Regulation of CBP by Synaptic Activity

描述（由申请人提供）： 活性调节的基因表达在两个过程中都起着重要作用。 突触回路的重塑和神经元存活。活性依赖 细胞内Ca 2+的增加触发了数百种或可能是 成千上万的基因。cAMP反应元件（CRE）被确定为主要的 通过立即早期基因（IEG）启动子分析的Ca 2+响应元件 许多（如果不是大多数的话）Ca 2+应答启动子含有克雷斯。碱性亮氨酸 拉链转录因子CREB作为二聚体与CRE结合， 通过Ca 2+、cAMP/PKA和生长因子信号通路激活。的 转录因子CREB是活性诱导的Ca 2+内流的主要靶点， 是神经元存活和适应性突触可塑性的关键调节因子。 然而，Ca 2+激活CREB的机制尚不清楚， 阐明。已经确定PKA磷酸化CREB的Ser 133， 产生磷酸-丝氨酸对接基序，其募集其共活化剂CBP（或 同源物，p300）。CBP的募集被认为可以增强转录水平， 激活或者通过其与一般转录因子的结合， 机械或通过其内在的组蛋白乙酰转移酶活性。虽然 活动诱导的Ca 2+内流被广泛认为调节CREB功能， 类似的方式，促进CBP招募的信号级联， 转录激活尚未明确定义。 该提案的一个主要主题是确定PKA-CREB-CBP是否 范例还涉及Ca 2+激活的转录。因此，最初的目标 这项研究的目的是确定神经元活动是否会诱导 招聘CBP到CREB，以及是否需要招聘CBP 转录激活提出了四个具体目标。(1)确定 突触活动是否诱导神经元中CBP向CREB的募集， CBP的募集是否足以完全激活转录。 (2)破译活动调节信号级联，促进 CREB和CBP的磷酸化和活化。(3)确定突触 活性和Ca 2+内流调节CBP功能。(4)分析了 CBP和CREB在CaM激酶IV缺陷小鼠中的功能。本研究旨在 剖析导致突触活动介导的生物化学事件 CREB的磷酸化、CBP的募集和CBP依赖性的 转录。深入了解活动介导的 转录与神经变性病理的治疗相关 这可能是由兴奋性毒性细胞死亡或凋亡引起的。通过一个指导 职业发展计划，首席研究员将获得更高的 在分子神经生物学领域具有生产力， 关于基因调控。这将通过进行上述 研究，出席和介绍地方和国家研讨会和会议， 并参与分子生物学和神经生理学的教学课程 技术.

DESCRIPTION (provided by applicant): Activity-regulated gene expression plays a fundamental role in both the remodeling of synaptic circuitry and in neuronal survival. Activity-dependent increases in intracellular Ca2+ trigger the expression of hundreds or perhaps thousands of genes. The cAMP response element (CRE) was identified as a major Ca2+ responsive element via analysis of immediate early gene (IEGs) promoters and many if not most Ca2+ responsive promoters contain CREs. The basic leucine zipper transcription factor, CREB, binds to the CRE as a dimer and can be activated by Ca2+, cAMP/PKA, and growth factor signaling pathways. The transcription factor CREB is a major target of activity-induced Ca2+ influx and is a key regulator of both neuronal survival and adaptive synaptic plasticity. Nevertheless, the mechanisms by which Ca2+ activates CREB have not been clearly elucidated. It is well established that PKA phosphorylates Ser 133 of CREB and creates a phospho-serine docking motif that recruits its coactivator CBP (or the homologue, p300). Recruitment of CBP is thought to enhance transcriptional activation either via its association with the general transcriptional machinery or via its intrinsic histone acetyltransferase activity. Although activity-induced Ca2+ influx is widely believed to regulate CREB function in an analogous manner, the signaling cascades that promote CBP recruitment and transcriptional activation have not been clearly defined. A major theme of this proposal is to determine whether the PKA-CREB-CBP paradigm also pertains to Ca2+-activated transcription. Thus, an initial goal of this study will be to determine whether neuronal activity induces the recruitment of CBP to CREB and whether CBP recruitment is required for transcriptional activation. Four specific aims are proposed. (1) Determine whether synaptic activity induces the recruitment of CBP to CREB in neurons and whether recruitment of CBP is sufficient for full transcriptional activation. (2) Decipher the activity-regulated signaling cascades that promote the phosphorylation and activation of CREB and CBP. (3) Determine how synaptic activity and Ca2+ influx regulate CBP function. (4) Analyze the regulation of CBP and CREB function in mice deficient for CaM kinase IV. This study seeks to dissect the biochemical events result in synaptic activity-mediated phosphorylation of CREB, recruitment of CBP, and "activation" of CBP-dependent transcription. Gaining insight into the mechanisms of activity-mediated transcription has relevance in the treatment of neurodegenerative pathologies that may result from excitotoxic cell death or apoptosis. Through a mentored career development plan, the Principal Investigator will gain an enhanced capacity to be productive in the field of molecular neurobiology with a focus on gene regulation. This will be accomplished by conducting the above described research, attending and presenting at local and national seminars and meetings, and participating in courses teaching molecular biology and neurophysiology techniques.