Molecular Analysis of Metalloprotease Disintegrin ADAM12

描述（由申请人提供）：亚当斯是一个含有去整合素和金属蛋白酶结构域的细胞表面蛋白家族，在涉及细胞表面蛋白水解、细胞-细胞或细胞-基质相互作用的许多生物学过程中发挥重要作用。我们的长期目标是了解ADAM蛋白的功能，并剖析它们在跨膜信号传导中的作用。目前，我们的研究重点是ADAM 12，ADAM家族成员，参与骨骼肌发育和/或再生。我们的初步结果表明，ADAM 12在体外成肌细胞分化过程中的GO期（静止）的诱导中起作用。在体外成肌分化过程中形成的静止的未分化细胞与体内的肌卫星细胞具有一些共同的特征。卫星细胞在肌肉再生、肌肉肥大和出生后肌肉生长过程中起着关键作用，但骨骼肌卫星细胞自我更新的机制尚不清楚。该提案的目标是了解ADAM 12在GO进入肌肉细胞过程中的作用。我们推测，ADAM 12诱导进入静止的机制涉及PI 3 K活性的下调，并依赖于ADAM 12介导的细胞-细胞相互作用。为了验证我们的假设，我们将进行一系列研究，以实现以下具体目标。在目的1中，我们将表征与细胞周期停滞相关的分子事件和导致ADAM 12在成肌细胞系和原代成肌细胞中上调静止细胞标志物（视网膜母细胞瘤相关蛋白p130和细胞周期抑制剂p27）的事件序列。在目的2中，我们将全面分析ADAM 12结合对培养成肌细胞中PI 3 K活性的影响。在目标3中，我们将研究ADAM 12的胞外亚结构域：去整合素，富含半胱氨酸和EGF样区，在ADAM 12诱导的细胞周期阻滞和上调p130和p27中的作用。我们的研究结果可能有助于了解卫星细胞的生物学及其在肌肉生长和修复中的作用。

DESCRIPTION (provided by applicant): ADAMs, a family of cell surface proteins containing a disintegrin and metalloprotease domains, play important roles in many biological processes involving cell surface proteolysis, cell-cell, or cell-matrix interactions. Our long-term goal is to understand the function of ADAM proteins and to dissect their roles in transmembrane signaling. Currently, we focus our studies on ADAM12, an ADAM family member that is involved in skeletal muscle development and/or regeneration. Our preliminary results suggest that ADAM12 plays a role in the induction of GO phase (quiescence) during myoblast differentiation in vitro. Quiescent, undifferentiated cells formed during myogenic differentiation in vitro share several characteristics with muscle satellite cells in vivo. Satellite cells play a pivotal role during muscle regeneration, muscle hypertrophy, and post-natal muscle growth, but the mechanism of self-renewal of the satellite cell compartment in skeletal muscle is poorly understood. The goal of this proposal is to understand the role of ADAM12 during GO entry in muscle cells. We hypothesize that the mechanism by which ADAM12 induces the entry into quiescence involves down-regulation of PI3K activity and depends on ADAM12-mediated cell-cell interactions. To test our hypothesis, we will perform a series of studies that will pursue the following Specific Aims. In Aim 1, we will characterize the molecular events associated with cell cycle arrest and the sequence of events leading to up-regulation of quiescent cell markers (retinoblastoma-related protein p130 and cell cycle inhibitor p27) by ADAM12 in myoblastic cell lines and in primary myoblasts. In Aim 2, we will perform a comprehensive analysis of the effect of ADAM12 binding on the activity of PI3K in cultured myoblasts. In Aim 3, we will examine the role of the extracellular sub-domains of ADAM12: disintegrin, cysteine-rich, and EGF-like region, in ADAM12-induced cell cycle arrest and up-regulation of p130 and p27. The results of our studies may help understand the biology of satellite cells and their role in muscle growth and repair.