Kiparaki
- Institute of Molecular Biology and Biotechnology, Foundation for Research and Technology Hellas, and Department of Biology, University of Crete, 70013 Heraklion, Crete, Greece
| | - Ioanna Zarifi
- Institute of Molecular Biology and Biotechnology, Foundation for Research and Technology Hellas, and Department of Biology, University of Crete, 70013 Heraklion, Crete, Greece
| | - Christos Delidakis
- Institute of Molecular Biology and Biotechnology, Foundation for Research and Technology Hellas, and Department of Biology, University of Crete, 70013 Heraklion, Crete, Greece
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46
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Chen YS, Aubee J, DiVito KA, Zhou H, Zhang W, Chou FP, Simbulan-Rosenthal CM, Rosenthal DS. Id3 induces an Elk-1-caspase-8-dependent apoptotic pathway in squamous carcinoma cells. Cancer Med 2015; 4:914-24. [PMID: 25693514 PMCID: PMC4472214 DOI: 10.1002/cam4.427] [Citation(s) in RCA: 10] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/11/2014] [Revised: 12/28/2014] [Accepted: 01/10/2015] [Indexed: 11/29/2022] Open
Abstract
Inhibitor of differentiation/DNA-binding (Id) proteins are helix–loop–helix (HLH) transcription factors. The Id protein family (Id1–Id4) mediates tissue homeostasis by regulating cellular processes including differentiation, proliferation, and apoptosis. Ids typically function as dominant negative HLH proteins, which bind other HLH proteins and sequester them away from DNA promoter regions. Previously, we have found that Id3 induced apoptosis in immortalized human keratinocytes upon UVB exposure, consistent with its role as a tumor suppressor. To investigate the role of Id3 in malignant squamous cell carcinoma (SCC) cells (A431), a tetracycline-regulated inducible system was used to induce Id3 in cell culture and mouse xenograft models. We found that upon Id3 induction, there was a decrease in cell number under low serum conditions, as well as in soft agar. Microarray, RT-PCR, immunoblot, siRNA, and inhibitor studies revealed that Id3 induced expression of Elk-1, an E-twenty-six (ETS)-domain transcription factor, inducing procaspase-8 expression and activation. Id3 deletion mutants revealed that 80 C-terminal amino acids, including the HLH, are important for Id3-induced apoptosis. In a mouse xenograft model, Id3 induction decreased tumor size by 30%. Using immunofluorescent analysis, we determined that the tumor size decrease was also mediated through apoptosis. Furthermore, we show that Id3 synergizes with 5-FU and cisplatin therapies for nonmelanoma skin cancer cells. Our studies have shown a molecular mechanism by which Id3 induces apoptosis in SCC, and this information can potentially be used to develop new treatments for SCC patients.
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Affiliation(s)
- You-Shin Chen
- Department of Biochemistry and Molecular & Cellular Biology, Georgetown University, Washington, District of Columbia, 20057
| | - Joseph Aubee
- Department of Biochemistry and Molecular & Cellular Biology, Georgetown University, Washington, District of Columbia, 20057
| | - Kyle A DiVito
- Department of Biochemistry and Molecular & Cellular Biology, Georgetown University, Washington, District of Columbia, 20057
| | - Hengbo Zhou
- Department of Biochemistry and Molecular & Cellular Biology, Georgetown University, Washington, District of Columbia, 20057
| | - Weiyi Zhang
- Department of Biochemistry and Molecular & Cellular Biology, Georgetown University, Washington, District of Columbia, 20057
| | - Fen-Pi Chou
- Institute of Biochemistry and Biotechnology, Chung Shan Medical University, Taichung, 402, Taiwan
| | - Cynthia M Simbulan-Rosenthal
- Department of Biochemistry and Molecular & Cellular Biology, Georgetown University, Washington, District of Columbia, 20057
| | - Dean S Rosenthal
- Department of Biochemistry and Molecular & Cellular Biology, Georgetown University, Washington, District of Columbia, 20057.,Lombardi Comprehensive Cancer Center, Georgetown University, Washington, District of Columbia, 20057
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47
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Troost T, Schneider M, Klein T. A re-examination of the selection of the sensory organ precursor of the bristle sensilla of Drosophila melanogaster. PLoS Genet 2015; 11:e1004911. [PMID: 25569355 PMCID: PMC4287480 DOI: 10.1371/journal.pgen.1004911] [Citation(s) in RCA: 25] [Impact Index Per Article: 2.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/27/2014] [Accepted: 11/20/2014] [Indexed: 12/31/2022] Open
Abstract
The bristle sensillum of the imago of Drosophila is made of four cells that arise from a sensory organ precursor cell (SOP). This SOP is selected within proneural clusters (PNC) through a mechanism that involves Notch signalling. PNCs are defined through the expression domains of the proneural genes, whose activities enables cells to become SOPs. They encode tissue specific bHLH proteins that form functional heterodimers with the bHLH protein Daughterless (Da). In the prevailing lateral inhibition model for SOP selection, a transcriptional feedback loop that involves the Notch pathway amplifies small differences of proneural activity between cells of the PNC. As a result only one or two cells accumulate sufficient proneural activity to adopt the SOP fate. Most of the experiments that sustained the prevailing lateral inhibition model were performed a decade ago. We here re-examined the selection process using recently available reagents. Our data suggest a different picture of SOP selection. They indicate that a band-like region of proneural activity exists. In this proneural band the activity of the Notch pathway is required in combination with Emc to define the PNCs. We found a sub-group in the PNCs from which a pre-selected SOP arises. Our data indicate that most imaginal disc cells are able to adopt a proneural state from which they can progress to become SOPs. They further show that bristle formation can occur in the absence of the proneural genes if the function of emc is abolished. These results suggest that the tissue specific proneural proteins of Drosophila have a similar function as in the vertebrates, which is to determine the time of emergence and position of the SOP and to stabilise the proneural state. The sensory organ precursor cell (SOP) that forms the mechanosensory bristles of the adult PNS of Drosophila is a paradigm to study neural precursor determination. The current model states that the SOP is selected in proneural clusters (PNCs) defined through the expression of the proneural genes. The selection occurs through lateral inhibition mediated by the Notch signalling pathway. The SOP is pre-selected by differential expression of Extramacrochaetae (Emc), the only member of the Id proteins in Drosophila, which inactivates the proneural factors. We have re-examined the selection process using novel markers and mutants. Our data suggest a different picture of SOP selection. We discovered a band–like region of varying proneural activity where the peaks constitute the proneural clusters. Within the PNC, a subgroup exists from which the SOP arises. The Notch pathway has two distinct functions in the subgroup and in the rest of the band. We show that so far one unappreciated essential role of the proneural genes is the neutralisation of the activity of Emc. Our data suggest that the selection of the SOP is more similar to neural selection in vertebrates than previously anticipated.
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Affiliation(s)
- Tobias Troost
- Institut fuer Genetik, Heinrich-Heine-Universitaet Duesseldorf, Duesseldorf, Germany
| | - Markus Schneider
- Institut fuer Genetik, Heinrich-Heine-Universitaet Duesseldorf, Duesseldorf, Germany
| | - Thomas Klein
- Institut fuer Genetik, Heinrich-Heine-Universitaet Duesseldorf, Duesseldorf, Germany
- * E-mail:
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48
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Wang LH, Baker NE. Salvador-Warts-Hippo pathway in a developmental checkpoint monitoring helix-loop-helix proteins. Dev Cell 2015; 32:191-202. [PMID: 25579975 DOI: 10.1016/j.devcel.2014.12.002] [Citation(s) in RCA: 14] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/15/2013] [Revised: 10/16/2014] [Accepted: 12/01/2014] [Indexed: 12/19/2022]
Abstract
The E proteins and Id proteins are, respectively, the positive and negative heterodimer partners for the basic-helix-loop-helix protein family and as such contribute to a remarkably large number of cell-fate decisions. E proteins and Id proteins also function to inhibit or promote cell proliferation and cancer. Using a genetic modifier screen in Drosophila, we show that the Id protein Extramacrochaetae enables growth by suppressing activation of the Salvador-Warts-Hippo pathway of tumor suppressors, activation that requires transcriptional activation of the expanded gene by the E protein Daughterless. Daughterless protein binds to an intronic enhancer in the expanded gene, both activating the SWH pathway independently of the transmembrane protein Crumbs and bypassing the negative feedback regulation that targets the same expanded enhancer. Thus, the Salvador-Warts-Hippo pathway has a cell-autonomous function to prevent inappropriate differentiation due to transcription factor imbalance and monitors the intrinsic developmental status of progenitor cells, distinct from any responses to cell-cell interactions.
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Affiliation(s)
- Lan-Hsin Wang
- Department of Genetics, Albert Einstein College of Medicine, 1300 Morris Park Avenue, Bronx, NY 10461, USA
| | - Nicholas E Baker
- Department of Genetics, Albert Einstein College of Medicine, 1300 Morris Park Avenue, Bronx, NY 10461, USA; Department of Developmental and Molecular Biology, Albert Einstein College of Medicine, 1300 Morris Park Avenue, Bronx, NY 10461, USA; Department of Ophthalmology and Visual Sciences, Albert Einstein College of Medicine, 1300 Morris Park Avenue, Bronx, NY 10461, USA.
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49
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Grubišić V, Parpura V. Diversity in the utilization of glucose and lactate in synthetic mammalian myotubes generated by engineered configurations of MyoD and E12 in otherwise non-differentiation growth conditions. Biomaterials 2014; 43:50-60. [PMID: 25591961 DOI: 10.1016/j.biomaterials.2014.12.002] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [Key Words] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/18/2014] [Accepted: 12/03/2014] [Indexed: 12/21/2022]
Abstract
We previously used the expression of various combinations and configurations of MyoD and E12, two basic helix-loop-helix transcription factors (TF), to produce populations of myotubes assuming distinct morphology, myofibrillar development and Ca2+ dynamics, from mammalian C2C12 myoblasts in non-differentiation growth conditions. Here, we assessed the synthetically generated myotubes in terms of energetics, otherwise necessary to sustain their mechanical output as bio-actuators. We found that the myotubes exhibit changed expression of key regulators for the uptake and utilization of two major cellular fuels, glucose and lactate. Furthermore, while lactate transport was uniformly slowed in all the populations of myotubes, glucose uptake and utilization were modified by particular TF configuration. Our approach allows the production of a class of biomaterials with predetermined energetics that could be applied in biorobotics, where fuel of choice could be used, and also in reparative medicine where, for example, particular population of myotubes could be additionally employed as glucose sinks to mitigate effects of secondary metabolic syndrome.
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Affiliation(s)
- Vladimir Grubišić
- Department of Neurobiology, University of Alabama at Birmingham, Birmingham, AL 35294, USA
| | - Vladimir Parpura
- Department of Neurobiology, University of Alabama at Birmingham, Birmingham, AL 35294, USA; Department of Biotechnology, University of Rijeka, 51000 Rijeka, Croatia.
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Fischer B, Azim K, Hurtado-Chong A, Ramelli S, Fernández M, Raineteau O. E-proteins orchestrate the progression of neural stem cell differentiation in the postnatal forebrain. Neural Dev 2014; 9:23. [PMID: 25352248 PMCID: PMC4274746 DOI: 10.1186/1749-8104-9-23] [Citation(s) in RCA: 34] [Impact Index Per Article: 3.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/16/2014] [Accepted: 10/08/2014] [Indexed: 11/10/2022] Open
Abstract
BACKGROUND Neural stem cell (NSC) differentiation is a complex multistep process that persists in specific regions of the postnatal forebrain and requires tight regulation throughout life. The transcriptional control of NSC proliferation and specification involves Class II (proneural) and Class V (Id1-4) basic helix-loop-helix (bHLH) proteins. In this study, we analyzed the pattern of expression of their dimerization partners, Class I bHLH proteins (E-proteins), and explored their putative role in orchestrating postnatal subventricular zone (SVZ) neurogenesis. RESULTS Overexpression of a dominant-negative form of the E-protein E47 (dnE47) confirmed a crucial role for bHLH transcriptional networks in postnatal neurogenesis by dramatically blocking SVZ NSC differentiation. In situ hybridization was used in combination with RT-qPCR to measure and compare the level of expression of E-protein transcripts (E2-2, E2A, and HEB) in the neonatal and adult SVZ as well as in magnetic affinity cell sorted progenitor cells and neuroblasts. Our results evidence that E-protein transcripts, in particular E2-2 and E2A, are enriched in the postnatal SVZ with expression levels increasing as cells engage towards neuronal differentiation. To investigate the role of E-proteins in orchestrating lineage progression, both in vitro and in vivo gain-of-function and loss-of-function experiments were performed for individual E-proteins. Overexpression of E2-2 and E2A promoted SVZ neurogenesis by enhancing not only radial glial cell differentiation but also cell cycle exit of their progeny. Conversely, knock-down by shRNA electroporation resulted in opposite effects. Manipulation of E-proteins and/or Ascl1 in SVZ NSC cultures indicated that those effects were Ascl1 dependent, although they could not solely be attributed to an Ascl1-induced switch from promoting cell proliferation to triggering cell cycle arrest and differentiation. CONCLUSIONS In contrast to former concepts, suggesting ubiquitous expression and subsidiary function for E-proteins to foster postnatal neurogenesis, this work unveils E-proteins as being active players in the orchestration of postnatal SVZ neurogenesis.
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Affiliation(s)
| | | | | | | | | | - Olivier Raineteau
- Brain Research Institute, ETH Zurich/University of Zurich, 8057 Zurich, Switzerland.
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