Mechanism of Tgfbr2 in Chondroprotection

Tgfbr2 软骨保护机制

基本信息

批准号：
10394844
负责人：
Rosa A. Serra
金额：
$ 32.34万
依托单位：
UNIVERSITY OF ALABAMA AT BIRMINGHAM
依托单位国家：
美国
项目类别：
财政年份：
2013
资助国家：
美国
起止时间：
2013-04-01 至 2025-03-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10394844
关键词：
Arthritis Binding Binding Proteins Biological Biological Assay Biological Process Bioreactors CRISPR/Cas technology Cartilage Cells Centers for Disease Control and Prevention (U.S.)Chondrocytes Cyclic AMP-Dependent Protein Kinases DNA Degenerative polyarthritis Development Disease Dominant-Negative Mutation Drug Targeting Drug usage Elements Engineering Extracellular Matrix Proteins Funding Future Gel Gene Expression Gene Expression Regulation Genes Genetic Transcription Half-Life Homeostasis Human In Vitro Industrialization Knock-in Knock-out Knowledge Laboratories Luciferases Maintenance Maps Mass Spectrum Analysis Mediating Messenger RNA Modeling Modification Molecular Mus Mutation Pathway interactions Peptides Pharmaceutical Preparations Pharmacology Phenotype Phosphorylation Phosphorylation Site Polysaccharides Post-Translational Modification Site Post-Translational Protein Processing Post-Translational Regulation Proteoglycan Regulation Reporter Role SOX9 protein Serine Signal Pathway Signal Transduction Site Skeletal Development Stains Stream Sulfate Sumoylation Pathway TGFBR2 gene Tissue-Specific Gene Expression Tissues Transforming Growth Factor beta Transgenic Mice Western Blotting articular cartilage cartilage degradation chromatin immunoprecipitation disability drug discovery experimental study genetic manipulation high throughput screening joint function loss of function mutant novel p38 Mitogen Activated Protein Kinase parathyroid hormone-related protein polysulfated glycosaminoglycan prevent response sex skeletal disorder skeletal tissue stable cell line transcription factor

项目摘要

Project Summary. Osteoarthritis (OA) is a leading cause of disability in the industrialized world but there are no approved disease modifying drugs for OA. The long-term objective of my laboratory is to understand the factors that mediate the development and maintenance of articular cartilage so that specific targets to prevent or treat OA can be identified. TGF-ß is a multifunctional peptide that has been shown to regulate skeletal development and tissue-specific gene expression. We were the first to identify TGF-ß as a chondroprotective factor. The importance of TGF-ß in human osteoarthritis has now been well established. We previously identified several down-stream targets of TGF-ß that regulate post-translational processing of the major extracellular matrix proteins in cartilage. One in particular, 3-Prime-Phoshoadenosine 5-Prime-Phosphosulfate Synthase 2 (Papss2), has been associated with osteoarthritis in humans and mice and is required for proper sulfation of glycosamino glycans in cartilage. The transcription factor Sox9 has similar roles in cartilage as TGF-ß and in the last funding period we showed that TGF-ß regulates post-translational modifications on Sox9 and increases the half-life of the Sox9 protein. Furthermore, we showed that Sox9 is sufficient and required for TGF-ß-mediated regulation of Papss2 transcription defining a novel signaling pathway for TGF-ß in cartilage. We also showed, using a bioreactor model, that expression of Sox9 in cartilage deficient for TGF-ß signaling was sufficient to restore proteoglycan staining to that cartilage. In this funding period, we propose to continue to determine the mechanisms by which TGF-ß post-translationally modifies the Sox9 protein and determine the biological consequences of the modifications. We will also determine the molecular mechanism whereby TGF- ß regulates Papss2 expression through a novel Sox9-dependent mechanism. Detailed molecular information about the mechanisms of TGF-ß action in articular cartilage will allow us to develop high throughput assays for future drug discovery screens. We propose the following specific aims: 1A) To determine how TGF-ß regulates the phosphorylation of Sox9 on Serine 181; 1B) Determine the role of Serine 211 in TGF-ß-mediated stabilization of Sox9 protein; 1C) Determine the mechanism whereby TGF-ß treatment results in sumoylation of Sox9; 2) Determine the biological functions of post translationally modified Sox9 3) Identify TGF-ß /Sox9 responsive DNA elements in the Papss2 gene; and 4) Determine if downstream targets of TGF-ß can prevent cartilage degeneration and OA. The information acquired during this project period will provide cartilage unique targets for drug discovery to prevent or treat osteoarthritis.

项目摘要。骨关节炎（OA）是工业化世界中疾病的主要原因，但没有批准的疾病修改OA的药物。我实验室的长期目标是了解介导的因素关节软骨的开发和维护，以便预防或治疗OA的特定目标可以是确定。 TGF-ß是一种多功能肽，已显示用于调节骨骼发育和组织特异性基因表达。我们是第一个将TGF-ß确定为软骨保护因素的人。这 TGF-ß在人骨关节炎中的重要性现已确定。我们以前确定了几个 TGF-ß的下游靶标调节主要细胞外基质的翻译后处理软骨中的蛋白质。尤其是3个p-Phoshoadenosine 5-磷脂硫酸盐合酶2 （PAPSS2），与人类和小鼠的骨关节炎有关软骨中的糖氨基聚糖。转录因子SOX9在软骨中的作用与TGF-ß和最后的资金期我们表明，TGF-ß调节了Sox9和增加Sox9蛋白的半衰期。此外，我们表明Sox9足够了，需要 TGF-β介导的PAPSSS2转录定义了软骨中TGF-ß的新信号通路的调节。我们还使用生物反应器模型显示了Sox9在软骨中的表达缺乏TGF-ß信号传导足以将蛋白聚糖染色恢复到该软骨。在这个资金期间，我们建议继续为了确定TGF-β后翻译修饰Sox9蛋白的机制，并确定修饰的生物学后果。我们还将确定TGF-的分子机制通过一种新型的Sox9依赖性机制来调节PAPSS2的表达。详细的分子信息关于关节软骨中TGF-ß动作的机制将使我们能够为未来的药物发现屏幕。我们提出以下特定目的：1a）确定TGF-ß如何调节 Sox9在丝氨酸181上的磷酸化； 1b）确定丝氨酸211在TGF-ß介导的作用 Sox9蛋白的稳定； 1C）确定TGF-ß处理导致Sumoylation的机制 sox9; 2）确定翻译后修改的生物学功能Sox9 3）识别TGF-ß /SOX9 PAPSSS2基因中的响应DNA元素； 4）确定TGF-ß下游目标是否可以防止软骨变性和OA。在此项目期间获得的信息将提供独特的软骨预防或治疗骨关节炎的药物发现靶标。