Role of Transcription Factor ERG in Skeletogenesis

描述(申请人提供)：在四肢骨骼发育过程中，软骨细胞遵循两种发育途径，形成永生的关节软骨或短暂的生长板软骨，在此过程中细胞成熟、肥大，并被骨取代。虽然这种发育分叉对于骨骼形成和功能的关键重要性已经被广泛认识，但潜在的调节机制仍然不清楚，特别是在分子水平上。在之前的资助期间，我们重点研究了转录因子ERG及其变异体C-1-1。用鸡和人的C-1-1进行的功能获得研究表明：(A)C-1-1能够在整个肢体软骨细胞群中施加稳定的关节样表型，阻止生长板的形成、软骨细胞的成熟和骨的形成；(B)C-1-1中和促成熟转录因子Runx2的作用；以及(C)关节主控调控因子GDF-5快速诱导ERG/C-1-1在发育中的滑膜关节中的表达。这些和其他发现导致了我们对这一竞争性延续方案的中心假设：(A)C-1-1作用于GDF-5的下游有助于永久关节软骨细胞的形成；以及(B)C-1-1反过来抑制Runx2功能，维持细胞的永久状态，并防止成熟和肥大。我们的目标是：(I)通过细胞、外植体培养和转基因方法对小鼠ERG变体进行功能鉴定；(Ii)利用GDF-5-Cre和GDF-5-Creer小鼠，确定有条件地去除ERG基因在发育或出生后生活中的后果；以及(Iii)确定GDF-5通过信号通路和启动子作用触发ERG表达的机制，以及ERG/C-1-1如何抑制Runx2功能。该项目将对关节软骨细胞的起源和功能产生基本的见解。它还应该产生退行性关节疾病的小鼠模型，可以用来测试未来针对骨关节炎患者和老年患者常见的关节疾病的基因和细胞疗法。

DESCRIPTION (provided by applicant): During limb skeletogenesis chondrocytes follow two developmental paths and produce permanent articular cartilage persisting through life or transient growth plate cartilage in which the cells mature, hypertrophy, and are replaced by bone. Though the critical importance of this developmental bifurcation for skeletal formation and function is widely recognized, the underlying mechanisms of regulation remain unclear, particularly at the molecular level. In the previous funding period, we focused on the transcription factor ERG and its variant C-1-1. Gain-of-function studies with chick and human C-1-1 show that: (a) C-1-1 is able to impose a stable and articular-like phenotype over the entire limb chondrocyte population, blocking growth plate formation, chondrocyte maturation and bone formation; (b) C-1-1 counteracts action of the pro-maturation transcription factor Runx2; and (c) the joint master regulator GDF-5 rapidly induces ERG/C-1-1 expression in developing synovial joints. These and other findings lead to our central hypotheses for this competitive continuation proposal: (a) C-1-1 acting down-stream of GDF-5 contributes to formation of permanent articular chondrocytes; and (b) C-1-1 in turn inhibits Runx2 function, maintains the permanent status of the cells, and prevents maturation and hypertrophy. Our aims are: (i) to functionally characterize murine ERG variants by cell and explant cultures and transgenic approaches; (ii) determine the consequences of conditional ERG gene ablation during development or postnatal life, using GDF-5-Cre and GDF-5-CreER mice; and (iii) determine the mechanisms by which GDF-5 triggers ERG expression by signaling pathways and promoter action, and how ERG/C-1-1 inhibits Runx2 function. The project will produce fundamental insights into genesis and function of articular chondrocytes. It should also generate mouse models of degenerative joint disease that could be used to test future gene- and cell-based therapies for joints conditions common to osteoarthritic patients and aging individuals.