NEUROENDOCRINE REGULATION OF CREB BY CAMP AND CALCIUM

cAMP和Ca 2+信号通路在整个细胞中使用。 神经内分泌系统调节分泌和基因表达。它有 最近表明，cAMP和Ca 2+可以收敛到调节 单个转录因子CREB的活性。这种趋同可能 允许CREB整合多种细胞外信号， 转录。推测Ca 2+对CREB激活的影响是 由Ca 2 +/钙调蛋白依赖性蛋白激酶（CaM激酶）介导。 该实验室最近的研究表明， 激酶对CREB的激活有非常不同的影响。在 特别地，我们发现CaM激酶II可以抑制CREB的激活， 而CaM激酶IV则能激活CREB。拟议的研究旨在 进一步表征特异性CaM激酶调节 激活CREB/ATF转录因子家族。具体目标 包括：i）检验CREB的Ser 142磷酸化是 生理调节。这将涉及抗体的制备 其检测Ser 142的磷酸化。2)确定抑制剂是否 也观察到CaMK II对ATF 1的影响，ATF 1是一种相关的亮氨酸拉链， 转录因子，这也被证明是响应的变化， cAMP和Ca 2+。CREB负调控位点周围区域 在Ser 142是保守的，但在ATF 1中不相同。3)考查能力 激活CREB。有可能是CaM激酶， CaMKIV可激活CREB。我们将使用转染试验来确定是否 CaMKI可激活CREB。4)检验抑制性的 CaMKII对CREB激活的影响是由于 在Ser 142磷酸化以阻断CREB和co- 激活蛋白，CREB结合蛋白（CBP）。5)映射CREB的部分 和Cl 3 P，其是高亲和力、PKA依赖性和CaM 激酶诱导的相互作用。6)形成游离型cAMP反应性 报告基因检测cAMP和Ca 2+对细胞内Ca ~（2+）相互作用的影响。 具有染色质结构的CREB。这些研究应该提供新的见解 进入允许cAMP和Ca 2+调节细胞膜成员的机制。 CREB/ATF家族。

The cAMP and Ca2+ signaling pathways are used throughout the neuroendocrine system to regulate secretion and gene expression. It has recently been shown that cAMP and Ca2+ can converge to regulate the activity of a single transcription factor, CREB. This convergence likely permits CREB to integrate multiple extracellular signals in the regulation of transcription. Presumably the effects of Ca2+ on CREB activation are mediated by Ca2+/calmodulin-dependent protein kinases (CaM Kinases). Recent studies from this laboratory have demonstrated that specific CaM Kinases can have very different effects on the activation of CREB. In particular, we found that CaM Kinase II can inhibit activation of CREB while CaM Kinase IV can activate CREB. The proposed studies seek to further characterize the ability of specific CaM Kinases to regulate activation of the CREB/ATF transcription factor family. The specific aims include: i) Test the hypothesis that phosphorylation of Ser142 of CREB is physiologically regulated. This will involve preparation of an antibody which detects phosphorylation of Ser142. 2) Determine if the inhibitory effects of CaMKII are also observed for ATF1, a related leucine-zipper transcription factor, which has also been shown to respond to changes in cAMP and Ca2+. The region surrounding the negative regulatory site of CREB at Ser142 is conserved, but not identical in ATF1. 3) Examine the ability of CaMKI to activate CREB. It is possible that CaM Kinases other than CaMKIV can activate CREB. We will use transfection assays to determine if CaMKI can activate CREB. 4) Test the hypothesis that the inhibitory effects of CaMKII on activation of CREB are due to the effects of phosphorylation at Ser142 to block interaction between CREB and the co- activator protein, CREB binding protein (CBP). 5) Map the portions of CREB and Cl3P which are required for high affinity, PKA-dependent and CaM Kinase-induced interactions. 6) Develop an episomal, cAMP-responsive reporter gene to examine effects of cAMP and Ca2+ on the interaction of CREB with chromatin structure. These studies should provide new insight into the mechanisms which permit cAMP and Ca2+ to regulate members of the CREB/ATF family.