ROLE OF CDK2 CELL CYCLE SIGNALING IN ISCHEMIC INJURY AND PROTECTION

主要研究者/项目负责人（最后，第一，中间）：Ping，Peipei（MacLellan，项目4） 本计划项目申请的主题是了解信号转导通路介导的 缺血性损伤和心脏保护，使用多学科方法结合生物物理学，生理学， 蛋白质组学和遗传学。虽然项目1 - 3集中在两个信号通路以前已知的调节 缺血性损伤和保护（即，PKCe和p38 MAPK），项目4关注的是一个最近被认可的通路， 在心肌缺血领域的重要性细胞周期调节蛋白，特别是，它将集中在三个 在该途径中功能相互关联的分子：细胞周期蛋白依赖性激酶2（Cdk 2）， 视网膜母细胞瘤基因产物（Rb）和调节细胞周期基因的转录因子（E2Fs 包括细胞周期蛋白A和E，Cdk2的催化伙伴。 项目4中提出的研究得到以下支持：1）最近的证据表明， 心脏伴随着包括Cdk 2在内的许多细胞周期调节蛋白的上调;以及2）通过 惊人的初步数据表明，心肌梗死面积增加Rb-null心肌受到区域性 缺血性损伤相反，体内用Cdk2抑制剂治疗阻断了Cdk2活性的预期增加 缺血性损伤，并导致梗死面积减少。这表明Cdk2在缺血性脑损伤中的关键作用。 损伤和Rb蛋白对损伤的心脏保护作用。尽管这些观察结果很有趣， 这些作用的细胞机制尚不清楚。 与项目1 - 3和核心项目合作，项目4提出了3个目标。目标1将阐明 Cdk2调节缺血性损伤能力的潜在机制;与心脏生物学核心合作， 目的是确定Cdk2如何调节线粒体功能和细胞死亡途径; 蛋白质组学核心，目标1将确定新的Cdk2底物。目标2将研究Rb的分子基础 在缺血性损伤中的心脏保护作用，它将决定Rb的p38 a MAPK依赖性调节，并将确定 E2F家族（Rb的主要靶点）在这些过程中的作用。最后，目标3将阐明机制， PKCe在心脏保护中调节Cdk2活性。这些研究将与项目合作进行 2和心脏生物学核心，并将采用两种完善的心脏保护小鼠模型：PKCe 转基因和一氧化氮供体诱导的晚期预适应。拟议的调查将 为理解Rb心脏保护作用的分子基础和机制做出了重要贡献 Cdk2调节缺血性损伤和凋亡性细胞死亡的能力。

Principal Investigator/ProgramDirector (Last, First, Middle): Ping, Peipei (MacLellan,Project 4) The theme of this Program Project application is to understand the signal transduction pathways mediating ischemic injury and cardioprotection using a multidisciplinary approach combining biophysics, physiology, proteomics and genetics. While Projects 1-3 are focused on two signaling pathways previously known to modulate ischemic injury and protection (i.e., PKCe and p38 MAPK), Project 4 focuses on a pathway with recently recognized importance in the field of myocardial ischemiacell cycle regulatory proteins, in particular, it will focus on three molecules whose functions are interrelated in this pathway: the cyclin-dependent kinase-2 (Cdk2), the retinoblastoma gene product (Rb), and the transcription factors (E2Fs) that regulate genes responsible for cell cycle entry including Cyclin A and E, the catalytic partners of Cdk2. Studies proposed in Project 4 are supported 1) by recent evidence demonstrating that ischemic injury to the heart are accompanied by the upregulation of a number of cell cycle regulatory proteins including Cdk2; and 2) by striking preliminary data demonstrating that infarct size was increased in Rb-null myocardium subjected to regional ischemic injury. Conversely, treatment with a Cdk2 inhibitor in vivo blocked the expected increase in Cdk2 activity with ischemic injury and led to a reduced infarct size formation. This suggests a critical role for Cdk2 in ischemic injury and a cardioprotective role of the Rb protein against injury. Although these observations are intriguing, as the cellular mechanisms underlying these effects are unknown. In collaboration with Projects 1-3 and the Cores, Project 4 proposes 3 Aims. Aim 1 will elucidate the mechanisms underlying Cdk2's ability to modulate ischemic injury; In collaboration with the Heart Biology Core, this aim will determine how Cdk2 modulates mitochondrial function and the cell death pathways; In collaboration with the Proteomic Core, Aim 1 will identify novel Cdk2 substrates. Aim 2 will examine the molecular basis forRb's cardioprotective role in ischemic injury, it will determine p38a MAPK dependent modulation of Rb, and will define the role of E2F family (theprimary targets of Rb) in these processes. Finally, Aim 3 will elucidate mechanisms by which PKCe regulates Cdk2 activity in cardioprotection. These studies will be performed in collaboration with Project 2 and the Heart Biology Core, and will employ two well-established murine models of cardioprotection: the PKCe transgenesis and the nitric oxide donor induced late phase of preconditioning. The proposed investigations will make key contributions to understanding the molecular basis for Rb's cardioprotective effect and the mechanism underlying Cdk2's ability to regulate ischemic damage and apoptotic cell death.