TRANSCRIPTIONAL REGULATION BY THE CAM CASCADE

细胞内钙升高是最重要的信号之一 细胞中的分子，尤其是神经组织中的分子。其中一个重要的 钙的作用是调节选定基因的转录， 从而改变细胞的表型。来自多项研究的 实验室，包括我们自己的，已经证明了一个家庭 钙调蛋白依赖的蛋白激酶介导许多钙离子- 依赖转录事件。特别是，我们关注的是 能磷酸化几种反式激活的CaM-Kinase IV CREB和SRF等蛋白质。在过去的两年里，我们有 部分地描述了由CaM-KK级联组成的CaM-KK级联 一种可以磷酸化和激活CaM-Kinase IV和 在目前的拨款申请中，我们将进一步 通过以下方式来表征这一CaM-激酶级联的参与：1) 确定级联的其他上游组件(例如，凸轮- 激酶激酶)或CaM-激酶的锚定蛋白。2) 通过以下方法鉴定CaM-Kinase IV的其他反式激活蛋白 检测CaM对NGFI-B基因转录调控的影响 激酶级联反应。3)确定部位和生理相关性 Cam-Kinase级联和MAP-Kinase之间的串扰 卡斯卡德。 这些研究将在各种培养细胞中进行， 尤其是PC12细胞，并将利用生化和分子 生物技术。这项研究的结果对 钙离子对细胞凋亡和突触可塑性的调控 依赖基因转录。

Elevated intracellular calcium is one of the most important signaling molecules in cells, especially in neural tissues. One of the important actions of calcium is to regulate the transcription of selected genes, thereby altering the phenotype of the cell. Studies from a number of laboratories, including our own, have demonstrated that a family of calmodulin-dependent protein kinases mediate many of these Ca2+- dependent transcriptional events. In particular, we have focused on CaM-kinase IV which can phosphorylate several transactivating proteins such as CREB and SRF. In the past two years we have partially characterized a CaM-kinase cascade consisting of a CaM- kinase kinase which can phosphorylate and activate CaM-kinase IV and CaM-kinase I. In the current grant application we will further characterize the involvement of this CaM-kinase cascade by: 1) identifying additional upstream components of the cascade (e.g. CaM- kinase kinase kinase) or anchoring proteins for CaM-kinase kinase. 2) identifying additional transactivating proteins for CaM-kinase IV by examining transcriptional regulation of the NGFI-B gene by the CaM- kinase cascade. 3) determining the sites and physiological relevance of cross-talk between the CaM-kinase cascade and the MAP-kinase cascade. These studies will be performed in a variety of cultured cells, especially PC12 cells, and will utilize biochemical and molecular biological techniques. The results of this study have implications for modulating cellular apoptosis and synaptic plasticity due to Ca2+- dependent gene transcription.