Identification of Mechanoresponsive Promoter Elements in Chondrogenesis

软骨形成中机械反应启动子元件的鉴定

• 批准号: 8461068
• 负责人: Dominik R Haudenschild
• 金额: $ 7.32万
• 依托单位: UNIVERSITY OF CALIFORNIA AT DAVIS
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-04-01 至 2016-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8461068
• 关键词: 3-Dimensional; Aging; Apoptotic; Arthritis; Base Pairing; Beryllium; Binding; Binding Proteins; Bioinformatics; Biological Assay; Biological Models; Bone Marrow Stem Cell; Cartilage; Cartilage Matrix; Cell Death; Cell Differentiation process; Cell membrane; Chondrocytes; Chondrogenesis; Code; Collagen Fibril; Collagen Gene; Collagen Type II; Complex; Cytoskeleton; DNA; DNA Binding; DNA-Binding Proteins; Data; Databases; Disease; Elements; Engineering; Ensure; Event; Extracellular Matrix; Focal Adhesions; Foundations; Functional disorder; Future; Gene Activation; Gene Targeting; Genes; Genetic Transcription; Goals; Grant; Growth; Human; Hybrids; Indium; Integrins; Knowledge; Lead; Length; Luciferases; Maintenance; Measurement; Mechanical Stimulation; Mechanics; Mediating; Mediator of activation protein; Membrane Proteins; Modeling; Osteogenesis; Outcome; Pathway interactions; Phosphotransferases; Physiological; Planet Mars; Process; Promoter Regions; Proteins; Regulatory Element; Reporting; Response Elements; Signal Transduction; Stem cells; Stimulus; Testing; Tissue Engineering; Tissues; Transcriptional Activation; Transcriptional Regulation; Transduction Gene; Work; Yeasts; aggrecan; base; bone; cDNA Library; cartilage repair; clinical application; improved; in vivo Model; inhibitor/antagonist; insight; lubricin; mammalian COMP; novel strategies; progenitor; promoter; protein function; response; small molecule; stem; transcription factor

DESCRIPTION (provided by applicant): Cartilage is constantly subjected to mechanical forces including compression, tension, and shear. These forces are converted into physiological responses that are critical to cartilage ma trix composition, cellular survival, and progenitor cell differentiation. Cellular responses to me- chanical forces vary greatly, and can range from proliferation, to differentiation, to apoptotic cell death, and from matrix synthesis to enzymatic matrix degradation. For the proper maintenance of cartilage tissue, the appropriate cellular response must be invoked. Inappropriate cellular re- sponse to mechanical loading contributes to pathological outcomes during growth and in aging. A clearer understanding of the mechanotransduction pathway has tremendous potential in many clinical applications. Traditional studies of mechanotransduction events have focused on cell membrane-associated proteins and cytosolic signal transduction proteins. These approaches have successfully identi- fied integrins, focal adhesion complexes, the cytoskeleton, and select kinase pathways as me- diators of mechanotransduction. However, we have still not identified the DNA elements in a gene promoter that confer the mechanoresponsiveness to the gene. Target genes of mechanotransduction pathways in cartilage include extracellular matrix genes such as Aggrecan and type II collagen and Cartilage Oligomeric Matrix Protein (COMP). COMP functions in the proper assembly of collagen fibrils in cartilage. COMP is expressed by chondro- cytes in endochondral bone formation, and continues to be expressed chondrocytes of mature cartilage. Importantly, COMP transcription is highly sensitive to mechanical stimulation both in chondrocytes and bone marrow stem cells. Our preliminary data demonstrates that the proximal 3kb of the human COMP promoter is suffi- cient to activate COMP transcription in response to mechanical stimulation. In this study we will identify the mechanoresponsive DNA element within the 3kb human COMP promoter. We ex- pect to identify new pathways that mediate the cellular response to mechanical forces, and we anticipate that the COMP mechanoresponse pathways will be conserved in other genes. This project is an integral part of our long-term strategy to gain insight into mechanotransduction pathways and improve our tissue engineering and cartilage repair abilities. Future goals directly stemming from this work include 1) identification of small molecule inhibitors/activators of the mechanoresponse element, 2) identification of additional mechanoresponsive genes and path- ways, and 3) an in vivo model that allows non-destructive measurement of mechanotransduc- tion activity.

描述（由申请人提供）：肱骨持续承受机械力，包括压缩、拉伸和剪切。这些力被转化为对软骨基质组成、细胞存活和祖细胞分化至关重要的生理反应。细胞对机械力的反应变化很大，从增殖到分化，再到凋亡性细胞死亡，从基质合成到酶促基质降解。为了正确维护软骨组织，必须调用适当的细胞反应。细胞对机械负荷的不适当反应导致生长和衰老过程中的病理结果。更清楚地了解机械传导途径在许多临床应用中具有巨大的潜力。传统的力学信号转导研究主要集中在细胞膜相关蛋白和胞浆信号转导蛋白。这些方法已经成功地鉴定了整合素、粘着斑复合物、细胞骨架和选择激酶途径作为机械转导的介质。然而，我们仍然没有确定基因启动子中赋予基因机械反应性的DNA元件。软骨中机械传导途径的靶基因包括细胞外基质基因，例如聚集蛋白聚糖和II型胶原以及软骨寡聚基质蛋白（COMP）。COMP在软骨中胶原纤维的正确组装中起作用。COMP在软骨内骨形成中由软骨细胞表达，并且在成熟软骨的软骨细胞中继续表达。重要的是，COMP转录在软骨细胞和骨髓干细胞中对机械刺激高度敏感。我们的初步数据表明，人COMP启动子的近端3 kb足以激活COMP转录以响应机械刺激。在这项研究中，我们将确定3 kb的人COMP启动子内的机械响应DNA元件。我们期望确定介导细胞对机械力反应的新途径，并且我们预计COMP机械反应途径将在其他基因中保守。该项目是我们长期战略的一个组成部分，以深入了解机械转导途径，提高我们的组织工程和软骨修复能力。直接源自该工作的未来目标包括1）鉴定机械应答元件的小分子抑制剂/活化剂，2）鉴定另外的机械应答基因和途径，和3）允许机械转导活性的非破坏性测量的体内模型。

项目成果

Effect of age and cytoskeletal elements on the indentation-dependent mechanical properties of chondrocytes.

• DOI: 10.1371/journal.pone.0061651
• 发表时间: 2013
• 期刊: PloS one
• 影响因子: 3.7
• 作者: Chahine NO;Blanchette C;Thomas CB;Lu J;Haudenschild D;Loots GG
• 通讯作者: Loots GG