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REGULATION OF SKELETAL MUSCLE DEVELOPMENT AND MAINTENANCE BY PROTEIN O-GLUCOSYLTRANSFERASE 1 (POGLUT1)

蛋白质 O-葡萄糖基转移酶 1 (POGLUT1) 调节骨骼肌发育和维持

基本信息

批准号：
10212971
负责人：
Radbod Darabi
金额：
$ 46.26万
依托单位：
BAYLOR COLLEGE OF MEDICINE
依托单位国家：
美国
项目类别：
财政年份：
2020
资助国家：
美国
起止时间：
2020-07-08 至 2025-06-30
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10212971
关键词：
Adult Affect Alleles Animals Biochemical Biological Biological Assay Biology CRISPR/Cas technology Carbohydrates Cell Communication Cell Culture Techniques Cell Surface Proteins Cell model Cells Complex Consensus Sequence Data Defect Development Disease Disease model Drosophila genus Embryonic Development Engraftment Enzymes Epidermal Growth Factor Family Functional disorder Future Genes Genetic Diseases Glucose Glucosyltransferase Growth Homing Human Immunodeficient Mouse Impairment In Vitro Injury Knock-in Knockout Mice Limb-Girdle Muscular Dystrophies Link Maintenance Mammals Mediating Molecular Mus Muscle Muscle Cells Muscle Development Muscle satellite cell Muscular Dystrophies Mutation Myoblasts Myopathy NOTCH3 gene Notch Signaling Pathway Pathology Pathway interactions Patients Phenotype Physiology Play Polysaccharides Post-Translational Protein Processing Process Proliferating Protein Glycosylation Proteins Proteomics Regulation Reporting Role Siblings Signal Transduction Site Skeletal Muscle Structure Techniques Testing Therapeutic Transgenes base body system c new causal variant cell type conditional knockout consanguineous family experimental study fly glycosylation glycosyltransferase human disease in vivo induced pluripotent stem cell insight migration mouse genetics muscle form mutant myogenesis notch protein novel novel therapeutic intervention postnatal progenitor receptor repaired response satellite cell self-renewal skeletal stem cells therapeutic target

项目摘要

SUMMARY Carbohydrate addition (glycosylation) is one of the most common posttranslational modifications of secreted and cell surface proteins. Glycosylation is critical for normal animal development and physiology, and mutations in genes involved in glycosylation cause more than a 100 human diseases with diverse phenotypes. However, glycan structures are complex, and each form of glycosylation can be found in tens to thousands of target proteins. Accordingly, understanding the molecular mechanisms underlying glycosylation disorders constitute a major challenge. One of the critical roles played by protein glycosylation is the regulation of a cell-to-cell communication mechanism called the Notch signaling pathway. Notch signaling regulates many processes during animal development and adult maintenance. For example, studies in mice and cell culture have shown that muscle development and muscle repair after injury depend on Notch signaling in mammals. However, mutations in Notch pathway components or modulators have not been reported in human patients with muscular dystrophy. We have recently reported a consanguineous family in which several siblings suffer from LGMD-2Z, which is a new form of limb-girdle muscular dystrophy. LGMD-2Z is caused by homozygosity for a recessive mutation in a gene called POGLUT1. POGLUT1 is a glycosyltransferase which adds O-linked glucose to a number of transmembrane and secreted proteins, including multiple components of the Notch signaling pathway. We have previously shown that POGLUT1 regulates Notch signaling in fruit flies and mice. Analysis of the muscle tissues and myoblasts isolated from the above-mentioned patients provided evidence suggesting that impaired Notch signaling plays an important role in the pathophysiology of this form of muscular dystrophy. However, the biologically-relevant targets of POGLUT1 in the Notch pathway and other pathways in the muscle are not known. In this proposal, we will use biochemical and cell culture assays, proteomic profiling, mouse genetic experiments and iPS cell experiments to determine the molecular mechanisms underlying the regulation of muscle development and maintenance by POGLUT1 and to identify its relevant targets. We will use iPS cells from patients, along with a CRISPR/Cas9-mediated corrected version of them, for in vitro disease modeling and in vivo engraftment experiments. These studies have the potential to provide novel insight into the pathophysiology of a muscular dystrophy caused by abnormal glycosylation and might establish a new framework for future therapeutic approaches for muscle diseases.

摘要糖加成(糖基化)是最常见的翻译后修饰之一分泌蛋白和细胞表面蛋白。糖基化对动物的正常发育和生理学和涉及糖基化的基因突变导致人类100多种疾病具有多样的表型。然而，多糖的结构是复杂的，并且每种形式的糖基化可以在数万到数千种目标蛋白质中找到。因此，理解分子糖基化紊乱的潜在机制构成了一个重大挑战。其中一个关键角色蛋白质糖基化作用是细胞间通讯机制的调节，称为 Noch信号通路。Notch信号在动物发育和发育过程中调节许多过程成人赡养费。例如，对老鼠和细胞培养的研究表明，肌肉哺乳动物肌肉损伤后的发育和修复依赖于Notch信号。然而， Notch通路组件或调节物的突变在人类患者中尚未报道患有肌肉营养不良。我们最近报道了一个血缘关系密切的家庭，其中几个人兄弟姐妹患有LGMD-2Z，这是一种新的肢体带状肌营养不良症。LGMD-2Z是由POGLUT1基因的隐性突变纯合子引起。POGLUT1是一种糖基转移酶，将O-连接的葡萄糖添加到一些跨膜并分泌蛋白质，包括Notch信号通路的多个组成部分。我们之前已经展示了 POGLUT1调控果蝇和小鼠的Notch信号。肌肉组织和组织的分析从上述患者中分离出的成肌细胞提供的证据表明， Notch信号在这种类型的肌营养不良的病理生理学中起着重要作用。然而，POGLUT1在Notch途径和其他途径中的生物相关靶点目前尚不清楚这些肌肉。在这项提案中，我们将使用生化和细胞培养分析，蛋白质组图谱、小鼠遗传学实验和iPS细胞实验确定分子 POGLUT1和TO调节肌肉发育和维持的机制确定其相关目标。我们将使用来自患者的iPS细胞，以及CRISPR/Cas9介导的它们的更正版本，用于体外疾病建模和体内植入实验。这些研究有可能为肌营养不良症的病理生理学提供新的见解。由糖基化异常引起，并可能为未来的治疗建立一个新的框架肌肉疾病的治疗方法。