Elucidation of the Role of Creb5 in Synovial Joint Formation

阐明 Creb5 在滑膜关节形成中的作用

• 批准号: 10020759
• 负责人: Andrew Bruce Lassar
• 金额: $ 68.07万
• 依托单位: HARVARD MEDICAL SCHOOL
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-09-18 至 2024-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10020759
• 关键词: ATAC-seq; Adult; Alleles; Area; Arthritis; Cattle; Cells; Chondrocytes; Competence; Coupled; DNA Binding Domain; Degenerative polyarthritis; Development; Ectopic Expression; Embryo; Engineering; Environment; Epidermal Growth Factor Receptor; Epiphysial cartilage; Event; Exons; Fibroblasts; Friction; Gene Expression; Genes; Genetic Recombination; Genetic Transcription; Goals; Guide RNA; Hallmark Cell; Induced Mutation; Infection; Injury; Joints; Knee joint; Limb Bud; Maintenance; Mechanics; Mediating; Meniscus structure of joint; Mesenchyme; Mus; Natural regeneration; Newborn Infant; Physiologic Ossification; Play; Protein Binding Domain; Proteins; Proteoglycan; Regulatory Element; Rheumatoid Arthritis; Role; Signal Pathway; Signal Transduction; Subfamily lentivirinae; Synovial joint; Tissues; Transforming Growth Factor Beta 2; Work; articular cartilage; bZIP Domain; insight; joint formation; knock-down; long bone; lubricin; novel; p38 Mitogen Activated Protein Kinase; programs; protein expression; response; small hairpin RNA; stem cell population; tissue regeneration; transcription factor; transcriptome sequencing

Project Summary/Abstract. The broad, long-term goal of this project is to develop a comprehensive understanding of the developmental events that dictate cell fate decisions in formation of the synovial joint. Within this broad area, this proposal focuses on the differentiation of articular cartilage, which plays a central role in maintaining the low-friction environment of the joint space. Indeed, a hallmark of cells comprising the articular cartilage is their expression of proteoglycans, such as the protein lubricin, encoded by the Prg4 gene, that lubricates the joint and protects against the development of arthritis. Prg4 is specifically expressed in the superficial-most layer of the articular cartilage. Findings by both the Lassar lab and others have established that Prg4-expressing cells in the superficial zone of articular cartilage (in embryonic and early post-natal mice) serve as a stem cell population for all deeper regions of the articular cartilage in the adult. Thus, to elucidate how the articular cartilage stem cell population is both generated during development and maintained in the adult, a key objective of this proposal is to identify the factors that regulate the expression of both Prg4 and other genes that are specifically expressed in the superficial zone of the articular cartilage. Recent findings in the Lassar lab indicate that the transcription factor Creb5 is uniquely expressed in superficial zone articular chondrocytes (as opposed to both deeper zone articular chondrocytes and growth plate chondrocytes) and is a crucial regulator of Prg4 expression. Most notably, ectopic expression of Creb5 in deep zone bovine articular chondrocytes (which do not expression Prg4) enabled TGF-b2 and EGFR signals to induce Prg4 expression in these cells, to a level equal to that expressed by superficial zone articular chondrocytes. These findings suggest that Creb5 establishes a competent state in chondrocytes to express Prg4 in response to these signaling pathways. In addition, the Lassar lab has found that mice engineered to lack functional Creb5 fail to form many synovial joints, and that mis-expression of Creb5 throughout the limb bud mesenchyme (with Prx1-Cre) results in a profound loss of growth plate development in long bones. Taken together, these findings indicate that Creb5 plays a critical role in both the formation of synovial joints and is a both a novel and crucial regulator of Prg4/lubricin expression in articular chondrocytes. This project will identify both Creb5-dependent genes and the regulatory elements that drive the expression of these genes in primary bovine superficial zone articular chondrocytes; and mechanistically determine how TGFb, EGFR, and p38 signaling modulate the transcriptional activity of Creb5 in these cells. In addition, this project will determine how Creb5 regulates the formation of synovial joints, and elucidate how Creb5 expression in the epiphyseal perichondrium blocks extension of the growth plate into the epiphyseal ends of the long bones.

项目摘要/摘要。 这个项目的广泛、长期的目标是发展对发展的全面理解 在滑膜关节的形成中决定细胞命运的事件。在这一广泛的领域内，这项提议 着重于关节软骨的分化，这在维持低摩擦中起着核心作用 关节空间的环境。事实上，构成关节软骨的细胞的一个特征就是它们的表达。 一种蛋白多糖，如由Prg4基因编码的蛋白润滑素，它能润滑关节并保护关节 抗关节炎的发展。Prg4在关节最浅层特异表达 软骨。Lassar实验室和其他实验室的研究发现，Prg4表达的细胞在浅表 关节软骨带(在胚胎和出生后早期的小鼠)是所有深层次的干细胞群。 成人关节软骨的区域。因此，为了阐明关节软骨干细胞群体是如何 既是在发育过程中产生的，也是在成人中保持的，本提案的一个关键目标是识别 调节Prg4和其他基因表达的因子 关节软骨的浅表区。Lassar实验室的最新发现表明，转录因子 Creb5仅在浅层关节软骨细胞中表达(与深层关节软骨细胞表达相反 软骨细胞和生长板软骨细胞)，是Prg4表达的关键调节因子。最值得注意的是，异类 深部区牛关节软骨细胞(不表达Prg4)中Creb5的表达可激活转化生长因子-b2 和EGFR信号诱导Prg4在这些细胞中的表达，达到与表面区表达相同的水平 关节软骨细胞。这些发现表明，Creb5在软骨细胞中建立了一种胜任状态 表达Prg4以响应这些信号通路。此外，拉萨实验室还发现，通过基因工程改造的小鼠 缺乏功能的Creb5不能形成许多滑膜关节，并且Creb5在肢体中的错误表达 芽间充质细胞(带有PRX1-Cre)导致长骨生长板发育严重丧失。已被占用 总之，这些发现表明，Creb5在滑膜关节的形成中起着关键作用，是一种 在关节软骨细胞中Prg4/润滑素表达的一个新的和关键的调节因子。该项目将确定 依赖于Creb5的基因和驱动这些基因表达的调控元件 并从机制上决定了TGFb、EGFR和p38 信号调节这些细胞中Creb5的转录活性。此外，该项目将确定 Creb5如何调控滑膜关节的形成，并阐明Creb5在骨痂中的表达 软骨膜阻止生长板延伸到长骨的骨骺末端。