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Zhang X, Liu F, Fang Q, Sun C, Fan J. E3 ligase HERC5-catalyzed UGDH isgylation promotes SNAI1-mediated tumor metastasis and cisplatin resistance in oral squamous cell carcinoma. Biol Direct 2025; 20:27. [PMID: 40045362 PMCID: PMC11883920 DOI: 10.1186/s13062-025-00622-1] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/13/2024] [Accepted: 02/22/2025] [Indexed: 03/09/2025] Open
Abstract
BACKGROUND Oral squamous cell carcinoma (OSCC) is one of the leading causes of cancer-related mortality worldwide due to its high aggressive potential and drug resistance. Previous studies have revealed an important function of HECT And RLD Domain Containing E3 Ubiquitin Protein Ligase 5 (HERC5) in cancer. Six GEO gene microarrays identified HERC5 as a significant upregulated gene in OSCC tissues or cells (log2 Fold change > 1 and adj.p < 0.05). This study aimed to explore the role and underlying mechanisms of HERC5 in OSCC development. RESULTS High HERC5 expression in OSCC tissues was confirmed by our hospital validation cohort and positively correlated with primary tumor stages. Subsequent functional studies demonstrated that knockdown of HERC5 inhibited the migratory and invasive capabilities with decrease of Vimentin and increase of E-cadherin in OSCC cells. In cisplatin treatment, cell survival rates were significantly reduced in HERC5-silencing OSCC cells, accompanied by the increase in cytotoxicity, DNA damage and apoptosis. OSCC cell-derived tumor xenograft displayed that HERC5 depletion inhibited pulmonary metastasis as well as restored the cisplatin-induced tumor burden. In line with this, overexpression of HERC5 yielded the opposite alterations both in vivo and in vitro. Mechanistically, UDP-glucose 6-dehydrogenase (UGDH) was identified as a HERC5-binding protein. Cysteine residue at position 994 in the HECT domain of HERC5 catalyzed the conjugation of ubiquitin-like protein Interferon-induced 15 kDa protein (ISG15) to UGDH (ISGylation of UGDH) and facilitated its phosphorylation, therefore enhancing SNAI1 mRNA stability. SNAI1 depletion inhibited HERC5 overexpression-triggered invasion and cisplatin resistance of OSCC cells. CONCLUSIONS Our study indicates that HERC5 may be a promising therapeutic target for OSCC.
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Affiliation(s)
- Xu Zhang
- Department of Head Neck and Thyroid, The Affiliated Cancer Hospital of Zhengzhou University & Henan Cancer Hospital, No. 127, Dongming Road, Zhengzhou, China
| | - Fayu Liu
- Department of Oromaxillofacial-Head and Neck Surgery, School and Hospital of Stomatology, Liaoning Province Key Laboratory of Oral Disease, China Medical University, Shenyang, China
| | - Qigen Fang
- Department of Head Neck and Thyroid, The Affiliated Cancer Hospital of Zhengzhou University & Henan Cancer Hospital, No. 127, Dongming Road, Zhengzhou, China
| | - Changfu Sun
- Department of Oromaxillofacial-Head and Neck Surgery, School and Hospital of Stomatology, Liaoning Province Key Laboratory of Oral Disease, China Medical University, Shenyang, China
| | - Jie Fan
- Department of Head Neck and Thyroid, The Affiliated Cancer Hospital of Zhengzhou University & Henan Cancer Hospital, No. 127, Dongming Road, Zhengzhou, China.
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Song W, Geng S, Qi Q, Lu X. Ugd Is Involved in the Synthesis of Glycans of Glycoprotein and LPS and Is Important for Cellulose Degradation in Cytophaga hutchinsonii. Microorganisms 2025; 13:395. [PMID: 40005761 PMCID: PMC11858162 DOI: 10.3390/microorganisms13020395] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/13/2025] [Revised: 02/06/2025] [Accepted: 02/09/2025] [Indexed: 02/27/2025] Open
Abstract
Cytophaga hutchinsonii, a member of the phylum Bacteroidetes, can rapidly degrade crystalline cellulose through direct cell-to-substrate contact. Most of its cellulases are secreted by the Type IX secretion system (T9SS) and anchored to the cell surface. Our previous study proved that the C-terminal domain (CTD) of the T9SS substrate cellulase Cel9A is glycosylated in C. hutchinsonii. However, its glycosylation mechanism has remained elusive. In this study, we found that chu_3394, which encodes UDP-glucose 6-dehydrogenase (Ugd), was important for the glycosylation of large amounts of periplasmic and outer membrane proteins in C. hutchinsonii. The contents of mannose, glucose, galactose, and xylose were detected to be reduced in the glycoproteins of the ∆ugd mutant compared to that of wild-type. They might be essential monosaccharides that contribute to the structure and function of glycans attached to proteins in C. hutchinsonii. The depletion of mannose, glucose, galactose, and xylose indicates a decrease in glycosylation modifications in the ∆ugd mutant strain. Then, we found that the deletion of ugd resulted in weakened glycosylation modification of the recombinant green fluorescent protein-tagged CTD of Cel9A. Additionally, the outer-membrane localization of Cel9A was affected in the mutant. Besides this, Ugd was also important for the synthesis of O-antigen of lipopolysaccharide (LPS). Thus, Ugd was involved in the synthesis of glycans in both glycoproteins and LPS in C. hutchinsonii. Moreover, the deletion of ugd affected the cellulose degradation, cell motility, and stress resistance of C. hutchinsonii.
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Affiliation(s)
| | | | | | - Xuemei Lu
- State Key Laboratory of Microbial Technology, Shandong University, Qingdao 266237, China; (W.S.); (S.G.); (Q.Q.)
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Klure DM, Greenhalgh R, Orr TJ, Shapiro MD, Dearing MD. Parallel gene expansions drive rapid dietary adaptation in herbivorous woodrats. Science 2025; 387:156-162. [PMID: 39787210 DOI: 10.1126/science.adp7978] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/11/2024] [Accepted: 11/15/2024] [Indexed: 01/12/2025]
Abstract
How mammalian herbivores evolve to feed on chemically defended plants remains poorly understood. In this study, we investigated the adaptation of two species of woodrats (Neotoma lepida and N. bryanti) to creosote bush (Larrea tridentata), a toxic shrub that expanded across the southwestern United States after the Last Glacial Maximum. We found that creosote-adapted woodrats have elevated gene dosage across multiple biotransformation enzyme families. These duplication events occurred independently across species and substantially increase expression of biotransformation genes, especially within the glucuronidation pathway. We propose that increased gene dosage resulting from duplication is an important mechanism by which animals initially adapt to novel environmental pressures.
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Affiliation(s)
- Dylan M Klure
- School of Biological Sciences, University of Utah, Salt Lake City, UT, USA
| | - Robert Greenhalgh
- School of Biological Sciences, University of Utah, Salt Lake City, UT, USA
| | - Teri J Orr
- School of Biological Sciences, University of Utah, Salt Lake City, UT, USA
- Department of Biology, New Mexico State University, Las Cruces, NM, USA
| | - Michael D Shapiro
- School of Biological Sciences, University of Utah, Salt Lake City, UT, USA
| | - M Denise Dearing
- School of Biological Sciences, University of Utah, Salt Lake City, UT, USA
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Jung I, Nam S, Lee DY, Park SY, Yu JH, Seo JA, Lee DH, Kim NH. Association of Succinate and Adenosine Nucleotide Metabolic Pathways with Diabetic Kidney Disease in Patients with Type 2 Diabetes Mellitus. Diabetes Metab J 2024; 48:1126-1134. [PMID: 38945526 PMCID: PMC11621657 DOI: 10.4093/dmj.2023.0377] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 10/23/2023] [Accepted: 05/06/2024] [Indexed: 07/02/2024] Open
Abstract
BACKGRUOUND Although the prevalence of diabetic kidney disease (DKD) is increasing, reliable biomarkers for its early detection are scarce. This study aimed to evaluate the association of adenosine and succinate levels and their related pathways, including hyaluronic acid (HA) synthesis, with DKD. METHODS We examined 235 participants and categorized them into three groups: healthy controls; those with diabetes but without DKD; and those with DKD, which was defined as estimated glomerular filtration rate (eGFR) <60 mL/min/1.73 m2. We compared the concentrations of urinary adenosine, succinate, and HA and the serum levels of cluster of differentiation 39 (CD39) and CD73, which are involved in adenosine generation, among the groups with DKD or albuminuria. In addition, we performed multiple logistic regression analysis to evaluate the independent association of DKD or albuminuria with the metabolites after adjusting for risk factors. We also showed the association of these metabolites with eGFR measured several years before enrollment. This study was registered with the Clinical Research Information Service (https://cris.nih.go.kr; Registration number: KCT0003573). RESULTS Urinary succinate and serum CD39 levels were higher in the DKD group than in the control and non-DKD groups. Correlation analysis consistently linked urinary succinate and serum CD39 concentrations with eGFR, albuminuria, and ΔeGFR, which was calculated retrospectively. However, among the various metabolites studied, only urinary succinate was identified as an independent indicator of DKD and albuminuria. CONCLUSION Among several potential metabolites, only urinary succinate was independently associated with DKD. These findings hold promise for clinical application in the management of DKD.
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Affiliation(s)
- Inha Jung
- Division of Endocrinology and Metabolism, Department of Internal Medicine, Korea University College of Medicine, Seoul, Korea
| | - Seungyoon Nam
- Department of Genome Medicine and Science, AI Convergence Center for Medical Science, Gachon Institute of Genome Medicine and Science, Gachon University Gil Medical Center, Gachon University College of Medicine, Incheon, Korea
- Department of Health Sciences and Technology, Gachon Advanced Institute for Health Sciences and Technology, Gachon University, Incheon, Korea
| | - Da Young Lee
- Division of Endocrinology and Metabolism, Department of Internal Medicine, Korea University College of Medicine, Seoul, Korea
| | - So Young Park
- Division of Endocrinology and Metabolism, Department of Internal Medicine, Korea University College of Medicine, Seoul, Korea
| | - Ji Hee Yu
- Division of Endocrinology and Metabolism, Department of Internal Medicine, Korea University College of Medicine, Seoul, Korea
| | - Ji A Seo
- Division of Endocrinology and Metabolism, Department of Internal Medicine, Korea University College of Medicine, Seoul, Korea
| | - Dae Ho Lee
- Department of Health Sciences and Technology, Gachon Advanced Institute for Health Sciences and Technology, Gachon University, Incheon, Korea
- Department of Internal Medicine, Gachon University Gil Medical Center, Gachon University College of Medicine, Incheon, Korea
| | - Nan Hee Kim
- Division of Endocrinology and Metabolism, Department of Internal Medicine, Korea University College of Medicine, Seoul, Korea
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Vitale DL, Parnigoni A, Viola M, Karousou E, Sevic I, Moretto P, Passi A, Alaniz L, Vigetti D. Deciphering Drug Resistance: Investigating the Emerging Role of Hyaluronan Metabolism and Signaling and Tumor Extracellular Matrix in Cancer Chemotherapy. Int J Mol Sci 2024; 25:7607. [PMID: 39062846 PMCID: PMC11276752 DOI: 10.3390/ijms25147607] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/07/2024] [Revised: 07/04/2024] [Accepted: 07/09/2024] [Indexed: 07/28/2024] Open
Abstract
Hyaluronan (HA) has gained significant attention in cancer research for its role in modulating chemoresistance. This review aims to elucidate the mechanisms by which HA contributes to chemoresistance, focusing on its interactions within the tumor microenvironment. HA is abundantly present in the extracellular matrix (ECM) and binds to cell-surface receptors such as CD44 and RHAMM. These interactions activate various signaling pathways, including PI3K/Akt, MAPK, and NF-κB, which are implicated in cell survival, proliferation, and drug resistance. HA also influences the physical properties of the tumor stroma, enhancing its density and reducing drug penetration. Additionally, HA-mediated signaling contributes to the epithelial-mesenchymal transition (EMT), a process associated with increased metastatic potential and resistance to apoptosis. Emerging therapeutic strategies aim to counteract HA-induced chemoresistance by targeting HA synthesis, degradation, metabolism, or its binding to CD44. This review underscores the complexity of HA's role in chemoresistance and highlights the potential for HA-targeted therapies to improve the efficacy of conventional chemotherapeutics.
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Affiliation(s)
- Daiana L. Vitale
- Laboratorio de Microambiente Tumoral, Centro de Investigaciones Básicas y Aplicadas (CIBA), Universidad Nacional del Noroeste de la Provincia de Buenos Aires, Junín B6000, Argentina; (D.L.V.); (I.S.); (L.A.)
- Centro de Investigaciones y Transferencia del Noroeste de la Provincia de Buenos Aires (CITNOBA), UNNOBA-UNSAdA-CONICET, Junín 6000, Argentina
| | - Arianna Parnigoni
- Department of Medical Biochemistry and Microbiology, Uppsala University, SE-751 23 Uppsala, Sweden;
| | - Manuela Viola
- Dipartimento di Medicina e Chirurgia, Universitá degli Studi dell’Insubria, 21100 Varese, Italy; (M.V.); (E.K.); (P.M.); (A.P.)
| | - Evgenia Karousou
- Dipartimento di Medicina e Chirurgia, Universitá degli Studi dell’Insubria, 21100 Varese, Italy; (M.V.); (E.K.); (P.M.); (A.P.)
| | - Ina Sevic
- Laboratorio de Microambiente Tumoral, Centro de Investigaciones Básicas y Aplicadas (CIBA), Universidad Nacional del Noroeste de la Provincia de Buenos Aires, Junín B6000, Argentina; (D.L.V.); (I.S.); (L.A.)
- Centro de Investigaciones y Transferencia del Noroeste de la Provincia de Buenos Aires (CITNOBA), UNNOBA-UNSAdA-CONICET, Junín 6000, Argentina
| | - Paola Moretto
- Dipartimento di Medicina e Chirurgia, Universitá degli Studi dell’Insubria, 21100 Varese, Italy; (M.V.); (E.K.); (P.M.); (A.P.)
| | - Alberto Passi
- Dipartimento di Medicina e Chirurgia, Universitá degli Studi dell’Insubria, 21100 Varese, Italy; (M.V.); (E.K.); (P.M.); (A.P.)
| | - Laura Alaniz
- Laboratorio de Microambiente Tumoral, Centro de Investigaciones Básicas y Aplicadas (CIBA), Universidad Nacional del Noroeste de la Provincia de Buenos Aires, Junín B6000, Argentina; (D.L.V.); (I.S.); (L.A.)
- Centro de Investigaciones y Transferencia del Noroeste de la Provincia de Buenos Aires (CITNOBA), UNNOBA-UNSAdA-CONICET, Junín 6000, Argentina
| | - Davide Vigetti
- Dipartimento di Medicina e Chirurgia, Universitá degli Studi dell’Insubria, 21100 Varese, Italy; (M.V.); (E.K.); (P.M.); (A.P.)
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Greenhalgh R, Klure DM, Orr TJ, Armstrong NM, Shapiro MD, Dearing MD. The desert woodrat (Neotoma lepida) induces a diversity of biotransformation genes in response to creosote bush resin. Comp Biochem Physiol C Toxicol Pharmacol 2024; 280:109870. [PMID: 38428625 PMCID: PMC11006593 DOI: 10.1016/j.cbpc.2024.109870] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 08/10/2023] [Revised: 01/26/2024] [Accepted: 02/24/2024] [Indexed: 03/03/2024]
Abstract
Liver biotransformation enzymes have long been thought to enable animals to feed on diets rich in xenobiotic compounds. However, despite decades of pharmacological research in humans and rodents, little is known about hepatic gene expression in specialized mammalian herbivores feeding on toxic diets. Leveraging a recently identified population of the desert woodrat (Neotoma lepida) found to be highly tolerant to toxic creosote bush (Larrea tridentata), we explored the expression changes of suites of biotransformation genes in response to diets enriched with varying amounts of creosote resin. Analysis of hepatic RNA-seq data indicated a dose-dependent response to these compounds, including the upregulation of several genes encoding transcription factors and numerous phase I, II, and III biotransformation families. Notably, elevated expression of five biotransformation families - carboxylesterases, cytochromes P450, aldo-keto reductases, epoxide hydrolases, and UDP-glucuronosyltransferases - corresponded to species-specific duplication events in the genome, suggesting that these genes play a prominent role in N. lepida's adaptation to creosote bush. Building on pharmaceutical studies in model rodents, we propose a hypothesis for how the differentially expressed genes are involved in the biotransformation of creosote xenobiotics. Our results provide some of the first details about how these processes likely operate in the liver of a specialized mammalian herbivore.
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Affiliation(s)
- Robert Greenhalgh
- School of Biological Sciences, University of Utah, 257 S 1400 E, Salt Lake City, UT 84112, USA.
| | - Dylan M Klure
- School of Biological Sciences, University of Utah, 257 S 1400 E, Salt Lake City, UT 84112, USA.
| | - Teri J Orr
- School of Biological Sciences, University of Utah, 257 S 1400 E, Salt Lake City, UT 84112, USA.
| | - Noah M Armstrong
- School of Biological Sciences, University of Utah, 257 S 1400 E, Salt Lake City, UT 84112, USA.
| | - Michael D Shapiro
- School of Biological Sciences, University of Utah, 257 S 1400 E, Salt Lake City, UT 84112, USA.
| | - M Denise Dearing
- School of Biological Sciences, University of Utah, 257 S 1400 E, Salt Lake City, UT 84112, USA.
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7
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Wang M, Zeng J, Tan H, Guo Q, Li X, Ling X, Zhang J, Song S, Deng Y. Anti-virulence and bactericidal activities of Stattic against Shigella sonnei. Appl Environ Microbiol 2023; 89:e0107423. [PMID: 38032177 PMCID: PMC10734500 DOI: 10.1128/aem.01074-23] [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: 06/24/2023] [Accepted: 10/13/2023] [Indexed: 12/01/2023] Open
Abstract
IMPORTANCE Shigella sonnei is a major human enteric pathogen that causes bacillary dysentery. The increasing spread of drug-resistant S. sonnei strains has caused an emergent need for the development of new antimicrobial agents against this pathogenic bacterium. In this study, we demonstrate that Stattic employs two antibacterial mechanisms against S. sonnei. It exerted both anti-virulence activity and bactericidal activity against S. sonnei, suggesting that it shows advantages over traditional antibiotics. Moreover, Stattic showed excellent synergistic effects with kanamycin, ampicillin, chloramphenicol, and gentamicin against S. sonnei. Our findings suggest that Stattic has promising potential for development as a new antibiotic or as an adjuvant to antibiotics for infections caused by S. sonnei.
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Affiliation(s)
- Mingfang Wang
- School of Pharmaceutical Sciences (Shenzhen), Shenzhen Campus of Sun Yat-sen University, Sun Yat-sen University, Shenzhen, China
| | - Jia Zeng
- School of Pharmaceutical Sciences (Shenzhen), Shenzhen Campus of Sun Yat-sen University, Sun Yat-sen University, Shenzhen, China
| | - Huihui Tan
- School of Pharmaceutical Sciences (Shenzhen), Shenzhen Campus of Sun Yat-sen University, Sun Yat-sen University, Shenzhen, China
| | - Quan Guo
- School of Pharmaceutical Sciences (Shenzhen), Shenzhen Campus of Sun Yat-sen University, Sun Yat-sen University, Shenzhen, China
| | - Xia Li
- School of Pharmaceutical Sciences (Shenzhen), Shenzhen Campus of Sun Yat-sen University, Sun Yat-sen University, Shenzhen, China
| | - Xiwen Ling
- School of Pharmaceutical Sciences (Shenzhen), Shenzhen Campus of Sun Yat-sen University, Sun Yat-sen University, Shenzhen, China
| | - Jinyue Zhang
- School of Pharmaceutical Sciences, Hainan University, Haikou, China
| | - Shihao Song
- School of Pharmaceutical Sciences (Shenzhen), Shenzhen Campus of Sun Yat-sen University, Sun Yat-sen University, Shenzhen, China
- School of Pharmaceutical Sciences, Hainan University, Haikou, China
| | - Yinyue Deng
- School of Pharmaceutical Sciences (Shenzhen), Shenzhen Campus of Sun Yat-sen University, Sun Yat-sen University, Shenzhen, China
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8
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Price MJ, Nguyen AD, Byemerwa JK, Flowers J, Baëta CD, Goodwin CR. UDP-glucose dehydrogenase (UGDH) in clinical oncology and cancer biology. Oncotarget 2023; 14:843-857. [PMID: 37769033 PMCID: PMC10538703 DOI: 10.18632/oncotarget.28514] [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: 08/02/2023] [Accepted: 09/04/2023] [Indexed: 09/30/2023] Open
Abstract
UDP-glucose-6-dehydrogenase (UGDH) is a cytosolic, hexameric enzyme that converts UDP-glucose to UDP-glucuronic acid (UDP-GlcUA), a key reaction in hormone and xenobiotic metabolism and in the production of extracellular matrix precursors. In this review, we classify UGDH as a molecular indicator of tumor progression in multiple cancer types, describe its involvement in key canonical cancer signaling pathways, and identify methods to inhibit UGDH, its substrates, and its downstream products. As such, we position UGDH as an enzyme to be exploited as a potential prognostication marker in oncology and a therapeutic target in cancer biology.
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Affiliation(s)
- Meghan J. Price
- Department of Internal Medicine, John Hopkins Hospital, Baltimore, MD 21287, USA
| | - Annee D. Nguyen
- Department of Neurosurgery, Duke University Medical Center, Durham, NC 27710, USA
| | - Jovita K. Byemerwa
- Department of Pharmacology and Cancer Biology, Duke University, Durham, NC 27708, USA
| | - Jasmine Flowers
- Department of Neurosurgery, Associated with Duke University Medical Center, Durham, NC 27710, USA
| | - César D. Baëta
- Department of Epidemiology and Clinical Research, Stanford University, Stanford, CA 94305, USA
| | - C. Rory Goodwin
- Department of Neurosurgery, Duke Center for Brain and Spine Metastasis and Duke Cancer Institute, Durham, NC 27710, USA
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9
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Warren A, Porter RM, Reyes-Castro O, Ali MM, Marques-Carvalho A, Kim HN, Gatrell LB, Schipani E, Nookaew I, O'Brien CA, Morello R, Almeida M. The NAD salvage pathway in mesenchymal cells is indispensable for skeletal development in mice. Nat Commun 2023; 14:3616. [PMID: 37330524 PMCID: PMC10276814 DOI: 10.1038/s41467-023-39392-7] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/13/2022] [Accepted: 06/09/2023] [Indexed: 06/19/2023] Open
Abstract
NAD is an essential co-factor for cellular energy metabolism and multiple other processes. Systemic NAD+ deficiency has been implicated in skeletal deformities during development in both humans and mice. NAD levels are maintained by multiple synthetic pathways but which ones are important in bone forming cells is unknown. Here, we generate mice with deletion of Nicotinamide Phosphoribosyltransferase (Nampt), a critical enzyme in the NAD salvage pathway, in all mesenchymal lineage cells of the limbs. At birth, NamptΔPrx1 exhibit dramatic limb shortening due to death of growth plate chondrocytes. Administration of the NAD precursor nicotinamide riboside during pregnancy prevents the majority of in utero defects. Depletion of NAD post-birth also promotes chondrocyte death, preventing further endochondral ossification and joint development. In contrast, osteoblast formation still occurs in knockout mice, in line with distinctly different microenvironments and reliance on redox reactions between chondrocytes and osteoblasts. These findings define a critical role for cell-autonomous NAD homeostasis during endochondral bone formation.
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Affiliation(s)
- Aaron Warren
- Division of Endocrinology and Metabolism, University of Arkansas for Medical Sciences, Little Rock, AR, USA
| | - Ryan M Porter
- Division of Endocrinology and Metabolism, University of Arkansas for Medical Sciences, Little Rock, AR, USA
- Center for Musculoskeletal Disease Research, University of Arkansas for Medical Sciences, Little Rock, AR, USA
- Department of Orthopedic Surgery, University of Arkansas for Medical Sciences, Little Rock, AR, USA
| | - Olivia Reyes-Castro
- Division of Endocrinology and Metabolism, University of Arkansas for Medical Sciences, Little Rock, AR, USA
| | - Md Mohsin Ali
- Division of Endocrinology and Metabolism, University of Arkansas for Medical Sciences, Little Rock, AR, USA
| | - Adriana Marques-Carvalho
- CNC-Center for Neuroscience and Cell Biology, University of Coimbra, UC-Biotech, Biocant Park, Cantanhede, Portugal
| | - Ha-Neui Kim
- Division of Endocrinology and Metabolism, University of Arkansas for Medical Sciences, Little Rock, AR, USA
- Center for Musculoskeletal Disease Research, University of Arkansas for Medical Sciences, Little Rock, AR, USA
| | - Landon B Gatrell
- Division of Endocrinology and Metabolism, University of Arkansas for Medical Sciences, Little Rock, AR, USA
| | - Ernestina Schipani
- Department of Orthopaedic Surgery, University of Pennsylvania, Philadelphia, PA, USA
| | - Intawat Nookaew
- Center for Musculoskeletal Disease Research, University of Arkansas for Medical Sciences, Little Rock, AR, USA
- Department of Biomedical Informatics, University of Arkansas for Medical Sciences, Little Rock, AR, USA
| | - Charles A O'Brien
- Division of Endocrinology and Metabolism, University of Arkansas for Medical Sciences, Little Rock, AR, USA
- Center for Musculoskeletal Disease Research, University of Arkansas for Medical Sciences, Little Rock, AR, USA
- Department of Orthopedic Surgery, University of Arkansas for Medical Sciences, Little Rock, AR, USA
| | - Roy Morello
- Center for Musculoskeletal Disease Research, University of Arkansas for Medical Sciences, Little Rock, AR, USA
- Department of Orthopedic Surgery, University of Arkansas for Medical Sciences, Little Rock, AR, USA
- Department of Physiology and Cell Biology, University of Arkansas for Medical Sciences, Little Rock, AR, USA
| | - Maria Almeida
- Division of Endocrinology and Metabolism, University of Arkansas for Medical Sciences, Little Rock, AR, USA.
- Center for Musculoskeletal Disease Research, University of Arkansas for Medical Sciences, Little Rock, AR, USA.
- Department of Orthopedic Surgery, University of Arkansas for Medical Sciences, Little Rock, AR, USA.
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10
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Liu Y, Li Y, Li G, Chu H. The molecular mechanism of Y473 phosphorylation of UGDH relieves the inhibition effect of UDP-glucose on HuR. Phys Chem Chem Phys 2023; 25:8714-8724. [PMID: 36896759 DOI: 10.1039/d3cp00227f] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 03/03/2023]
Abstract
Uridine diphosphate glucose (UDP-Glc) is able to accelerate the decay of snail family transcriptional repressor 1 (SNAI1) mRNA by inhibiting Hu antigen R (HuR, an RNA-binding protein), thereby preventing cancer invasiveness and drug resistance. Nevertheless, the phosphorylation of tyrosine 473 (Y473) of UDP-glucose dehydrogenase (UGDH is capable of converting UDP-Glc to uridine diphosphate glucuronic acid (UDP-GlcUA)) weakens the inhibition of UDP-Glc to HuR, thus initiating the epithelial-mesenchymal transformation of tumor cells and promoting tumor cell migration and metastasis. To address the mechanism, we performed molecular dynamics simulations combined with molecular mechanics generalized Born surface area (MM/GBSA) analysis on wild-type and Y473 phosphorylated UGDH and HuR, UDP-Glc, UDP-GlcUA complexes. We demonstrated that Y473 phosphorylation was able to enhance the binding between UGDH and the HuR/UDP-Glc complex. Compared with HuR, UGDH has a stronger binding ability with UDP-Glc; therefore, UDP-Glc was inclined to bind to UGDH and then was catalyzed to UDP-GlcUA by UGDH, which relieved the inhibition of UDP-Glc to HuR. In addition, the binding ability of HuR for UDP-GlcUA was lower than its affinity for UDP-Glc, significantly reducing the inhibition of HuR. Hence, HuR bound to SNAI1 mRNA more easily to increase the stability of mRNA. Our results revealed the micromolecular mechanism of Y473 phosphorylation of UGDH regulating the interaction between UGDH and HuR as well as relieving the inhibition of UDP-Glc on HuR, which contributed to understanding the role of UGDH and HuR in tumor metastasis and developing small molecule drugs targeting the interaction between UGDH and HuR.
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Affiliation(s)
- Ye Liu
- Laboratory of Molecular Modeling and Design, State Key Laboratory of Molecular Reaction Dynamics, Dalian Institute of Chemical Physics, Chinese Academy of Sciences, Dalian, 116023, China.
| | - Yan Li
- Laboratory of Molecular Modeling and Design, State Key Laboratory of Molecular Reaction Dynamics, Dalian Institute of Chemical Physics, Chinese Academy of Sciences, Dalian, 116023, China.
| | - Guohui Li
- Laboratory of Molecular Modeling and Design, State Key Laboratory of Molecular Reaction Dynamics, Dalian Institute of Chemical Physics, Chinese Academy of Sciences, Dalian, 116023, China.
| | - Huiying Chu
- Laboratory of Molecular Modeling and Design, State Key Laboratory of Molecular Reaction Dynamics, Dalian Institute of Chemical Physics, Chinese Academy of Sciences, Dalian, 116023, China.
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11
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Salzano A, Fioriniello S, D'Onofrio N, Balestrieri ML, Aiese Cigliano R, Neglia G, Della Ragione F, Campanile G. Transcriptomic profiles of the ruminal wall in Italian Mediterranean dairy buffaloes fed green forage. BMC Genomics 2023; 24:133. [PMID: 36941576 PMCID: PMC10029215 DOI: 10.1186/s12864-023-09215-6] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/07/2022] [Accepted: 02/28/2023] [Indexed: 03/23/2023] Open
Abstract
BACKGROUND Green feed diet in ruminants exerts a beneficial effect on rumen metabolism and enhances the content of milk nutraceutical quality. At present, a comprehensive analysis focused on the identification of genes, and therefore, biological processes modulated by the green feed in buffalo rumen has never been reported. We performed RNA-sequencing in the rumen of buffaloes fed a total mixed ration (TMR) + the inclusion of 30% of ryegrass green feed (treated) or TMR (control), and identified differentially expressed genes (DEGs) using EdgeR and NOISeq tools. RESULTS We found 155 DEGs using EdgeR (p-values < 0.05) and 61 DEGs using NOISeq (prob ≥0.8), 30 of which are shared. The rt-qPCR validation suggested a higher reliability of EdgeR results as compared with NOISeq data, in our biological context. Gene Ontology analysis of DEGs identified using EdgeR revealed that green feed modulates biological processes relevant for the rumen physiology and, then, health and well-being of buffaloes, such as lipid metabolism, response to the oxidative stress, immune response, and muscle structure and function. Accordingly, we found: (i) up-regulation of HSD17B13, LOC102410803 (or PSAT1) and HYKK, and down-regulation of CDO1, SELENBP1 and PEMT, encoding factors involved in energy, lipid and amino acid metabolism; (ii) enhanced expression of SIM2 and TRIM14, whose products are implicated in the immune response and defense against infections, and reduced expression of LOC112585166 (or SAAL1), ROR2, SMOC2, and S100A11, encoding pro-inflammatory factors; (iii) up-regulation of NUDT18, DNAJA4 and HSF4, whose products counteract stressful conditions, and down-regulation of LOC102396388 (or UGT1A9) and LOC102413340 (or MRP4/ABCC4), encoding detoxifying factors; (iv) increased expression of KCNK10, CACNG4, and ATP2B4, encoding proteins modulating Ca2+ homeostasis, and reduced expression of the cytoskeleton-related MYH11 and DES. CONCLUSION Although statistically unpowered, this study suggests that green feed modulates the expression of genes involved in biological processes relevant for rumen functionality and physiology, and thus, for welfare and quality production in Italian Mediterranean dairy buffaloes. These findings, that need to be further confirmed through the validation of additional DEGs, allow to speculate a role of green feed in the production of nutraceutical molecules, whose levels might be enhanced also in milk.
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Affiliation(s)
- Angela Salzano
- Department of Veterinary Medicine and Animal Production, Federico II University, Naples, Italy
| | | | - Nunzia D'Onofrio
- Department of Precision Medicine, University of Campania Luigi Vanvitelli, Naples, Italy
| | | | | | - Gianluca Neglia
- Department of Veterinary Medicine and Animal Production, Federico II University, Naples, Italy
| | - Floriana Della Ragione
- Institute of Genetics and Biophysics 'A. Buzzati-Traverso', CNR, Naples, Italy.
- IRCCS Istituto Neurologico Mediterraneo Neuromed, Pozzilli, Isernia, Italy.
| | - Giuseppe Campanile
- Department of Veterinary Medicine and Animal Production, Federico II University, Naples, Italy
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12
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Park S, Kim OH, Lee K, Park IB, Kim NH, Moon S, Im J, Sharma SP, Oh BC, Nam S, Lee DH. Plasma and urinary extracellular vesicle microRNAs and their related pathways in diabetic kidney disease. Genomics 2022; 114:110407. [PMID: 35716820 DOI: 10.1016/j.ygeno.2022.110407] [Citation(s) in RCA: 6] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/27/2022] [Revised: 04/22/2022] [Accepted: 06/10/2022] [Indexed: 11/30/2022]
Abstract
To explore extracellular vesicle microRNAs (EV miRNAs) and their target mRNAs in relation to diabetic kidney disease (DKD), we performed paired plasma and urinary EV small RNA sequencing (n = 18) in patients with type 2 diabetes and DKD (n = 5) and healthy subjects (n = 4) and metabolic network analyses using our own miRNA and public mRNA datasets. We found 13 common differentially expressed EV miRNAs in both fluids and 17 target mRNAs, including RRM2, NT5E, and UGDH. Because succinate dehydrogenase B was suggested to interact with proteins encoded by these three genes, we measured urinary succinate and adenosine in a validation study (n = 194). These two urinary metabolite concentrations were associated with DKD progression. In addition, renal expressions of NT5E and UGDH proteins were increased in db/db mice with DKD compared to control mice. In conclusion, we profiled DKD-related EV miRNAs in plasma and urine samples and found their relevant target pathways.
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Affiliation(s)
- Sungjin Park
- Department of Genome Medicine and Science, AI Convergence Center for Medical Science, Gachon Institute of Genome Medicine and Science, Gachon University Gil Medical Center, Gachon University College of Medicine, Incheon, Republic of Korea
| | - Ok-Hee Kim
- Department of Physiology, Lee Gil Ya Cancer and Diabetes Institute, College of Medicine, Gachon University, Incheon, Republic of Korea
| | - Kiyoung Lee
- Department of Internal Medicine, Gachon University Gil Medical Center, Incheon, Republic of Korea; Department of Internal Medicine, Gachon University College of Medicine, Incheon, Republic of Korea
| | - Ie Byung Park
- Department of Internal Medicine, Gachon University Gil Medical Center, Incheon, Republic of Korea; Department of Internal Medicine, Gachon University College of Medicine, Incheon, Republic of Korea
| | - Nan Hee Kim
- Division of Endocrinology and Metabolism, Department of Internal Medicine, Korea University College of Medicine, Seoul, Republic of Korea
| | - Seongryeol Moon
- Department of Health Sciences and Technology, Gachon Advanced Institute for Health Sciences and Technology (GAIHST), Gachon University, Incheon, Republic of Korea
| | - Jaebeen Im
- Department of Health Sciences and Technology, Gachon Advanced Institute for Health Sciences and Technology (GAIHST), Gachon University, Incheon, Republic of Korea
| | - Satya Priya Sharma
- Department of Internal Medicine, Gachon University Gil Medical Center, Incheon, Republic of Korea
| | - Byung-Chul Oh
- Department of Physiology, Lee Gil Ya Cancer and Diabetes Institute, College of Medicine, Gachon University, Incheon, Republic of Korea
| | - Seungyoon Nam
- Department of Genome Medicine and Science, AI Convergence Center for Medical Science, Gachon Institute of Genome Medicine and Science, Gachon University Gil Medical Center, Gachon University College of Medicine, Incheon, Republic of Korea; Department of Health Sciences and Technology, Gachon Advanced Institute for Health Sciences and Technology (GAIHST), Gachon University, Incheon, Republic of Korea.
| | - Dae Ho Lee
- Department of Internal Medicine, Gachon University Gil Medical Center, Incheon, Republic of Korea; Department of Internal Medicine, Gachon University College of Medicine, Incheon, Republic of Korea; Department of Health Sciences and Technology, Gachon Advanced Institute for Health Sciences and Technology (GAIHST), Gachon University, Incheon, Republic of Korea.
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13
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Zhan D, Yalcin F, Ma D, Fu Y, Wei S, Lal B, Li Y, Dzaye O, Laterra J, Ying M, Lopez-Bertoni H, Xia S. Targeting UDP-α-d-glucose 6-dehydrogenase alters the CNS tumor immune microenvironment and inhibits glioblastoma growth. Genes Dis 2022; 9:717-730. [PMID: 35782977 PMCID: PMC9243400 DOI: 10.1016/j.gendis.2021.08.008] [Citation(s) in RCA: 9] [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: 05/20/2021] [Revised: 08/14/2021] [Accepted: 08/26/2021] [Indexed: 12/15/2022] Open
Abstract
Glioblastoma (GBM, WHO grade IV glioma) is the most common and lethal malignant brain tumor in adults with a dismal prognosis. The extracellular matrix (ECM) supports GBM progression by promoting tumor cell proliferation, migration, and immune escape. Uridine diphosphate (UDP)-glucose 6-dehydrogenase (UGDH) is the rate-limiting enzyme that catalyzes the biosynthesis of glycosaminoglycans that are the principal component of the CNS ECM. We investigated how targeting UGDH in GBM influences the GBM immune microenvironment, including tumor-associated microglia/macrophages (TAMs) and T cells. TAMs are the main immune effector cells in GBM and can directly target tumor cells if properly activated. In co-cultures of GBM cells and human primary macrophages, UGDH knockdown in GBM cells promoted macrophage phagocytosis and M1-like polarization. In orthotropic human GBM xenografts and syngeneic mouse glioma models, targeting UGDH decreased ECM deposition, increased TAM phagocytosis marker expression, reduced M2-like TAMs and inhibited tumor growth. UGDH knockdown in GBM cells also promoted cytotoxic T cell infiltration and activation in orthotopic syngeneic mouse glioma models. The potent and in-human-use small molecule GAG synthesis inhibitor 4-methylumbelliferone (4-MU) was found to inhibit GBM cell proliferation and migration in vitro, mimic the macrophage and T-cell responses to UGDH knockdown in vitro and in vivo and inhibit growth of orthotopic murine GBM. Our study shows that UGDH supports GBM growth through multiple mechanisms and supports the development of ECM-based therapeutic strategies to simultaneously target tumor cells and their microenvironment.
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Affiliation(s)
- Daqian Zhan
- Neurology, Hugo W. Moser Research Institute at Kennedy Krieger, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA
- Department of Critical Care Medicine, Tongji Hospital, Huazhong University of Science and Technology, Wuhan, Hubei 430074, PR China
| | - Fatih Yalcin
- Department of Radiology and Neuroradiology, Charité, Berlin 10117, Germany
- Russell H. Morgan Department of Radiology and Radiological Science, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA
| | - Ding Ma
- Neurology, Hugo W. Moser Research Institute at Kennedy Krieger, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA
- Department of Respiratory and Critical Care Medicine, Tongji Hospital, Huazhong University of Science and Technology, Wuhan, Hubei 430074, PR China
| | - Yi Fu
- Neurology, Hugo W. Moser Research Institute at Kennedy Krieger, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA
| | - Shuang Wei
- University Hospital Center Schleswig Holstein, Department of Neurosurgery, Kiel, Schleswig-Holstein 24105, Germany
| | - Bachchu Lal
- Neurology, Hugo W. Moser Research Institute at Kennedy Krieger, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA
| | - Yunqing Li
- Neurology, Hugo W. Moser Research Institute at Kennedy Krieger, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA
| | - Omar Dzaye
- Department of Radiology and Neuroradiology, Charité, Berlin 10117, Germany
- Departments of Neurology, Oncology and Neuroscience, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA
| | - John Laterra
- Neurology, Hugo W. Moser Research Institute at Kennedy Krieger, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA
- Blood and Cell Therapy Institute, University of Science and Technology of China, Anhui Provincial Hospital, Hefei, Anhui 230026, PR China
| | - Mingyao Ying
- Neurology, Hugo W. Moser Research Institute at Kennedy Krieger, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA
| | - Hernando Lopez-Bertoni
- Neurology, Hugo W. Moser Research Institute at Kennedy Krieger, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA
| | - Shuli Xia
- Neurology, Hugo W. Moser Research Institute at Kennedy Krieger, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA
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14
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Zimmer BM, Barycki JJ, Simpson MA. Mechanisms of coordinating hyaluronan and glycosaminoglycan production by nucleotide sugars. Am J Physiol Cell Physiol 2022; 322:C1201-C1213. [PMID: 35442826 DOI: 10.1152/ajpcell.00130.2022] [Citation(s) in RCA: 3] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/22/2022]
Abstract
Hyaluronan is a versatile macromolecule capable of an exceptional range of functions from cushioning and hydration to dynamic signaling in development and disease. Because of its critical roles, hyaluronan production is regulated at multiple levels including epigenetic, transcriptional, and post-translational control of the three hyaluronan synthase (HAS) enzymes. Precursor availability can dictate the rate and amount of hyaluronan synthesized and shed by the cells producing it. However, the nucleotide-activated sugar substrates for hyaluronan synthesis by HAS also participate in exquisitely fine tuned cross talking pathways that intersect with central carbohydrate metabolism. Multiple UDP-sugars have alternative metabolic fates and exhibit coordinated and reciprocal allosteric control of enzymes within their biosynthetic pathways to preserve appropriate precursor ratios for accurate partitioning among downstream products, while also sensing and maintaining energy homeostasis. Since the dysregulation of nucleotide sugar and hyaluronan synthesis is associated with multiple pathologies, these pathways offer opportunities for therapeutic intervention. Recent structures of several key rate-limiting enzymes in the UDP-sugar synthesis pathways have offered new insights to the overall regulation of hyaluronan production by precursor fate decisions. The details of UDP-sugar control and the structural basis for underlying mechanisms are discussed in this review.
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Affiliation(s)
- Brenna M Zimmer
- Department of Molecular and Structural Biochemistry, North Carolina State University, Raleigh, NC, United States
| | - Joseph J Barycki
- Department of Molecular and Structural Biochemistry, North Carolina State University, Raleigh, NC, United States
| | - Melanie A Simpson
- Department of Molecular and Structural Biochemistry, North Carolina State University, Raleigh, NC, United States
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15
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Wang W, Cui J, Ma H, Lu W, Huang J. Targeting Pyrimidine Metabolism in the Era of Precision Cancer Medicine. Front Oncol 2021; 11:684961. [PMID: 34123854 PMCID: PMC8194085 DOI: 10.3389/fonc.2021.684961] [Citation(s) in RCA: 70] [Impact Index Per Article: 17.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/24/2021] [Accepted: 04/27/2021] [Indexed: 12/26/2022] Open
Abstract
Metabolic rewiring is considered as a primary feature of cancer. Malignant cells reprogram metabolism pathway in response to various intrinsic and extrinsic drawback to fuel cell survival and growth. Among the complex metabolic pathways, pyrimidine biosynthesis is conserved in all living organism and is necessary to maintain cellular fundamental function (i.e. DNA and RNA biosynthesis). A wealth of evidence has demonstrated that dysfunction of pyrimidine metabolism is closely related to cancer progression and numerous drugs targeting pyrimidine metabolism have been approved for multiple types of cancer. However, the non-negligible side effects and limited efficacy warrants a better strategy for negating pyrimidine metabolism in cancer. In recent years, increased studies have evidenced the interplay of oncogenic signaling and pyrimidine synthesis in tumorigenesis. Here, we review the recent conceptual advances on pyrimidine metabolism, especially dihydroorotate dehydrogenase (DHODH), in the framework of precision oncology medicine and prospect how this would guide the development of new drug precisely targeting the pyrimidine metabolism in cancer.
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Affiliation(s)
- Wanyan Wang
- State Key Laboratory of Bioreactor Engineering, Shanghai Key Laboratory of New Drug Design, School of Pharmacy, East China University of Science and Technology, Shanghai, China
| | - Jiayan Cui
- State Key Laboratory of Bioreactor Engineering, Shanghai Key Laboratory of New Drug Design, School of Pharmacy, East China University of Science and Technology, Shanghai, China
| | - Hui Ma
- State Key Laboratory of Bioreactor Engineering, Shanghai Key Laboratory of New Drug Design, School of Pharmacy, East China University of Science and Technology, Shanghai, China
| | - Weiqiang Lu
- Shanghai Key Laboratory of Regulatory Biology, Institute of Biomedical Sciences and School of Life Sciences, East China Normal University, Shanghai, China
| | - Jin Huang
- State Key Laboratory of Bioreactor Engineering, Shanghai Key Laboratory of New Drug Design, School of Pharmacy, East China University of Science and Technology, Shanghai, China
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16
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Zhang S, Cao L, Sun X, Yu J, Xu X, Chang R, Suo J, Liu G, Xu Z, Qu C. Genome-wide analysis of UGDH genes in Populus trichocarpa and responsiveness to nitrogen treatment. 3 Biotech 2021; 11:149. [PMID: 33732570 DOI: 10.1007/s13205-021-02697-9] [Citation(s) in RCA: 4] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/27/2020] [Accepted: 02/16/2021] [Indexed: 12/20/2022] Open
Abstract
Plant UDP-glucose 6-dehydrogenase (UGDH) is an important enzyme for the formation of hemicellulose and pectin. Previous studies on UGDH have primarily focused on the biosynthesis of cell wall polysaccharides, while few studies have focused on their regulation by exogenous nitrogen. In the present study, four genes encoding PtUGDH proteins were analyzed by bioinformatics methods. And, the expression profiles of PtUGDH genes under different nitrogen treatments were evaluated with qRT-PCR. The results showed that PtUGDHs have conserved NAD coenzyme binding motif GAGYVGG and the catalytic motif GFGGSCFQKDIL. According to the phylogenetic analysis, PtUGDHs were divided into two subgroups. PtUGDH3 and PtUGDH4 were closely related to AtUGDH1 (important for normal development of Arabidopsis cell wall structure). Chromosomal distribution and genome synteny analysis revealed four segmental-duplicated gene pairs on chr4, 8, 10 and 17. Tissue-specific data from PlantGenIE showed that PtUGDH3 and PtUGDH4 were highly expressed in stems. The qRT-PCR detection showed that the expression of PtUGDH3 in the lower stem and PtUGDH2 of upper leaves were significantly increased induced by low ammonium or nitrate condition. This comprehensive analysis of the UGDH family in poplar provides new insights into their regulation by nitrogen, and would increase our understanding of the roles of UGDHs in hemicellulose and pectin biosynthesis in the cell wall and during poplar development. SUPPLEMENTARY INFORMATION The online version contains supplementary material available at 10.1007/s13205-021-02697-9.
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17
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Vitale DL, Caon I, Parnigoni A, Sevic I, Spinelli FM, Icardi A, Passi A, Vigetti D, Alaniz L. Initial Identification of UDP-Glucose Dehydrogenase as a Prognostic Marker in Breast Cancer Patients, Which Facilitates Epirubicin Resistance and Regulates Hyaluronan Synthesis in MDA-MB-231 Cells. Biomolecules 2021; 11:biom11020246. [PMID: 33572239 PMCID: PMC7914570 DOI: 10.3390/biom11020246] [Citation(s) in RCA: 14] [Impact Index Per Article: 3.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/29/2020] [Revised: 02/01/2021] [Accepted: 02/02/2021] [Indexed: 12/11/2022] Open
Abstract
UDP-glucose-dehydrogenase (UGDH) synthesizes UDP-glucuronic acid. It is involved in epirubicin detoxification and hyaluronan synthesis. This work aimed to evaluate the effect of UGDH knockdown on epirubicin response and hyaluronan metabolism in MDA-MB-231 breast cancer cells. Additionally, the aim was to determine UGDH as a possible prognosis marker in breast cancer. We studied UGDH expression in tumors and adjacent tissue from breast cancer patients. The prognostic value of UGDH was studied using a public Kaplan–Meier plotter. MDA-MB-231 cells were knocked-down for UGDH and treated with epirubicin. Epirubicin-accumulation and apoptosis were analyzed by flow cytometry. Hyaluronan-coated matrix and metabolism were determined. Autophagic-LC3-II was studied by Western blot and confocal microscopy. Epirubicin accumulation increased and apoptosis decreased during UGDH knockdown. Hyaluronan-coated matrix increased and a positive modulation of autophagy was detected. Higher levels of UGDH were correlated with worse prognosis in triple-negative breast cancer patients that received chemotherapy. High expression of UGDH was found in tumoral tissue from HER2--patients. However, UGDH knockdown contributes to epirubicin resistance, which might be associated with increases in the expression, deposition and catabolism of hyaluronan. The results obtained allowed us to propose UGDH as a new prognostic marker in breast cancer, positively associated with development of epirubicin resistance and modulation of extracellular matrix.
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Affiliation(s)
- Daiana L. Vitale
- Laboratorio de Microambiente Tumoral, Centro de Investigaciones Básicas y Aplicadas (CIBA), Universidad Nacional del Noroeste de la Provincia de Buenos Aires, Junín 6000, Argentina; (D.L.V.); (I.S.); (F.M.S.); (A.I.)
- Centro de Investigaciones y Transferencia del Noroeste de la Provincia de Buenos Aires (CITNOBA), UNNOBA-UNSAdA-CONICET, Junín 6000, Argentina
| | - Ilaria Caon
- Dipartimento di Medicina e Chirurgia, Università degli Studio dell’Insubria, 21100 Varese, Italy; (I.C.); (A.P.); (A.P.)
| | - Arianna Parnigoni
- Dipartimento di Medicina e Chirurgia, Università degli Studio dell’Insubria, 21100 Varese, Italy; (I.C.); (A.P.); (A.P.)
| | - Ina Sevic
- Laboratorio de Microambiente Tumoral, Centro de Investigaciones Básicas y Aplicadas (CIBA), Universidad Nacional del Noroeste de la Provincia de Buenos Aires, Junín 6000, Argentina; (D.L.V.); (I.S.); (F.M.S.); (A.I.)
- Centro de Investigaciones y Transferencia del Noroeste de la Provincia de Buenos Aires (CITNOBA), UNNOBA-UNSAdA-CONICET, Junín 6000, Argentina
| | - Fiorella M. Spinelli
- Laboratorio de Microambiente Tumoral, Centro de Investigaciones Básicas y Aplicadas (CIBA), Universidad Nacional del Noroeste de la Provincia de Buenos Aires, Junín 6000, Argentina; (D.L.V.); (I.S.); (F.M.S.); (A.I.)
- Centro de Investigaciones y Transferencia del Noroeste de la Provincia de Buenos Aires (CITNOBA), UNNOBA-UNSAdA-CONICET, Junín 6000, Argentina
| | - Antonella Icardi
- Laboratorio de Microambiente Tumoral, Centro de Investigaciones Básicas y Aplicadas (CIBA), Universidad Nacional del Noroeste de la Provincia de Buenos Aires, Junín 6000, Argentina; (D.L.V.); (I.S.); (F.M.S.); (A.I.)
- Centro de Investigaciones y Transferencia del Noroeste de la Provincia de Buenos Aires (CITNOBA), UNNOBA-UNSAdA-CONICET, Junín 6000, Argentina
| | - Alberto Passi
- Dipartimento di Medicina e Chirurgia, Università degli Studio dell’Insubria, 21100 Varese, Italy; (I.C.); (A.P.); (A.P.)
| | - Davide Vigetti
- Dipartimento di Medicina e Chirurgia, Università degli Studio dell’Insubria, 21100 Varese, Italy; (I.C.); (A.P.); (A.P.)
- Correspondence: (D.V.); (L.A.); Tel.: + 39-332-307170 (D.V.); +54-236-4-407750 (ext. 11625) (L.A.)
| | - Laura Alaniz
- Laboratorio de Microambiente Tumoral, Centro de Investigaciones Básicas y Aplicadas (CIBA), Universidad Nacional del Noroeste de la Provincia de Buenos Aires, Junín 6000, Argentina; (D.L.V.); (I.S.); (F.M.S.); (A.I.)
- Centro de Investigaciones y Transferencia del Noroeste de la Provincia de Buenos Aires (CITNOBA), UNNOBA-UNSAdA-CONICET, Junín 6000, Argentina
- Correspondence: (D.V.); (L.A.); Tel.: + 39-332-307170 (D.V.); +54-236-4-407750 (ext. 11625) (L.A.)
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Tenascin-C Function in Glioma: Immunomodulation and Beyond. ADVANCES IN EXPERIMENTAL MEDICINE AND BIOLOGY 2020; 1272:149-172. [PMID: 32845507 DOI: 10.1007/978-3-030-48457-6_9] [Citation(s) in RCA: 24] [Impact Index Per Article: 4.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 12/12/2022]
Abstract
First identified in the 1980s, tenascin-C (TNC) is a multi-domain extracellular matrix glycoprotein abundantly expressed during the development of multicellular organisms. TNC level is undetectable in most adult tissues but rapidly and transiently induced by a handful of pro-inflammatory cytokines in a variety of pathological conditions including infection, inflammation, fibrosis, and wound healing. Persistent TNC expression is associated with chronic inflammation and many malignancies, including glioma. By interacting with its receptor integrin and a myriad of other binding partners, TNC elicits context- and cell type-dependent function to regulate cell adhesion, migration, proliferation, and angiogenesis. TNC operates as an endogenous activator of toll-like receptor 4 and promotes inflammatory response by inducing the expression of multiple pro-inflammatory factors in innate immune cells such as microglia and macrophages. In addition, TNC drives macrophage differentiation and polarization predominantly towards an M1-like phenotype. In contrast, TNC shows immunosuppressive function in T cells. In glioma, TNC is expressed by tumor cells and stromal cells; high expression of TNC is correlated with tumor progression and poor prognosis. Besides promoting glioma invasion and angiogenesis, TNC has been found to affect the morphology and function of tumor-associated microglia/macrophages in glioma. Clinically, TNC can serve as a biomarker for tumor progression; and TNC antibodies have been utilized as an adjuvant agent to deliver anti-tumor drugs to target glioma. A better mechanistic understanding of how TNC impacts innate and adaptive immunity during tumorigenesis and tumor progression will open new therapeutic avenues to treat brain tumors and other malignancies.
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Zimmer BM, Barycki JJ, Simpson MA. Integration of Sugar Metabolism and Proteoglycan Synthesis by UDP-glucose Dehydrogenase. J Histochem Cytochem 2020; 69:13-23. [PMID: 32749901 DOI: 10.1369/0022155420947500] [Citation(s) in RCA: 21] [Impact Index Per Article: 4.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/08/2023] Open
Abstract
Regulation of proteoglycan and glycosaminoglycan synthesis is critical throughout development, and to maintain normal adult functions in wound healing and the immune system, among others. It has become increasingly clear that these processes are also under tight metabolic control and that availability of carbohydrate and amino acid metabolite precursors has a role in the control of proteoglycan and glycosaminoglycan turnover. The enzyme uridine diphosphate (UDP)-glucose dehydrogenase (UGDH) produces UDP-glucuronate, an essential precursor for new glycosaminoglycan synthesis that is tightly controlled at multiple levels. Here, we review the cellular mechanisms that regulate UGDH expression, discuss the structural features of the enzyme, and use the structures to provide a context for recent studies that link post-translational modifications and allosteric modulators of UGDH to its function in downstream pathways.
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Affiliation(s)
- Brenna M Zimmer
- Department of Molecular and Structural Biochemistry, North Carolina State University, Raleigh, North Carolina
| | - Joseph J Barycki
- Department of Molecular and Structural Biochemistry, North Carolina State University, Raleigh, North Carolina
| | - Melanie A Simpson
- Department of Molecular and Structural Biochemistry, North Carolina State University, Raleigh, North Carolina
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20
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Rocha B, Cillero-Pastor B, Eijkel G, Calamia V, Fernandez-Puente P, Paine MRL, Ruiz-Romero C, Heeren RMA, Blanco FJ. Integrative Metabolic Pathway Analysis Reveals Novel Therapeutic Targets in Osteoarthritis. Mol Cell Proteomics 2020; 19:574-588. [PMID: 31980557 PMCID: PMC7124476 DOI: 10.1074/mcp.ra119.001821] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/11/2019] [Revised: 01/08/2020] [Indexed: 01/15/2023] Open
Abstract
In osteoarthritis (OA), impairment of cartilage regeneration can be related to a defective chondrogenic differentiation of mesenchymal stromal cells (MSCs). Therefore, understanding the proteomic- and metabolomic-associated molecular events during the chondrogenesis of MSCs could provide alternative targets for therapeutic intervention. Here, a SILAC-based proteomic analysis identified 43 proteins related with metabolic pathways whose abundance was significantly altered during the chondrogenesis of OA human bone marrow MSCs (hBMSCs). Then, the level and distribution of metabolites was analyzed in these cells and healthy controls by matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI-MSI), leading to the recognition of characteristic metabolomic profiles at the early stages of differentiation. Finally, integrative pathway analysis showed that UDP-glucuronic acid synthesis and amino sugar metabolism were downregulated in OA hBMSCs during chondrogenesis compared with healthy cells. Alterations in these metabolic pathways may disturb the production of hyaluronic acid (HA) and other relevant cartilage extracellular matrix (ECM) components. This work provides a novel integrative insight into the molecular alterations of osteoarthritic MSCs and potential therapeutic targets for OA drug development through the enhancement of chondrogenesis.
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Affiliation(s)
- Beatriz Rocha
- Grupo de Investigación de Reumatología (GIR), Unidad de Proteómica, INIBIC - Complejo Hospitalario Universitario de A Coruña, SERGAS, Universidad de A Coruña, A Coruña, Spain
| | - Berta Cillero-Pastor
- The Maastricht Multimodal Molecular Imaging Institute (M4I), Division of Imaging Mass Spectrometry, Maastricht University, The Netherlands
| | - Gert Eijkel
- The Maastricht Multimodal Molecular Imaging Institute (M4I), Division of Imaging Mass Spectrometry, Maastricht University, The Netherlands
| | - Valentina Calamia
- Grupo de Investigación de Reumatología (GIR), Unidad de Proteómica, INIBIC - Complejo Hospitalario Universitario de A Coruña, SERGAS, Universidad de A Coruña, A Coruña, Spain.
| | - Patricia Fernandez-Puente
- Grupo de Investigación de Reumatología, INIBIC-Complejo Hospitalario Universitario de A Coruña, SERGAS, Agrupación CICA-INIBIC, Universidad de A Coruña, A Coruña, Spain
| | - Martin R L Paine
- The Maastricht Multimodal Molecular Imaging Institute (M4I), Division of Imaging Mass Spectrometry, Maastricht University, The Netherlands
| | - Cristina Ruiz-Romero
- Grupo de Investigación de Reumatología (GIR), Unidad de Proteómica, INIBIC - Complejo Hospitalario Universitario de A Coruña, SERGAS, Universidad de A Coruña, A Coruña, Spain
| | - Ron M A Heeren
- The Maastricht Multimodal Molecular Imaging Institute (M4I), Division of Imaging Mass Spectrometry, Maastricht University, The Netherlands
| | - Francisco J Blanco
- Grupo de Investigación de Reumatología, INIBIC-Complejo Hospitalario Universitario de A Coruña, SERGAS, Departamento de Medicina Universidad de A Coruña, A Coruña, Spain.
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21
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UDP-glucose 6-dehydrogenase expression as a predictor of survival in
patients with pulmonary adenocarcinoma. INTERNATIONAL JOURNAL OF SURGERY-ONCOLOGY 2020. [DOI: 10.1097/ij9.0000000000000085] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 12/25/2022]
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22
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Hengel H, Bosso-Lefèvre C, Grady G, Szenker-Ravi E, Li H, Pierce S, Lebigot É, Tan TT, Eio MY, Narayanan G, Utami KH, Yau M, Handal N, Deigendesch W, Keimer R, Marzouqa HM, Gunay-Aygun M, Muriello MJ, Verhelst H, Weckhuysen S, Mahida S, Naidu S, Thomas TG, Lim JY, Tan ES, Haye D, Willemsen MAAP, Oegema R, Mitchell WG, Pierson TM, Andrews MV, Willing MC, Rodan LH, Barakat TS, van Slegtenhorst M, Gavrilova RH, Martinelli D, Gilboa T, Tamim AM, Hashem MO, AlSayed MD, Abdulrahim MM, Al-Owain M, Awaji A, Mahmoud AAH, Faqeih EA, Asmari AA, Algain SM, Jad LA, Aldhalaan HM, Helbig I, Koolen DA, Riess A, Kraegeloh-Mann I, Bauer P, Gulsuner S, Stamberger H, Ng AYJ, Tang S, Tohari S, Keren B, Schultz-Rogers LE, Klee EW, Barresi S, Tartaglia M, Mor-Shaked H, Maddirevula S, Begtrup A, Telegrafi A, Pfundt R, Schüle R, Ciruna B, Bonnard C, Pouladi MA, Stewart JC, Claridge-Chang A, Lefeber DJ, Alkuraya FS, Mathuru AS, Venkatesh B, Barycki JJ, Simpson MA, Jamuar SS, Schöls L, Reversade B. Loss-of-function mutations in UDP-Glucose 6-Dehydrogenase cause recessive developmental epileptic encephalopathy. Nat Commun 2020; 11:595. [PMID: 32001716 PMCID: PMC6992768 DOI: 10.1038/s41467-020-14360-7] [Citation(s) in RCA: 33] [Impact Index Per Article: 6.6] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/20/2017] [Accepted: 12/19/2019] [Indexed: 12/16/2022] Open
Abstract
Developmental epileptic encephalopathies are devastating disorders characterized by intractable epileptic seizures and developmental delay. Here, we report an allelic series of germline recessive mutations in UGDH in 36 cases from 25 families presenting with epileptic encephalopathy with developmental delay and hypotonia. UGDH encodes an oxidoreductase that converts UDP-glucose to UDP-glucuronic acid, a key component of specific proteoglycans and glycolipids. Consistent with being loss-of-function alleles, we show using patients' primary fibroblasts and biochemical assays, that these mutations either impair UGDH stability, oligomerization, or enzymatic activity. In vitro, patient-derived cerebral organoids are smaller with a reduced number of proliferating neuronal progenitors while mutant ugdh zebrafish do not phenocopy the human disease. Our study defines UGDH as a key player for the production of extracellular matrix components that are essential for human brain development. Based on the incidence of variants observed, UGDH mutations are likely to be a frequent cause of recessive epileptic encephalopathy.
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Affiliation(s)
- Holger Hengel
- Department of Neurology and Hertie-Institute for Clinical Brain Research, University of Tübingen, Tübingen, Germany
- German Center of Neurodegenerative Diseases (DZNE), Tübingen, Germany
| | - Célia Bosso-Lefèvre
- Institute of Medical Biology, A*STAR, Biopolis, Singapore, 138648, Singapore
- National University of Singapore, Department of Paediatrics, Yong Loo Lin School of Medicine, Biopolis, Singapore, Singapore
| | - George Grady
- Department of Molecular and Structural Biochemistry North Carolina State University, Raleigh, NC, 27607, USA
| | | | - Hankun Li
- Yale-NUS College, 12 College Avenue West, Biopolis, Singapore, Singapore
| | - Sarah Pierce
- Division of Medical Genetics, Department of Medicine, University of Washington, Seattle, WA, USA
| | - Élise Lebigot
- Service De Biochimie, Hopital Bicêtre, Assistance publique-Hôpitaux de Paris, 78 avenue du general leclerc, Le Kremlin Bicêtre, France
| | - Thong-Teck Tan
- Institute of Medical Biology, Singapore Stem Cell Bank, A∗STAR, Biopolis, Singapore, 138648, Singapore
| | - Michelle Y Eio
- Institute of Medical Biology, Singapore Stem Cell Bank, A∗STAR, Biopolis, Singapore, 138648, Singapore
| | - Gunaseelan Narayanan
- Institute of Medical Biology, Singapore Stem Cell Bank, A∗STAR, Biopolis, Singapore, 138648, Singapore
| | - Kagistia Hana Utami
- Translational Laboratory in Genetic Medicine, Agency for Science, Technology, and Research, Singapore (A*STAR), 8A Biomedical Grove, Immunos, Level 5, Singapore, 138648, Singapore
| | - Monica Yau
- Program in Developmental & Stem Cell Biology, The Hospital for Sick Children, Department of Molecular Genetics, The University of Toronto, Toronto, ON, Canada
| | - Nader Handal
- Caritas Baby Hospital Bethlehem, Bethlehem, State of Palestine
| | | | - Reinhard Keimer
- Ped Neurology, Staufer Hospital, Wetzgauer Straße 85, Schwäbisch-Gmünd, Germany
| | | | - Meral Gunay-Aygun
- McKusick-Nathans Institute of Genetic Medicine, Johns Hopkins University School of Medicine, Baltimore, MD, 21205, USA
| | - Michael J Muriello
- McKusick-Nathans Institute of Genetic Medicine, Johns Hopkins University School of Medicine, Baltimore, MD, 21205, USA
| | - Helene Verhelst
- Department of Paediatric Neurology, Ghent University Hospital, Ghent, Belgium
| | - Sarah Weckhuysen
- Center for Molecular Neurology, VIB, Antwerp, Belgium
- Laboratory of Neurogenetics, Institute Born-Bunge, University of Antwerp, Antwerp, Belgium
- Department of Neurology, University Hospital Antwerp, Antwerp, Belgium
| | - Sonal Mahida
- Division of Neurology and Neurogenetics, Kennedy Krieger Institute, Baltimore, MD, USA
| | - Sakkubai Naidu
- Division of Neurology and Neurogenetics, Kennedy Krieger Institute, Baltimore, MD, USA
| | - Terrence G Thomas
- Neurology Service, Department of Paediatrics, KK Women's and Children's Hospital, Singapore, Singapore
| | - Jiin Ying Lim
- Genetics Service, Department of Paediatrics, KK Women's and Children's Hospital, Singapore, Singapore
- Paediatric Academic Clinical Programme, Duke-NUS Medical School, Singapore, Singapore
- SingHealth Duke-NUS Genomic Medicine Centre, Singapore, Singapore
| | - Ee Shien Tan
- Genetics Service, Department of Paediatrics, KK Women's and Children's Hospital, Singapore, Singapore
- Paediatric Academic Clinical Programme, Duke-NUS Medical School, Singapore, Singapore
- SingHealth Duke-NUS Genomic Medicine Centre, Singapore, Singapore
| | - Damien Haye
- Service de Génétique Médicale, CHU De Nice Hôpital de l'Archet 2, 151 route Saint Antoine de la Ginestière, CS 23079 062002, Nice, Cedex 3, France
| | - Michèl A A P Willemsen
- Department of Pediatric Neurology, Radboud University Medical Center, Nijmegen, The Netherlands
| | - Renske Oegema
- Department of Genetics, University Medical Center Utrecht, Utrecht, The Netherlands
| | - Wendy G Mitchell
- Neurology Division, Childrens Hospital Los Angeles & Department of Neurology, Keck School of Medicine of University of Southern California, Los Angeles, CA, 90033, USA
| | - Tyler Mark Pierson
- Department of Pediatrics, Department of Neurology, & the Board of Governors Regenerative Medicine Institute, Cedars-Sinai Medical Center, Los Angeles, CA, USA
| | - Marisa V Andrews
- Division of Genetics and Genomic Medicine, Department of Pediatrics, Washington University School of Medicine, St. Louis, MO, USA
| | - Marcia C Willing
- Division of Genetics and Genomic Medicine, Department of Pediatrics, Washington University School of Medicine, St. Louis, MO, USA
| | - Lance H Rodan
- Division of Genetics and Genomics and Department of Neurology, Boston Children's Hospital, Boston, MA, USA
| | - Tahsin Stefan Barakat
- Department of Clinical Genetics, Erasmus MC, University Medical Center, Wytemaweg 80, 3015 CN, Rotterdam, The Netherlands
| | - Marjon van Slegtenhorst
- Department of Clinical Genetics, Erasmus MC, University Medical Center, Wytemaweg 80, 3015 CN, Rotterdam, The Netherlands
| | - Ralitza H Gavrilova
- Department of Clinical Genomics, Mayo Clinic, 200 First Street SW, Rochester, MN, USA
| | - Diego Martinelli
- Genetics and Rare Diseases Research Division, Ospedale Pediatrico Bambino Gesù, IRCCS, viale San Paolo 15, 00146, Rome, Italy
| | - Tal Gilboa
- Child Neurology Unit, Hadassah-Hebrew University Medical Center, 9112001, Jerusalem, Israel
| | - Abdullah M Tamim
- Pediatric Neurology, King Faisal Specialist Hospital and Research Center, Riyadh, Saudi Arabia
| | - Mais O Hashem
- Department of Genetics, King Faisal Specialist Hospital and Research Center, Riyadh, Saudi Arabia
| | - Moeenaldeen D AlSayed
- Department of Medical Genetics, King Faisal Specialist Hospital and Research Center, Riyadh, Saudi Arabia
| | - Maha M Abdulrahim
- Department of Medical Genetics, King Faisal Specialist Hospital and Research Center, Riyadh, Saudi Arabia
| | - Mohammed Al-Owain
- Department of Medical Genetics, King Faisal Specialist Hospital and Research Center, Riyadh, Saudi Arabia
| | - Ali Awaji
- Department of Pediatrics, King Fahad Central Hospital in Jizan, Abu Arish, Saudi Arabia
| | - Adel A H Mahmoud
- Pediatric Neurology Department, National Neuroscience Institute, King Fahad Medical City, Riyadh, Saudi Arabia
| | - Eissa A Faqeih
- Section of Medical Genetics, Children's Hospital, King Fahad Medical City, Riyadh, Saudi Arabia
| | - Ali Al Asmari
- Section of Medical Genetics, Children's Hospital, King Fahad Medical City, Riyadh, Saudi Arabia
| | - Sulwan M Algain
- General Pediatrics and Adolescents, King Fahad Medical City, Riyadh, Saudi Arabia
| | - Lamyaa A Jad
- Pediatric Neurology Department, National Neuroscience Institute, King Fahad Medical City, Riyadh, Saudi Arabia
| | - Hesham M Aldhalaan
- Neuroscience Department King Faisal Specialist Hospital and Research Center, Riyadh, Saudi Arabia
| | - Ingo Helbig
- Division of Neurology, The Children's Hospital of Philadelphia, Philadelphia, PA, USA
| | - David A Koolen
- Department of Human Genetics, Radboud University Medical Center, Nijmegen, The Netherlands
| | - Angelika Riess
- Institute of Medical Genetics and Applied Genomics (Tübingen) and Centogene AG (Rostock), Rostock, Germany
| | | | - Peter Bauer
- Institute of Medical Genetics and Applied Genomics (Tübingen) and Centogene AG (Rostock), Rostock, Germany
| | - Suleyman Gulsuner
- Division of Medical Genetics, Department of Medicine, University of Washington, Seattle, WA, USA
| | - Hannah Stamberger
- Center for Molecular Neurology, VIB, Antwerp, Belgium
- Laboratory of Neurogenetics, Institute Born-Bunge, University of Antwerp, Antwerp, Belgium
- Department of Neurology, University Hospital Antwerp, Antwerp, Belgium
| | - Alvin Yu Jin Ng
- Institute of Molecular and Cell Biology, A*STAR, Biopolis, Singapore, 138673, Singapore
| | - Sha Tang
- Division of Clinical Genomics, Ambry Genetics, Aliso Viejo, CA, USA
| | - Sumanty Tohari
- Institute of Molecular and Cell Biology, A*STAR, Biopolis, Singapore, 138673, Singapore
| | - Boris Keren
- APHP, GH Pitié Salpêtrière, Department of Genetics, Unit of Development Genomics, Paris, France
| | | | - Eric W Klee
- Department of Clinical Genomics, Mayo Clinic, 200 First Street SW, Rochester, MN, USA
| | - Sabina Barresi
- Genetics and Rare Diseases Research Division, Ospedale Pediatrico Bambino Gesù, IRCCS, viale San Paolo 15, 00146, Rome, Italy
| | - Marco Tartaglia
- Genetics and Rare Diseases Research Division, Ospedale Pediatrico Bambino Gesù, IRCCS, viale San Paolo 15, 00146, Rome, Italy
| | - Hagar Mor-Shaked
- Department of Genetic and Metabolic Diseases, Hadassah-Hebrew University Medical Center, 9112001, Jerusalem, Israel
| | - Sateesh Maddirevula
- Department of Genetics, King Faisal Specialist Hospital and Research Center, Riyadh, Saudi Arabia
| | - Amber Begtrup
- GeneDx, 207 Perry Parkway, Gaithersburg, MD, 20877, USA
| | | | - Rolph Pfundt
- Department of Human Genetics, Radboud University Medical Center, Nijmegen, The Netherlands
| | - Rebecca Schüle
- Department of Neurology and Hertie-Institute for Clinical Brain Research, University of Tübingen, Tübingen, Germany
- German Center of Neurodegenerative Diseases (DZNE), Tübingen, Germany
| | - Brian Ciruna
- Program in Developmental & Stem Cell Biology, The Hospital for Sick Children, Department of Molecular Genetics, The University of Toronto, Toronto, ON, Canada
| | - Carine Bonnard
- Institute of Medical Biology, A*STAR, Biopolis, Singapore, 138648, Singapore
| | - Mahmoud A Pouladi
- Translational Laboratory in Genetic Medicine, Agency for Science, Technology, and Research, Singapore (A*STAR), 8A Biomedical Grove, Immunos, Level 5, Singapore, 138648, Singapore
- Department of Physiology, National University of Singapore, Singapore, 117597, Singapore
- Department of Medicine, National University of Singapore, Singapore, 117597, Singapore
| | - James C Stewart
- Institute of Molecular and Cell Biology, A*STAR, Biopolis, Singapore, 138673, Singapore
| | - Adam Claridge-Chang
- Institute of Molecular and Cell Biology, A*STAR, Biopolis, Singapore, 138673, Singapore
- Program in Neuroscience and Behavioral Disorders, Duke-NUS Medical School, Singapore, Singapore
| | - Dirk J Lefeber
- Department of Neurology, Donders Center for Brain, Cognition, and Behavior, Nijmegen, The Netherlands
- Department of Laboratory Medicine, Translational Metabolic Laboratory, Nijmegen, The Netherlands
| | - Fowzan S Alkuraya
- Department of Genetics, King Faisal Specialist Hospital and Research Center, Riyadh, Saudi Arabia
| | - Ajay S Mathuru
- Yale-NUS College, 12 College Avenue West, Biopolis, Singapore, Singapore
- Institute of Molecular and Cell Biology, A*STAR, Biopolis, Singapore, 138673, Singapore
| | - Byrappa Venkatesh
- National University of Singapore, Department of Paediatrics, Yong Loo Lin School of Medicine, Biopolis, Singapore, Singapore
- Institute of Molecular and Cell Biology, A*STAR, Biopolis, Singapore, 138673, Singapore
| | - Joseph J Barycki
- Department of Molecular and Structural Biochemistry North Carolina State University, Raleigh, NC, 27607, USA
| | - Melanie A Simpson
- Department of Molecular and Structural Biochemistry North Carolina State University, Raleigh, NC, 27607, USA
| | - Saumya S Jamuar
- Genetics Service, Department of Paediatrics, KK Women's and Children's Hospital, Singapore, Singapore
- Paediatric Academic Clinical Programme, Duke-NUS Medical School, Singapore, Singapore
- SingHealth Duke-NUS Genomic Medicine Centre, Singapore, Singapore
- SingHealth Duke-NUS Institute of Precision Medicine, Singapore, Singapore
| | - Ludger Schöls
- Department of Neurology and Hertie-Institute for Clinical Brain Research, University of Tübingen, Tübingen, Germany.
- German Center of Neurodegenerative Diseases (DZNE), Tübingen, Germany.
| | - Bruno Reversade
- Institute of Medical Biology, A*STAR, Biopolis, Singapore, 138648, Singapore.
- National University of Singapore, Department of Paediatrics, Yong Loo Lin School of Medicine, Biopolis, Singapore, Singapore.
- Institute of Molecular and Cell Biology, A*STAR, Biopolis, Singapore, 138673, Singapore.
- Medical Genetics Department, Koç University School of Medicine, 34010, Istanbul, Turkey.
- Reproductive Biology Laboratory, Obstetrics and Gynaecology, Academic Medical Center (AMC), Meibergdreef 9, 1105 AZ, Amsterdam-Zuidoost, The Netherlands.
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23
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Abasht B, Zhou N, Lee WR, Zhuo Z, Peripolli E. The metabolic characteristics of susceptibility to wooden breast disease in chickens with high feed efficiency. Poult Sci 2019; 98:3246-3256. [PMID: 30995306 DOI: 10.3382/ps/pez183] [Citation(s) in RCA: 45] [Impact Index Per Article: 7.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/29/2018] [Accepted: 03/15/2019] [Indexed: 01/11/2023] Open
Abstract
This study was conducted to characterize metabolic differences between high feed efficiency (HFE) and low feed efficiency (LFE) chickens to investigate why feed efficient chickens are more susceptible to muscle abnormalities such as wooden breast disease. Gene expression profiles were generated by RNA sequencing of pectoralis major muscle samples from 10 HFE and 13 LFE broiler chickens selected from a modern broiler population. Metabolism-associated differentially expressed genes were identified and interpreted by Ingenuity Pathway Analysis and literature mining. Our RNA-seq data indicate decreased glycolytic capacity, increased fatty acid uptake, mitochondrial oxidation of fatty acids, and several other metabolic alterations in the pectoralis major muscle of HFE chickens. We also quantified glycogen content of the pectoralis major muscle and found that the HFE chickens had a significantly (P ≤ 0.05) lower glycogen content. Collectively, this study indicates extensive metabolic differences in the pectoralis major muscle between HFE and LFE chickens and helps identify metabolic features of susceptibility to muscle disorders in modern broiler chickens.
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Affiliation(s)
- Behnam Abasht
- Department of Animal and Food Sciences, University of Delaware, 531 South College Ave, Newark, DE 19716
| | - Nan Zhou
- Department of Animal and Food Sciences, University of Delaware, 531 South College Ave, Newark, DE 19716
| | | | - Zhu Zhuo
- Department of Animal and Food Sciences, University of Delaware, 531 South College Ave, Newark, DE 19716
| | - Elisa Peripolli
- Department of Animal and Food Sciences, University of Delaware, 531 South College Ave, Newark, DE 19716
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24
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Catalytic mechanism of UDP-glucose dehydrogenase. Biochem Soc Trans 2019; 47:945-955. [PMID: 31189734 DOI: 10.1042/bst20190257] [Citation(s) in RCA: 10] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [Key Words] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/29/2019] [Revised: 05/15/2019] [Accepted: 05/21/2019] [Indexed: 11/17/2022]
Abstract
UDP-glucose dehydrogenase (UGDH), an oxidoreductase, catalyzes the NAD+-dependent four-electron oxidation of UDP-glucose to UDP-glucuronic acid. The catalytic mechanism of UGDH remains controversial despite extensive investigation and is classified into two types according to whether an aldehyde intermediate is generated in the first oxidation step. The first type, which involves the presence of this putative aldehyde, is inconsistent with some experimental findings. In contrast, the second type, which indicates that the first oxidation step bypasses the aldehyde via an NAD+-dependent bimolecular nucleophilic substitution (SN2) reaction, is consistent with the experimental phenomena, including those that cannot be explained by the first type. This NAD+-dependent SN2 mechanism is thus more reasonable and likely applicable to other oxidoreductases that catalyze four-electron oxidation reactions.
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25
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Goodwin CR, Ahmed AK, Xia S. UDP-α-D-glucose 6-dehydrogenase: a promising target for glioblastoma. Oncotarget 2019; 10:1542-1543. [PMID: 30899420 PMCID: PMC6422181 DOI: 10.18632/oncotarget.26670] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Key Words] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/01/2018] [Accepted: 02/03/2019] [Indexed: 11/25/2022] Open
Affiliation(s)
- C Rory Goodwin
- Department of Neurology, Hugo W. Moser Research Institute at Kennedy Krieger, Johns Hopkins School of Medicine, Baltimore, MD, USA
| | - A Karim Ahmed
- Department of Neurology, Hugo W. Moser Research Institute at Kennedy Krieger, Johns Hopkins School of Medicine, Baltimore, MD, USA
| | - Shuli Xia
- Department of Neurology, Hugo W. Moser Research Institute at Kennedy Krieger, Johns Hopkins School of Medicine, Baltimore, MD, USA
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26
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Beattie NR, Pioso BJ, Sidlo AM, Keul ND, Wood ZA. Hysteresis and Allostery in Human UDP-Glucose Dehydrogenase Require a Flexible Protein Core. Biochemistry 2018; 57:6848-6859. [PMID: 30457329 PMCID: PMC6312000 DOI: 10.1021/acs.biochem.8b00497] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/07/2023]
Abstract
Human UDP-glucose dehydrogenase (hUGDH) oxidizes UDP-glucose to UDP-glucuronic acid, an essential substrate in the phase II metabolism of drugs. The activity of hUGDH is regulated by the conformation of a buried allosteric switch (T131 loop/α6 helix). Substrate binding induces the allosteric switch to slowly isomerize from an inactive E* conformation to the active E state, which can be observed as enzyme hysteresis. When the feedback inhibitor UDP-xylose binds, the allosteric switch and surrounding residues in the protein core repack, converting the hexamer into an inactive, horseshoe-shaped complex (EΩ). This allosteric transition is facilitated by large cavities and declivities in the protein core that provide the space required to accommodate the alternate packing arrangements. Here, we have used the A104L substitution to fill a cavity in the E state and sterically prevent repacking of the core into the EΩ state. Steady state analysis shows that hUGDHA104L binds UDP-xylose with lower affinity and that the inhibition is no longer cooperative. This means that the allosteric transition to the high-UDP-xylose affinity EΩ state is blocked by the substitution. The crystal structures of hUGDHA104L show that the allosteric switch still adopts the E and E* states, albeit with a more rigid protein core. However, the progress curves of hUGDHA104L do not show hysteresis, which suggests that the E* and E states are now in rapid equilibrium. Our data suggest that hysteresis in native hUGDH originates from the conformational entropy of the E* state protein core.
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Affiliation(s)
- Nathaniel R. Beattie
- Department of Biochemistry & Molecular Biology, University of Georgia, Athens, GA 30602, USA
| | - Brittany J. Pioso
- Department of Biochemistry & Molecular Biology, University of Georgia, Athens, GA 30602, USA
| | - Andrew M. Sidlo
- Department of Biochemistry & Molecular Biology, University of Georgia, Athens, GA 30602, USA
| | - Nicholas D. Keul
- Department of Biochemistry & Molecular Biology, University of Georgia, Athens, GA 30602, USA
| | - Zachary A. Wood
- Department of Biochemistry & Molecular Biology, University of Georgia, Athens, GA 30602, USA
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27
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Bechard ME, Word AE, Tran AV, Liu X, Locasale JW, McDonald OG. Pentose conversions support the tumorigenesis of pancreatic cancer distant metastases. Oncogene 2018; 37:5248-5256. [PMID: 29849117 PMCID: PMC6692915 DOI: 10.1038/s41388-018-0346-5] [Citation(s) in RCA: 19] [Impact Index Per Article: 2.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/21/2017] [Revised: 04/17/2018] [Accepted: 05/11/2018] [Indexed: 12/14/2022]
Abstract
Pancreatic ductal adenocarcinoma (PDAC) adopts several unique metabolic strategies to support primary tumor growth. Whether additional metabolic strategies are adopted to support metastatic tumorigenesis is less clear. This could be particularly relevant for distant metastasis, which often follows a rapidly progressive clinical course. Here we report that PDAC distant metastases evolve a unique series of metabolic reactions to maintain activation of the anabolic glucose enzyme phosphogluconate dehydrogenase (PGD). PGD catalytic activity was recurrently elevated across distant metastases, and modulating PGD activity levels dictated tumorigenic capacity. Metabolomics data raised the possibility that distant metastases evolved a core pentose conversion pathway (PCP) that converted glucose-derived metabolites into PGD substrate, thereby hyperactivating the enzyme. Consistent with this, each individual metabolite in the PCP stimulated PGD catalysis in distant metastases, and knockdown of each individual PCP enzyme selectively impaired tumorigenesis. We propose that the PCP manufactures PGD substrate outside of the rate-limiting oxidative pentose phosphate pathway (oxPPP). This enables PGD-dependent tumorigenesis by providing adequate substrate to fuel high catalytic activity, and raises the possibility that PDAC distant metastases adopt their own unique metabolic strategies to support tumor growth.
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Affiliation(s)
- Matthew E Bechard
- Department of Pathology, Microbiology, and Immunology, Vanderbilt University Medical Center, Nashville, TN, USA
| | - Anna E Word
- Department of Pathology, Microbiology, and Immunology, Vanderbilt University Medical Center, Nashville, TN, USA
- Department of Biomedical Engineering, Vanderbilt University, Nashville, TN, USA
| | - Amanda V Tran
- Department of Pathology, Microbiology, and Immunology, Vanderbilt University Medical Center, Nashville, TN, USA
| | - Xiaojing Liu
- Duke Cancer Institute, Duke University School of Medicine, Durham, NC, USA
- Duke Molecular Physiology Institute, Duke University School of Medicine, Durham, NC, USA
- Department of Pharmacology and Cancer Biology, Duke University School of Medicine, Durham, NC, USA
| | - Jason W Locasale
- Duke Cancer Institute, Duke University School of Medicine, Durham, NC, USA
- Duke Molecular Physiology Institute, Duke University School of Medicine, Durham, NC, USA
- Department of Pharmacology and Cancer Biology, Duke University School of Medicine, Durham, NC, USA
| | - Oliver G McDonald
- Department of Pathology, Microbiology, and Immunology, Vanderbilt University Medical Center, Nashville, TN, USA.
- Epithelial Biology Center, Vanderbilt University Medical Center, Nashville, TN, USA.
- Vanderbilt-Ingram Cancer Center, Vanderbilt University Medical Center, Nashville, TN, USA.
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28
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Oyinlade O, Wei S, Lal B, Laterra J, Zhu H, Goodwin CR, Wang S, Ma D, Wan J, Xia S. Targeting UDP-α-D-glucose 6-dehydrogenase inhibits glioblastoma growth and migration. Oncogene 2018; 37:2615-2629. [PMID: 29479058 PMCID: PMC5957772 DOI: 10.1038/s41388-018-0138-y] [Citation(s) in RCA: 37] [Impact Index Per Article: 5.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/14/2017] [Revised: 11/21/2017] [Accepted: 12/24/2017] [Indexed: 01/07/2023]
Abstract
UDP-glucose 6-dehydrogenase (UGDH) produces UDP-α-D-glucuronic acid, the precursors for glycosaminoglycans (GAGs) and proteoglycans of the extracellular matrix. Elevated GAG formation has been implicated in a variety of human diseases, including glioblastoma (GBM). In our previous study, we found that Krüppel-like factor 4 (KLF4) promotes GBM cell migration by binding to methylated DNA, mainly methylated CpGs (mCpG) and transactivating gene expression. We identified UDGH as one of the downstream targets of KLF4-mCpG binding activity. In this study, we show that KLF4 upregulates UGDH expression in a mCpG-dependent manner, and UGDH is required for KLF4-induced cell migration in vitro. UGDH knockdown decreases GAG abundance in GBM cells, as well as cell proliferation and migration in vitro. In intracranial xenografts, reduced UGDH inhibits tumor growth and migration, accompanied by a decrease in the expression of extracellular matrix proteins such as tenascin C, brevican. Our studies demonstrate a novel DNA methylation-dependent UGDH upregulation by KLF4. Developing UGDH antagonists to decrease the synthesis of extracellular matrix components will be a useful strategy for GBM therapy.
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Affiliation(s)
- Olutobi Oyinlade
- Hugo W. Moser Research Institute at Kennedy Krieger, Baltimore, MD, 21205, USA
- Department of Pharmacology and Molecular Sciences, Johns Hopkins School of Medicine, Baltimore, MD, 21205, USA
| | - Shuang Wei
- Hugo W. Moser Research Institute at Kennedy Krieger, Baltimore, MD, 21205, USA
- Department of Neurology, Johns Hopkins School of Medicine, Baltimore, MD, 21205, USA
- Department of Respiratory and Critical Care Medicine, Tongji Hospital, Tongji Medical College Huazhong University of Science and Technology, Wuhan, 430030, China
| | - Bachchu Lal
- Hugo W. Moser Research Institute at Kennedy Krieger, Baltimore, MD, 21205, USA
- Department of Neurology, Johns Hopkins School of Medicine, Baltimore, MD, 21205, USA
| | - John Laterra
- Hugo W. Moser Research Institute at Kennedy Krieger, Baltimore, MD, 21205, USA
- Department of Neurology, Johns Hopkins School of Medicine, Baltimore, MD, 21205, USA
- Department of Neurosurgery, Johns Hopkins School of Medicine, Baltimore, MD, 21205, USA
- Department of Oncology, Johns Hopkins School of Medicine, Baltimore, MD, 21205, USA
| | - Heng Zhu
- Department of Pharmacology and Molecular Sciences, Johns Hopkins School of Medicine, Baltimore, MD, 21205, USA
- High throughput Biology Center, Johns Hopkins School of Medicine, Baltimore, MD, 21205, USA
| | - C Rory Goodwin
- Department of Neurosurgery, Duke University Medical Center, Durham, NC, 27710, USA
| | - Shuyan Wang
- Hugo W. Moser Research Institute at Kennedy Krieger, Baltimore, MD, 21205, USA
- Department of Neurology, Johns Hopkins School of Medicine, Baltimore, MD, 21205, USA
| | - Ding Ma
- Hugo W. Moser Research Institute at Kennedy Krieger, Baltimore, MD, 21205, USA
- Department of Neurology, Johns Hopkins School of Medicine, Baltimore, MD, 21205, USA
| | - Jun Wan
- Department of Medical and Molecular Genetics, Indiana University School of Medicine, Indianapolis, IN, 46202, USA
| | - Shuli Xia
- Hugo W. Moser Research Institute at Kennedy Krieger, Baltimore, MD, 21205, USA.
- Department of Neurology, Johns Hopkins School of Medicine, Baltimore, MD, 21205, USA.
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Vallet JL, Miles JR, Freking BA, Meyer S. Glucosamine supplementation during late gestation alters placental development and increases litter size. J Anim Sci Biotechnol 2017; 8:68. [PMID: 28883913 PMCID: PMC5580301 DOI: 10.1186/s40104-017-0198-9] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/12/2017] [Accepted: 07/11/2017] [Indexed: 11/17/2022] Open
Abstract
Background During late gestation the placental epithelial interface becomes highly folded, which involves changes in stromal hyaluronan. Hyaluronan is composed of glucoronate and N-acetyl-glucosamine. We hypothesized that supplementing gestating dams with glucosamine during this time would support placental folded-epithelial-bilayer development and increase litter size. In Exp. 1, gilts were unilaterally hysterectomized-ovariectomized (UHO). UHO gilts were mated and then supplemented daily with 10 g glucosamine (n = 16) or glucose (control, n = 17) from d 85 of gestation until slaughter (d 105). At slaughter, the number of live fetuses was recorded and each live fetus and its placenta was weighed. Uterine wall samples adjacent to the largest and smallest fetuses within each litter were processed for histology. In Exp. 2, pregnant sows in a commercial sow farm were supplemented with either 10 g glucosamine or glucose daily from d 85 of gestation to farrowing. Total piglets born and born alive were recorded for each litter. In Exp. 3, the same commercial farm and same protocol were used except that the dose of glucosamine and glucose was doubled to 20 g/d. Results In Exp. 1, the number of live fetuses tended to be greater in glucosamine-treated UHO gilts (P = 0.098). Placental morphometry indicated that the width of the folded bilayer was greater (P = 0.05) in glucosamine-treated gilts. In Exp. 2, litter size did not differ between glucosamine- and glucose-treated sows. However in Exp. 3, the increased dose of glucosamine resulted in a significant treatment by parity interaction (P ≤ 0.01), in which total piglets born and born alive were greater in glucosamine treated sows of later parity (5 and 6). Conclusions These results indicated that glucosamine supplementation increased the width of the folds of the placental bilayer and increased litter size in later parity, intact pregnant commercial sows.
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Affiliation(s)
- Jeffrey L Vallet
- USDA, ARS, U.S. Meat Animal Research Center (USMARC), P.O. Box 166, Clay Center NE, Nebraska, 68933 USA
| | - Jeremy R Miles
- USDA, ARS, U.S. Meat Animal Research Center (USMARC), P.O. Box 166, Clay Center NE, Nebraska, 68933 USA
| | - Bradley A Freking
- USDA, ARS, U.S. Meat Animal Research Center (USMARC), P.O. Box 166, Clay Center NE, Nebraska, 68933 USA
| | - Shane Meyer
- Plymouth Ag Group, Diller NE, Nebraska, 68342 USA
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Beattie NR, Keul ND, Sidlo AM, Wood ZA. Allostery and Hysteresis Are Coupled in Human UDP-Glucose Dehydrogenase. Biochemistry 2017; 56:202-211. [PMID: 27966912 PMCID: PMC5293408 DOI: 10.1021/acs.biochem.6b01044] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/28/2022]
Abstract
Human UDP-glucose dehydrogenase (hUGDH) is regulated by an atypical allosteric mechanism in which the feedback inhibitor UDP-xylose (UDP-Xyl) competes with the substrate for the active site. Binding of UDP-Xyl triggers the T131-loop/α6 allosteric switch, which converts the hexameric structure of hUGDH into an inactive, horseshoe-shaped complex (EΩ). This allosteric transition buries residue A136 in the protein core to produce a subunit interface that favors the EΩ structure. Here we use a methionine substitution to prevent the burial of A136 and trap the T131-loop/α6 switch in the active conformation. We show that hUGDHA136M does not exhibit substrate cooperativity, which is strong evidence that the methionine substitution prevents the formation of the low-UDP-Glc-affinity EΩ state. In addition, the inhibitor affinity of hUGDHA136M is reduced 14-fold, which most likely represents the Ki for competitive inhibition in the absence of the allosteric transition to the higher-affinity EΩ state. hUGDH also displays a lag in progress curves, which is caused by a slow, substrate-induced isomerization that activates the enzyme. Stopped-flow analysis shows that hUGDHA136M does not exhibit hysteresis, which suggests that the T131-loop/α6 switch is the source of the slow isomerization. This interpretation is supported by the 2.05 Å resolution crystal structure of hUGDHA136M, which shows that the A136M substitution has stabilized the active conformation of the T131-loop/α6 allosteric switch. This work shows that the T131-loop/α6 allosteric switch couples allostery and hysteresis in hUGDH.
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Affiliation(s)
- Nathaniel R. Beattie
- Department of Biochemistry & Molecular Biology, University of Georgia, Athens, GA 30602, USA
| | - Nicholas D. Keul
- Department of Biochemistry & Molecular Biology, University of Georgia, Athens, GA 30602, USA
| | - Andrew M. Sidlo
- Department of Biochemistry & Molecular Biology, University of Georgia, Athens, GA 30602, USA
| | - Zachary A. Wood
- Department of Biochemistry & Molecular Biology, University of Georgia, Athens, GA 30602, USA
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31
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Liver glucose metabolism in humans. Biosci Rep 2016; 36:BSR20160385. [PMID: 27707936 PMCID: PMC5293555 DOI: 10.1042/bsr20160385] [Citation(s) in RCA: 227] [Impact Index Per Article: 25.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/06/2016] [Revised: 09/19/2016] [Accepted: 10/04/2016] [Indexed: 12/16/2022] Open
Abstract
Information about normal hepatic glucose metabolism may help to understand pathogenic mechanisms underlying obesity and diabetes mellitus. In addition, liver glucose metabolism is involved in glycosylation reactions and connected with fatty acid metabolism. The liver receives dietary carbohydrates directly from the intestine via the portal vein. Glucokinase phosphorylates glucose to glucose 6-phosphate inside the hepatocyte, ensuring that an adequate flow of glucose enters the cell to be metabolized. Glucose 6-phosphate may proceed to several metabolic pathways. During the post-prandial period, most glucose 6-phosphate is used to synthesize glycogen via the formation of glucose 1-phosphate and UDP–glucose. Minor amounts of UDP–glucose are used to form UDP–glucuronate and UDP–galactose, which are donors of monosaccharide units used in glycosylation. A second pathway of glucose 6-phosphate metabolism is the formation of fructose 6-phosphate, which may either start the hexosamine pathway to produce UDP-N-acetylglucosamine or follow the glycolytic pathway to generate pyruvate and then acetyl-CoA. Acetyl-CoA may enter the tricarboxylic acid (TCA) cycle to be oxidized or may be exported to the cytosol to synthesize fatty acids, when excess glucose is present within the hepatocyte. Finally, glucose 6-phosphate may produce NADPH and ribose 5-phosphate through the pentose phosphate pathway. Glucose metabolism supplies intermediates for glycosylation, a post-translational modification of proteins and lipids that modulates their activity. Congenital deficiency of phosphoglucomutase (PGM)-1 and PGM-3 is associated with impaired glycosylation. In addition to metabolize carbohydrates, the liver produces glucose to be used by other tissues, from glycogen breakdown or from de novo synthesis using primarily lactate and alanine (gluconeogenesis).
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AglM and VNG1048G, Two Haloarchaeal UDP-Glucose Dehydrogenases, Show Different Salt-Related Behaviors. Life (Basel) 2016; 6:life6030031. [PMID: 27527219 PMCID: PMC5041007 DOI: 10.3390/life6030031] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/11/2016] [Revised: 07/27/2016] [Accepted: 07/29/2016] [Indexed: 11/16/2022] Open
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Abasht B, Mutryn MF, Michalek RD, Lee WR. Oxidative Stress and Metabolic Perturbations in Wooden Breast Disorder in Chickens. PLoS One 2016; 11:e0153750. [PMID: 27097013 PMCID: PMC4838225 DOI: 10.1371/journal.pone.0153750] [Citation(s) in RCA: 192] [Impact Index Per Article: 21.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/08/2015] [Accepted: 04/04/2016] [Indexed: 11/30/2022] Open
Abstract
This study was conducted to characterize metabolic features of the breast muscle (pectoralis major) in chickens affected with the Wooden Breast myopathy. Live birds from two purebred chicken lines and one crossbred commercial broiler population were clinically examined by manual palpation of the breast muscle (pectoralis major) at 47–48 days of age. Metabolite abundance was determined by gas chromatography/mass spectrometry (GC/MS) and liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS) using breast muscle tissue samples from 16 affected and 16 unaffected chickens. Muscle glycogen content was also quantified in breast muscle tissue samples from affected and unaffected chickens. In total, levels of 140 biochemicals were significantly different (FDR < 0.1 and fold-change A/U > 1.3 or < 0.77) between affected and unaffected chickens. Glycogen content measurements were considerably lower (1.7-fold) in samples taken from Wooden Breast affected birds when compared with samples from unaffected birds. Affected tissues exhibited biomarkers related to increased oxidative stress, elevated protein levels, muscle degradation, and altered glucose utilization. Affected muscle also showed elevated levels of hypoxanthine, xanthine, and urate molecules, the generation of which can contribute to altered redox homeostasis. In conclusion, our findings show that Wooden Breast affected tissues possess a unique metabolic signature. This unique profile may identify candidate biomarkers for diagnostic utilization and provide mechanistic insight into altered biochemical processes contributing to tissue hardening associated with the Wooden Breast myopathy in commercial chickens.
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Affiliation(s)
- Behnam Abasht
- Department of Animal and Food Sciences, University of Delaware, Newark, DE, United States of America
- * E-mail:
| | - Marie F. Mutryn
- Department of Animal and Food Sciences, University of Delaware, Newark, DE, United States of America
| | | | - William R. Lee
- Maple Leaf Farms, Leesburg, IN, United States of America
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Chu X, Han J, Guo D, Fu Z, Liu W, Tao Y. Characterization of UDP-glucose dehydrogenase from Pasteurella multocida CVCC 408 and its application in hyaluronic acid biosynthesis. Enzyme Microb Technol 2016; 85:64-70. [DOI: 10.1016/j.enzmictec.2015.12.009] [Citation(s) in RCA: 7] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/21/2015] [Revised: 12/17/2015] [Accepted: 12/18/2015] [Indexed: 10/22/2022]
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35
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Paul D, Chanukuppa V, Reddy PJ, Taunk K, Adhav R, Srivastava S, Santra MK, Rapole S. Global proteomic profiling identifies etoposide chemoresistance markers in non-small cell lung carcinoma. J Proteomics 2016; 138:95-105. [DOI: 10.1016/j.jprot.2016.02.008] [Citation(s) in RCA: 17] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/27/2015] [Revised: 12/31/2015] [Accepted: 02/11/2016] [Indexed: 02/05/2023]
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36
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Scoglio S, Lo Curcio V, Catalani S, Palma F, Battistelli S, Benedetti S. Inhibitory effects of Aphanizomenon flos-aquae constituents on human UDP-glucose dehydrogenase activity. J Enzyme Inhib Med Chem 2016; 31:1492-7. [PMID: 26903444 DOI: 10.3109/14756366.2016.1149478] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/16/2023] Open
Abstract
OBJECTIVE The purpose of this study was to investigate the in vitro inhibitory effects of the edible microalga Aphanizomenon flos-aquae (AFA) on human UDP-α-d-glucose 6-dehydrogenase (UGDH) activity, a cytosolic enzyme involved both in tumor progression and in phytochemical bioavailability. METHODS Both the hydrophilic and ethanolic AFA extracts as well as the constitutive active principles phycocyanin (PC), phycocyanobilin (PCB) and mycosporine-like amino acids (MAAs) were tested. RESULTS Among AFA components, PCB presented the strongest inhibitory effect on UGDH activity, acting as a competitive inhibitor with respect to UDP-glucose and a non-competitive inhibitor with respect to NAD(+). In preliminary experiments, AFA PCB was also effective in reducing the colony formation capacity of PC-3 prostate cancer cells and FTC-133 thyroid cancer cells. CONCLUSIONS Overall, these findings confirmed that AFA and its active principles are natural compounds with high biological activity. Further studies evaluating the effects of AFA PCB in reducing tumor cell growth and phytochemical glucuronidation are encouraged.
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Affiliation(s)
| | - Valeria Lo Curcio
- b Department of Biomolecular Sciences , University of Urbino "Carlo Bo" , Urbino , Italy
| | - Simona Catalani
- b Department of Biomolecular Sciences , University of Urbino "Carlo Bo" , Urbino , Italy
| | - Francesco Palma
- b Department of Biomolecular Sciences , University of Urbino "Carlo Bo" , Urbino , Italy
| | - Serafina Battistelli
- b Department of Biomolecular Sciences , University of Urbino "Carlo Bo" , Urbino , Italy
| | - Serena Benedetti
- b Department of Biomolecular Sciences , University of Urbino "Carlo Bo" , Urbino , Italy
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37
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Weyler C, Bureik M, Heinzle E. Selective oxidation of UDP-glucose to UDP-glucuronic acid using permeabilized Schizosaccharomyces pombe expressing human UDP-glucose 6-dehydrogenase. Biotechnol Lett 2015; 38:477-81. [PMID: 26582015 DOI: 10.1007/s10529-015-1995-x] [Citation(s) in RCA: 8] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/01/2015] [Accepted: 11/03/2015] [Indexed: 10/22/2022]
Abstract
OBJECTIVES To use permeabilized cells of the fission yeast, Schizosaccharomyces pombe, that expresses human UDP-glucose 6-dehydrogenase (UGDH, EC 1.1.1.22), for the production of UDP-glucuronic acid from UDP-glucose. RESULTS In cell extracts no activity was detected. Therefore, cells were permeabilized with 0.3 % (v/v) Triton X-100. After washing away all low molecular weight metabolites, the permeabilized cells were directly used as whole cell biocatalyst. Substrates were 5 mM UDP-glucose and 10 mM NAD(+). Divalent cations were not added to the reaction medium as they promoted UDP-glucose hydrolysis. With this reaction system 5 mM UDP-glucose were converted into 5 mM UDP-glucuronic acid within 3 h. CONCLUSIONS Recombinant permeabilized cells of S. pombe can be used to synthesize UDP-glucuronic acid with 100 % yield and selectivity.
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Affiliation(s)
- Christian Weyler
- Biochemical Engineering Institute, Saarland University, Campus A1.5, 66123, Saarbrücken, Germany
| | - Matthias Bureik
- School of Pharmaceutical Science and Technology (SPST), Tianjin University, Building 24, 92 Weijin Road, Nankai District, Tianjin, 300072, People's Republic of China
| | - Elmar Heinzle
- Biochemical Engineering Institute, Saarland University, Campus A1.5, 66123, Saarbrücken, Germany.
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38
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Singh S, Michalska K, Bigelow L, Endres M, Kharel MK, Babnigg G, Yennamalli RM, Bingman CA, Joachimiak A, Thorson JS, Phillips GN. Structural Characterization of CalS8, a TDP-α-D-Glucose Dehydrogenase Involved in Calicheamicin Aminodideoxypentose Biosynthesis. J Biol Chem 2015; 290:26249-58. [PMID: 26240141 DOI: 10.1074/jbc.m115.673459] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/18/2015] [Indexed: 11/06/2022] Open
Abstract
Classical UDP-glucose 6-dehydrogenases (UGDHs; EC 1.1.1.22) catalyze the conversion of UDP-α-d-glucose (UDP-Glc) to the key metabolic precursor UDP-α-d-glucuronic acid (UDP-GlcA) and display specificity for UDP-Glc. The fundamental biochemical and structural study of the UGDH homolog CalS8 encoded by the calicheamicin biosynthetic gene is reported and represents one of the first studies of a UGDH homolog involved in secondary metabolism. The corresponding biochemical characterization of CalS8 reveals CalS8 as one of the first characterized base-permissive UGDH homologs with a >15-fold preference for TDP-Glc over UDP-Glc. The corresponding structure elucidations of apo-CalS8 and the CalS8·substrate·cofactor ternary complex (at 2.47 and 1.95 Å resolution, respectively) highlight a notably high degree of conservation between CalS8 and classical UGDHs where structural divergence within the intersubunit loop structure likely contributes to the CalS8 base permissivity. As such, this study begins to provide a putative blueprint for base specificity among sugar nucleotide-dependent dehydrogenases and, in conjunction with prior studies on the base specificity of the calicheamicin aminopentosyltransferase CalG4, provides growing support for the calicheamicin aminopentose pathway as a TDP-sugar-dependent process.
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Affiliation(s)
- Shanteri Singh
- From the Center for Pharmaceutical Research and Innovation, University of Kentucky College of Pharmacy, Lexington, Kentucky 40536-0596
| | - Karolina Michalska
- the Midwest Center for Structural Genomics and Structural Biology Center, Biosciences Division, Argonne National Laboratory, Argonne, Illinois 60439
| | - Lance Bigelow
- the Midwest Center for Structural Genomics and Structural Biology Center, Biosciences Division, Argonne National Laboratory, Argonne, Illinois 60439
| | - Michael Endres
- the Midwest Center for Structural Genomics and Structural Biology Center, Biosciences Division, Argonne National Laboratory, Argonne, Illinois 60439
| | - Madan K Kharel
- the School of Pharmacy, University of Maryland Eastern Shore, Princess Anne, Maryland 21853
| | - Gyorgy Babnigg
- the Midwest Center for Structural Genomics and Structural Biology Center, Biosciences Division, Argonne National Laboratory, Argonne, Illinois 60439
| | - Ragothaman M Yennamalli
- the Department of BioSciences, Department of Chemistry, Rice University, Houston, Texas 77005
| | - Craig A Bingman
- the Department of Biochemistry, University of Wisconsin-Madison, Madison, Wisconsin 53706, and
| | - Andrzej Joachimiak
- the Midwest Center for Structural Genomics and Structural Biology Center, Biosciences Division, Argonne National Laboratory, Argonne, Illinois 60439
| | - Jon S Thorson
- From the Center for Pharmaceutical Research and Innovation, University of Kentucky College of Pharmacy, Lexington, Kentucky 40536-0596,
| | - George N Phillips
- the Department of BioSciences, Department of Chemistry, Rice University, Houston, Texas 77005 the Department of Biochemistry, University of Wisconsin-Madison, Madison, Wisconsin 53706, and
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Duan XC, Lu AM, Gu B, Cai ZP, Ma HY, Wei S, Laborda P, Liu L, Voglmeir J. Functional characterization of the UDP-xylose biosynthesis pathway in Rhodothermus marinus. Appl Microbiol Biotechnol 2015; 99:9463-72. [PMID: 26033773 DOI: 10.1007/s00253-015-6683-1] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/12/2015] [Revised: 04/20/2015] [Accepted: 05/07/2015] [Indexed: 10/23/2022]
Abstract
UDP-glucuronic acid dehydrogenase (UGD) and UDP-xylose synthase (UXS) are the two enzymes responsible for the biosynthesis of UDP-xylose from UDP-glucose. Several UGDs from bacterial sources, which oxidize UDP-glucose to glucuronic acid, have been found and functionally characterized whereas only few reports on bacterial UXS isoforms exist. Rhodothermus marinus, a halothermophilic bacterium commonly found in hot springs, proved to be a valuable source of carbohydrate active enzymes of biotechnological interest, such as xylanases, mannanases, and epimerases. However, no enzymes of R. marinus involved in the biosynthesis or modification of nucleotide sugars have been reported yet. Herein, we describe the cloning and characterization of two putative UGD (RmUGD1 and RmUGD2) and one UXS (RmUXS) isoform from this organism. All three enzymes could be expressed in recombinant form and purified to near homogeneity. UPLC- and NMR-based activity tests showed that RmUGD1 and RmUXS are indeed active enzymes, whereas no enzymatic activity could be detected by RmUGD2. Both RmUGD1 and RmUXS showed a temperature optimum of 60 °C, with almost no loss of activity after 1 h exposure at 70 °C. No metal ions were required for enzymatic activities. Zn(2+) ions strongly inhibited both enzymes. RmUGD1 showed higher salt tolerance and had a higher pH optimum than RmUXS. Furthermore, RmUGD1 was inhibited by UDP-xylose at higher concentrations. By coupling recombinant RmUXS and RmUGD1, UDP-xylose could be successfully synthesized directly from UDP-glucose. The high activity of the herein described enzymes make RmUGD1 and RmUXS the first thermo-tolerant biocatalysts for the synthesis of UDP-glucuronic acid and UDP-xylose.
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Affiliation(s)
- Xu C Duan
- Glycomics and Glycan Bioengineering Research Center (GGBRC), College of Food Science and Technology, Nanjing Agricultural University, Nanjing, People's Republic of China
| | - Ai M Lu
- College of Sciences, Nanjing Agricultural University, Nanjing, People's Republic of China
| | - Bin Gu
- Glycomics and Glycan Bioengineering Research Center (GGBRC), College of Food Science and Technology, Nanjing Agricultural University, Nanjing, People's Republic of China
| | - Zhi P Cai
- Glycomics and Glycan Bioengineering Research Center (GGBRC), College of Food Science and Technology, Nanjing Agricultural University, Nanjing, People's Republic of China
| | - Hong Y Ma
- Department of Plant Pathology, Nanjing Agricultural University, Nanjing, People's Republic of China
| | - Shuang Wei
- Glycomics and Glycan Bioengineering Research Center (GGBRC), College of Food Science and Technology, Nanjing Agricultural University, Nanjing, People's Republic of China
| | - Pedro Laborda
- Glycomics and Glycan Bioengineering Research Center (GGBRC), College of Food Science and Technology, Nanjing Agricultural University, Nanjing, People's Republic of China
| | - Li Liu
- Glycomics and Glycan Bioengineering Research Center (GGBRC), College of Food Science and Technology, Nanjing Agricultural University, Nanjing, People's Republic of China.
| | - Josef Voglmeir
- Glycomics and Glycan Bioengineering Research Center (GGBRC), College of Food Science and Technology, Nanjing Agricultural University, Nanjing, People's Republic of China.
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Shi L, Ji B, Kolar-Znika L, Boskovic A, Jadeau F, Combet C, Grangeasse C, Franjevic D, Talla E, Mijakovic I. Evolution of bacterial protein-tyrosine kinases and their relaxed specificity toward substrates. Genome Biol Evol 2015; 6:800-17. [PMID: 24728941 PMCID: PMC4007543 DOI: 10.1093/gbe/evu056] [Citation(s) in RCA: 33] [Impact Index Per Article: 3.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/04/2023] Open
Abstract
It has often been speculated that bacterial protein-tyrosine kinases (BY-kinases) evolve rapidly and maintain relaxed substrate specificity to quickly adopt new substrates when evolutionary pressure in that direction arises. Here, we report a phylogenomic and biochemical analysis of BY-kinases, and their relationship to substrates aimed to validate this hypothesis. Our results suggest that BY-kinases are ubiquitously distributed in bacterial phyla and underwent a complex evolutionary history, affected considerably by gene duplications and horizontal gene transfer events. This is consistent with the fact that the BY-kinase sequences represent a high level of substitution saturation and have a higher evolutionary rate compared with other bacterial genes. On the basis of similarity networks, we could classify BY kinases into three main groups with 14 subgroups. Extensive sequence conservation was observed only around the three canonical Walker motifs, whereas unique signatures proposed the functional speciation and diversification within some subgroups. The relationship between BY-kinases and their substrates was analyzed using a ubiquitous substrate (Ugd) and some Firmicute-specific substrates (YvyG and YjoA) from Bacillus subtilis. No evidence of coevolution between kinases and substrates at the sequence level was found. Seven BY-kinases, including well-characterized and previously uncharacterized ones, were used for experimental studies. Most of the tested kinases were able to phosphorylate substrates from B. subtilis (Ugd, YvyG, and YjoA), despite originating from very distant bacteria. Our results are consistent with the hypothesis that BY-kinases have evolved relaxed substrate specificity and are probably maintained as rapidly evolving platforms for adopting new substrates.
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Affiliation(s)
- Lei Shi
- INRA-AgroParisTech UMR 1319, Micalis-CBAI, Thiverval-Grignon, France
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Puchner C, Eixelsberger T, Nidetzky B, Brecker L. Saturation transfer difference NMR to study substrate and product binding to human UDP-xylose synthase (hUXS1A) during catalytic event. RSC Adv 2015. [DOI: 10.1039/c5ra18284k] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/21/2022] Open
Abstract
The human form of UDP-xylose synthase (hUXS1A) is studied with respect to its substrate and co-enzyme binding in binary and ternary complexes using saturation transfer difference (STD) NMR and in situ NMR.
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Affiliation(s)
- Claudia Puchner
- University of Vienna
- Institute of Organic Chemistry
- A-1090 Vienna
- Austria
| | - Thomas Eixelsberger
- Graz University of Technology
- Institute of Biotechnology and Biochemical Engineering
- A-8010 Graz
- Austria
| | - Bernd Nidetzky
- Graz University of Technology
- Institute of Biotechnology and Biochemical Engineering
- A-8010 Graz
- Austria
| | - Lothar Brecker
- University of Vienna
- Institute of Organic Chemistry
- A-1090 Vienna
- Austria
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42
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Kadirvelraj R, Custer GS, Keul ND, Sennett NC, Sidlo AM, Walsh RM, Wood ZA. Hysteresis in human UDP-glucose dehydrogenase is due to a restrained hexameric structure that favors feedback inhibition. Biochemistry 2014; 53:8043-51. [PMID: 25478983 DOI: 10.1021/bi500594x] [Citation(s) in RCA: 8] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/30/2022]
Abstract
Human UDP-α-d-glucose-6-dehydrogenase (hUGDH) displays hysteresis because of a slow isomerization from an inactive state (E*) to an active state (E). Here we show that the structure of E* constrains hUGDH in a conformation that favors feedback inhibition at physiological pH. The feedback inhibitor UDP-α-d-xylose (UDP-Xyl) competes with the substrate UDP-α-d-glucose for the active site. Upon binding, UDP-Xyl triggers an allosteric switch that changes the structure and affinity of the intersubunit interface to form a stable but inactive horseshoe-shaped hexamer. Using sedimentation velocity studies and a new crystal structure, we show that E* represents a stable conformational intermediate between the active and feedback-inhibited conformations. Because the allosteric switch occludes the cofactor and substrate binding sites in the inactive hexamer, the intermediate conformation observed in the crystal structure is consistent with the E* transient observed in relaxation studies. Steady-state analysis shows that the E* conformation enhances the affinity of hUGDH for the allosteric inhibitor UDP-Xyl by 8.6-fold (Ki = 810 nM). We present a model in which the constrained quaternary structure permits a small effector molecule to leverage a disproportionately large allosteric response.
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Affiliation(s)
- Renuka Kadirvelraj
- Department of Biochemistry & Molecular Biology, University of Georgia , Athens, Georgia 30602, United States
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Wen Y, Li J, Wang L, Tie K, Magdalou J, Chen L, Wang H. UDP-glucose dehydrogenase modulates proteoglycan synthesis in articular chondrocytes: its possible involvement and regulation in osteoarthritis. Arthritis Res Ther 2014; 16:484. [PMID: 25465897 PMCID: PMC4298080 DOI: 10.1186/s13075-014-0484-2] [Citation(s) in RCA: 11] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/19/2014] [Accepted: 11/05/2014] [Indexed: 11/20/2022] Open
Abstract
Introduction The objective of this study was to investigate the possible role of UDP-glucose dehydrogenase (UGDH) in osteoarthritis (OA) and uncover whether, furthermore how interleukin-1beta (IL-1β) affects UGDH gene expression. Methods UGDH specific siRNAs were applied to determine the role of UGDH in proteoglycan (PG) synthesis in human articular chondrocytes. Protein levels of UGDH and Sp1 in human and rat OA cartilage were detected. Then, human primary chondrocytes were treated with IL-1β to find out whether and how IL-1β could regulate the gene expression of UGDH and its trans-regulators, that is Sp1, Sp3 and c-Krox. Finally, p38 mitogen-activated protein kinase (MAPK) inhibitor SB203580 and stress-activated protein kinase/c-Jun N-terminal kinase (SAP/JNK) inhibitor SP600125 were used to pick out the pathway that mediated the IL-1β-modulated PGs synthesis and gene expression of UGDH, Sp1, Sp3 and c-Krox. Results UGDH specific siRNAs markedly inhibited UGDH mRNA and protein expression, and thus led to an obvious suppression of PGs synthesis in human articular chondrocytes. UGDH protein level in human and rat OA cartilage were much lower than the corresponding controls and negatively correlated to the degree of OA. Decrease in Sp1 protein level was also observed in human and rat OA cartilage respectively. Meanwhile, IL-1β suppressed UGDH gene expression in human articular chondrocytes in the late phase, which also modulated gene expression of Sp1, Sp3 and c-Krox and increased both Sp3/Sp1 and c-Krox/Sp1 ratio. Moreover, the inhibition of SAP/JNK and p38 MAPK pathways both resulted in an obvious attenuation of the IL-1β-induced suppression on the UGDH gene expression. Conclusions UGDH is essential in the PGs synthesis of articular chondrocytes, while the suppressed expression of UGDH might probably be involved in advanced OA, partly due to the modulation of p38 MAPK and SAP/JNK pathways and its trans-regulators by IL-1β. Electronic supplementary material The online version of this article (doi:10.1186/s13075-014-0484-2) contains supplementary material, which is available to authorized users.
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Park JM, Han NY, Han YM, Chung MK, Lee HK, Ko KH, Kim EH, Hahm KB. Predictive proteomic biomarkers for inflammatory bowel disease-associated cancer: Where are we now in the era of the next generation proteomics? World J Gastroenterol 2014; 20:13466-13476. [PMID: 25309077 PMCID: PMC4188898 DOI: 10.3748/wjg.v20.i37.13466] [Citation(s) in RCA: 6] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 12/27/2013] [Revised: 03/10/2014] [Accepted: 06/17/2014] [Indexed: 02/06/2023] Open
Abstract
Recent advances in genomic medicine have opened up the possibility of tailored medicine that may eventually replace traditional “one-size-fits all” approaches to the treatment of inflammatory bowel disease (IBD). In addition to exploring the interactions between hosts and microbes, referred to as the microbiome, a variety of strategies that can be tailored to an individual in the coming era of personalized medicine in the treatment of IBD are being investigated. These include prompt genomic screening of patients at risk of developing IBD, the utility of molecular discrimination of IBD subtypes among patients diagnosed with IBD, and the discovery of proteome biomarkers to diagnose or predict cancer risks. Host genetic factors influence the etiology of IBD, as do microbial ecosystems in the human bowel, which are not uniform, but instead represent many different microhabitats that can be influenced by diet and might affect processes essential to bowel metabolism. Further advances in basic research regarding intestinal inflammation may reveal new insights into the role of inflammatory mediators, referred to as the inflammasome, and the macromolecular complex of metabolites formed by intestinal bacteria. Collectively, knowledge of the inflammasome and metagenomics will lead to the development of biomarkers for IBD that target specific pathogenic mechanisms involved in the spontaneous progress of IBD. In this review article, our recent results regarding the discovery of potential proteomic biomarkers using a label-free quantification technique are introduced and on-going projects contributing to either the discrimination of IBD subtypes or to the prediction of cancer risks are accompanied by updated information from IBD biomarker research.
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Wen Y, Li J, Tan Y, Qin J, Xie X, Wang L, Mei Q, Wang H, Magdalou J, Chen L. Angelica Sinensis polysaccharides stimulated UDP-sugar synthase genes through promoting gene expression of IGF-1 and IGF1R in chondrocytes: promoting anti-osteoarthritic activity. PLoS One 2014; 9:e107024. [PMID: 25202993 PMCID: PMC4159308 DOI: 10.1371/journal.pone.0107024] [Citation(s) in RCA: 19] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/19/2014] [Accepted: 08/06/2014] [Indexed: 11/17/2022] Open
Abstract
BACKGROUND Osteoarthritis (OA) is a chronic joints disease characterized by progressive degeneration of articular cartilage due to the loss of cartilage matrix. Previously, we found, for the first time, that an acidic glycan from Angelica Sinensis Polysaccharides (APSs), namely the APS-3c, could protect rat cartilage from OA due to promoting glycosaminoglycan (GAG) synthesis in chondrocytes. In the present work, we tried to further the understanding of ASP-3c's anti-OA activity. METHODOLOGY/PRINCIPAL FINDINGS Human primary chondrocytes were treated with APS-3c or/and recombinant human interleukin 1β (IL-1β). It turned out that APS-3c promoted synthesis of UDP-xylose and GAG, as well as the gene expression of UDP-sugar synthases (USSs), insulin like growth factor 1 (IGF1) and IGF1 receptor (IGF1R), and attenuated the degenerative phenotypes, suppressed biosynthesis of UDP-sugars and GAG, and inhibited the gene expression of USSs, IGF1 and IGF1R induced by IL-1β. Then, we induced a rat OA model with papain, and found that APS-3c also stimulated GAG synthesis and gene expression of USSs, IGF1 and IGF1R in vivo. Additionally, recombinant human IGF1 and IGF1R inhibitor NP-AEW541 were applied to figure out the correlation between stimulated gene expression of USSs, IGF1 and IGF1R induced by APS-3c. It tuned out that the promoted GAG synthesis and USSs gene expression induced by APS-3c was mediated by the stimulated IGF1 and IGF1R gene expression, but not through directly activation of IGF1R signaling pathway. CONCLUSIONS/SIGNIFICANCES We demonstrated for the first time that APS-3c presented anti-OA activity through stimulating IGF-1 and IGF1R gene expression, but not directly activating the IGF1R signaling pathway, which consequently promoted UDP-sugars and GAG synthesis due to up-regulating gene expression of USSs. Our findings presented a better understanding of APS-3c's anti-OA activity and suggested that APS-3c could potentially be a novel therapeutic agent for OA.
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Affiliation(s)
- Yinxian Wen
- Department of Orthopedic Surgery, Zhongnan Hospital of Wuhan University, Wuhan, China
| | - Jing Li
- Department of pharmacology, Basic Medical School of Wuhan University, Wuhan, China
| | - Yang Tan
- Department of Orthopedic Surgery, Zhongnan Hospital of Wuhan University, Wuhan, China
| | - Jun Qin
- Department of Orthopedic Surgery, Zhongnan Hospital of Wuhan University, Wuhan, China
| | - Xianfei Xie
- Department of pharmacology, Basic Medical School of Wuhan University, Wuhan, China
| | - Linlong Wang
- Department of Orthopedic Surgery, Zhongnan Hospital of Wuhan University, Wuhan, China
| | - Qibing Mei
- Department of Pharmacology, School of Pharmacy, The Fourth Military Medical University, Xi'an, China
| | - Hui Wang
- Department of pharmacology, Basic Medical School of Wuhan University, Wuhan, China; Hubei Provincial Key Laboratory of Developmentally Originated Disease, Wuhan, China
| | - Jacques Magdalou
- UMR 7365 CNRS-Université de Lorraine, Faculté de Médecine, Vandœuvre-lès-Nancy, France
| | - Liaobin Chen
- Department of Orthopedic Surgery, Zhongnan Hospital of Wuhan University, Wuhan, China; Hubei Provincial Key Laboratory of Developmentally Originated Disease, Wuhan, China
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Schreier VN, Pethő L, Orbán E, Marquardt A, Petre BA, Mező G, Manea M. Protein expression profile of HT-29 human colon cancer cells after treatment with a cytotoxic daunorubicin-GnRH-III derivative bioconjugate. PLoS One 2014; 9:e94041. [PMID: 24718594 PMCID: PMC3981732 DOI: 10.1371/journal.pone.0094041] [Citation(s) in RCA: 7] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/09/2013] [Accepted: 03/10/2014] [Indexed: 11/18/2022] Open
Abstract
Targeted delivery of chemotherapeutic agents is a new approach for the treatment of cancer, which provides increased selectivity and decreased systemic toxicity. We have recently developed a promising drug delivery system, in which the anticancer drug daunorubicin (Dau) was attached via oxime bond to a gonadotropin-releasing hormone-III (GnRH-III) derivative used as a targeting moiety (Glp-His-Trp-Lys(Ac)-His-Asp-Trp-Lys(Dau = Aoa)-Pro-Gly-NH2; Glp = pyroglutamic acid, Ac = acetyl; Aoa = aminooxyacetyl). This bioconjugate exerted in vitro cytostatic/cytotoxic effect on human breast, prostate and colon cancer cells, as well as significant in vivo tumor growth inhibitory effect on colon carcinoma bearing mice. In our previous studies, H-Lys(Dau = Aoa)-OH was identified as the smallest metabolite produced in the presence of rat liver lysosomal homogenate, which was able to bind to DNA in vitro. To get a deeper insight into the mechanism of action of the bioconjugate, changes in the protein expression profile of HT-29 human colon cancer cells after treatment with the bioconjugate or free daunorubicin were investigated by mass spectrometry-based proteomics. Our results indicate that several metabolism-related proteins, molecular chaperons and proteins involved in signaling are differently expressed after targeted chemotherapeutic treatment, leading to the conclusion that the bioconjugate exerts its cytotoxic action by interfering with multiple intracellular processes.
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Affiliation(s)
| | - Lilla Pethő
- Department of Chemistry, University of Konstanz, Konstanz, Germany
- MTA-ELTE Research Group of Peptide Chemistry, Budapest, Hungary
| | - Erika Orbán
- MTA-ELTE Research Group of Peptide Chemistry, Budapest, Hungary
| | | | | | - Gábor Mező
- MTA-ELTE Research Group of Peptide Chemistry, Budapest, Hungary
| | - Marilena Manea
- Department of Chemistry, University of Konstanz, Konstanz, Germany
- Zukunftskolleg, University of Konstanz, Konstanz, Germany
- * E-mail:
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Mainprize IL, Bean JD, Bouwman C, Kimber MS, Whitfield C. The UDP-glucose dehydrogenase of Escherichia coli K-12 displays substrate inhibition by NAD that is relieved by nucleotide triphosphates. J Biol Chem 2013; 288:23064-74. [PMID: 23792965 DOI: 10.1074/jbc.m113.486613] [Citation(s) in RCA: 14] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/10/2023] Open
Abstract
UDP-glucose dehydrogenase (Ugd) generates UDP-glucuronic acid, an important precursor for the production of many hexuronic acid-containing bacterial surface glycostructures. In Escherichia coli K-12, Ugd is important for biosynthesis of the environmentally regulated exopolysaccharide known as colanic acid, whereas in other E. coli isolates, the same enzyme is required for production of the constitutive group 1 capsular polysaccharides, which act as virulence determinants. Recent studies have implicated tyrosine phosphorylation in the activation of Ugd from E. coli K-12, although it is not known if this is a feature shared by bacterial Ugd proteins. The activities of Ugd from E. coli K-12 and from the group 1 capsule prototype (serotype K30) were compared. Surprisingly, for both enzymes, site-directed Tyr → Phe mutants affecting the previously proposed phosphorylation site retained similar kinetic properties to the wild-type protein. Purified Ugd from E. coli K-12 had significant levels of NAD substrate inhibition, which could be alleviated by the addition of ATP and several other nucleotide triphosphates. Mutations in a previously identified UDP-glucuronic acid allosteric binding site decreased the binding affinity of the nucleotide triphosphate. Ugd from E. coli serotype K30 was not inhibited by NAD, but its activity still increased in the presence of ATP.
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Affiliation(s)
- Iain L Mainprize
- Department of Molecular and Cellular Biology, University of Guelph, Guelph, Ontario N1G 2W1, Canada
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Barbas A, Popescu A, Frazão C, Arraiano CM, Fialho AM. Rossmann-fold motifs can confer multiple functions to metabolic enzymes: RNA binding and ribonuclease activity of a UDP-glucose dehydrogenase. Biochem Biophys Res Commun 2012; 430:218-24. [PMID: 23137539 DOI: 10.1016/j.bbrc.2012.10.091] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/17/2012] [Accepted: 10/25/2012] [Indexed: 12/25/2022]
Abstract
Metabolic enzymes are usually characterized to have one specific function, and this is the case of UDP-glucose dehydrogenase that catalyzes the twofold NAD(+)-dependent oxidation of UDP-glucose into UDP-glucuronic acid. We have determined that this enzyme is also capable of participating in other cellular processes. Here, we report that the bacterial UDP-glucose dehydrogenase (UgdG) from Sphingomonas elodea ATCC 31461, which provides UDP-glucuronic acid for the synthesis of the exopolysaccharide gellan, is not only able to bind RNA but also acts as a ribonuclease. The ribonucleolytic activity occurs independently of the presence of NAD(+) and the RNA binding site does not coincide with the NAD(+) binding region. We have also performed the kinetics of interaction between UgdG and RNA. Moreover, computer analysis reveals that the N- and C-terminal domains of UgdG share structural features with ancient mitochondrial ribonucleases named MAR. MARs are present in lower eukaryotic microorganisms, have a Rossmannoid-fold and belong to the isochorismatase superfamily. This observation reinforces that the Rossmann structural motifs found in NAD(+)-dependent dehydrogenases can have a dual function working as a nucleotide cofactor binding domain and as a ribonuclease.
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Affiliation(s)
- Ana Barbas
- Instituto de Tecnologia Química e Biológica/Universidade Nova de Lisboa, Oeiras, Portugal
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Sakuraba H, Kawai T, Yoneda K, Ohshima T. Structure of a UDP-glucose dehydrogenase from the hyperthermophilic archaeon Pyrobaculum islandicum. Acta Crystallogr Sect F Struct Biol Cryst Commun 2012; 68:1003-7. [PMID: 22949183 PMCID: PMC3433186 DOI: 10.1107/s1744309112030667] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/13/2012] [Accepted: 07/04/2012] [Indexed: 06/01/2023]
Abstract
The crystal structure of an extremely thermostable UDP-glucose dehydrogenase (UDP-GDH) from the hyperthermophilic archaeon Pyrobaculum islandicum was determined at a resolution of 2.0 Å. The overall fold was comprised of an N-terminal NAD(+) dinucleotide binding domain and a C-terminal UDP-sugar binding domain connected by a long α-helix, and the main-chain coordinates of the enzyme were similar to those of previously studied UDP-GDHs, including the enzymes from Burkholderia cepacia, Streptococcus pyogenes and Klebsiella pneumoniae. However, the sizes of several surface loops in P. islandicum UDP-GDH were much smaller than the corresponding loops in B. cepacia UDP-GDH but were comparable to those of the S. pyogenes and K. pneumoniae enzymes. Structural comparison revealed that the presence of extensive intersubunit hydrophobic interactions, as well as the formation of an intersubunit aromatic pair network, is likely to be the main factor contributing to the hyperthermostability of P. islandicum UDP-GDH.
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Affiliation(s)
- Haruhiko Sakuraba
- Department of Applied Biological Science, Faculty of Agriculture, Kagawa University, 2393 Ikenobe, Miki-cho, Kita-gun, Kagawa 761-0795, Japan
| | - Tomoyuki Kawai
- Department of Applied Biological Science, Faculty of Agriculture, Kagawa University, 2393 Ikenobe, Miki-cho, Kita-gun, Kagawa 761-0795, Japan
| | - Kazunari Yoneda
- Department of Bioscience, School of Agriculture, Tokai University, Aso, Kumamoto 869-1404, Japan
| | - Toshihisa Ohshima
- Microbial Genetics Division, Institute of Genetic Resources, Faculty of Agriculture, Kyushu University, 6-10-1 Hakozaki, Higashi-ku, Fukuoka 812-8581, Japan
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Eixelsberger T, Brecker L, Nidetzky B. Catalytic mechanism of human UDP-glucose 6-dehydrogenase: in situ proton NMR studies reveal that the C-5 hydrogen of UDP-glucose is not exchanged with bulk water during the enzymatic reaction. Carbohydr Res 2012; 356:209-14. [PMID: 22525098 PMCID: PMC3387377 DOI: 10.1016/j.carres.2012.03.028] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/01/2012] [Revised: 03/23/2012] [Accepted: 03/24/2012] [Indexed: 11/18/2022]
Abstract
Human UDP-glucose 6-dehydrogenase (hUGDH) catalyzes the biosynthetic oxidation of UDP-glucose into UDP-glucuronic acid. The catalytic reaction proceeds in two NAD+-dependent steps via covalent thiohemiacetal and thioester enzyme intermediates. Formation of the thiohemiacetal adduct occurs through attack of Cys276 on C-6 of the UDP-gluco-hexodialdose produced in the first oxidation step. Because previous studies of the related enzyme from bovine liver had suggested loss of the C-5 hydrogen from UDP-gluco-hexodialdose due to keto-enol tautomerism, we examined incorporation of solvent deuterium into product(s) of UDP-glucose oxidation by hUGDH. We used wild-type enzyme and a slow-reacting Glu161→Gln mutant that accumulates the thioester adduct at steady state. In situ proton NMR measurements showed that UDP-glucuronic acid was the sole detectable product of both enzymatic transformations. The product contained no deuterium at C-5 within the detection limit (⩽2%). The results are consistent with the proposed mechanistic idea for hUGDH that incipient UDP-gluco-hexodialdose is immediately trapped by thiohemiacetal adduct formation.
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Affiliation(s)
- Thomas Eixelsberger
- Institute of Biotechnology and Biochemical Engineering, Graz University of Technology, Petersgasse 12/1, A-8010 Graz, Austria
| | - Lothar Brecker
- Institute of Organic Chemistry, University of Vienna, Währingerstraße 38, A-1090 Vienna, Austria
| | - Bernd Nidetzky
- Institute of Biotechnology and Biochemical Engineering, Graz University of Technology, Petersgasse 12/1, A-8010 Graz, Austria
- Corresponding author. Tel.: +43 316 873 8400; fax: +43 316 873 8434.
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