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: 10534104
• 负责人: Andrew Bruce Lassar
• 金额: $ 67.73万
• 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/10534104
• 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; Lentivirus; 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; Repression; Rheumatoid Arthritis; Role; Signal Pathway; Signal Transduction; Synovial joint; Tissues; Transforming Growth Factor Beta 2; Work; articular cartilage; bZIP Domain; inducible Cre; insight; joint formation; knock-down; long bone; lubricin; novel; p38 Mitogen Activated Protein Kinase; postnatal; 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.

项目概要/摘要。 该项目的广泛，长期目标是全面了解发展 决定滑膜关节形成中细胞命运的事件。在这一广阔的领域内，这项建议 重点是关节软骨的分化，这在维持低摩擦中起着核心作用。 关节空间的环境。事实上，组成关节软骨的细胞的标志是它们的表达 蛋白聚糖，如蛋白质润滑素，由Prg 4基因编码，润滑关节和保护 防止关节炎的发展。Prg 4特异性表达于关节软骨的最表层， 软骨Lassar实验室和其他人的发现已经确定，在表皮细胞中表达Prg 4的细胞， 关节软骨区（在胚胎和出生后早期的小鼠中）作为所有更深处的干细胞群， 成人关节软骨的区域。因此，为了阐明关节软骨干细胞群如何 是在发育过程中产生的，并在成人中维持，本提案的一个关键目标是确定 调节Prg 4和其他基因表达的因子，这些基因特异性地表达在细胞中。 关节软骨的浅表区。拉萨尔实验室的最新发现表明， Creb 5在浅区关节软骨细胞中独特地表达（与深区关节软骨细胞和浅区关节软骨细胞相反）。 软骨细胞和生长板软骨细胞），并且是Prg 4表达的关键调节剂。最明显的是， Creb 5在深层牛关节软骨细胞（不表达Prg 4）中的表达使TGF-β 2 EGFR信号诱导Prg 4在这些细胞中表达，达到与表浅区表达的水平相等的水平 关节软骨细胞这些发现表明，Creb 5在软骨细胞中建立了一种能力状态， 表达Prg 4以响应这些信号通路。此外，拉萨尔实验室发现， 缺乏功能性Creb 5不能形成许多滑膜关节， 芽间充质（具有Prx 1-Cre）导致长骨中生长板发育的严重丧失。采取 总之，这些发现表明Creb 5在滑膜关节的形成中起着关键作用， 两者都是关节软骨细胞中Prg 4/润滑素表达的新的和关键的调节剂。该项目将确定 Creb 5依赖性基因和驱动这些基因表达的调控元件在原发性肝癌中的表达， 牛浅区关节软骨细胞;并机械地确定TGF β，EGFR和p38 信号转导调节这些细胞中Creb 5的转录活性。此外，该项目将确定 Creb 5如何调节滑膜关节的形成，并阐明Creb 5在骨骺中的表达 软骨膜阻止生长板延伸到长骨的骺端。