Ca Regulation of Ca Channels

Ca 通道的 Ca 调节

基本信息

批准号：
7265541
负责人：
DAVID T YUE
金额：
$ 34.85万
依托单位：
JOHNS HOPKINS UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2007
资助国家：
美国
起止时间：
2007-07-01 至 2012-06-30
项目状态：
已结题

项目摘要

DESCRIPTION (provided by applicant): Over the past grant cycle, general rules for calmodulin (CaM) regulation of the family of Ca channels were discerned. CaM has two lobes, each with two Ca2+ binding sites. A first general aspect was that each lobe can autonomously trigger a form of channel regulation. Secondly, whenever the C-terminal lobe of CaM (C-lobe) triggers channel regulation, there is preferential responsiveness to local Ca2+ signals. Conversely, wherever the N-terminal lobe (N-lobe) initiates regulation, there is selectivity for global Ca2+ signals. Thirdly, there is reason to expect that this rule of operation generalizes beyond Ca channels, to many complexes in which CaM is preassociated with target molecules. Because CaM regulation of Ca channels (and other signaling molecules) is crucial for normal neuroprocessing, and likely important for therapeutics relating to pain, psychosis, and cardiac arrhythmogenesis, answering how these general rules occur is the overarching thrust for the next cycle of research. Three aims will address this overall theme. 1. To develop and perform elementary tests of a kinetic Ca2+ decoding mechanism for the CaM/Ca channel complex. This aim formulates a 'kinetic Ca2+ decoding' theory of how the CaM decoding occurs, and devises novel Voltage-block' experiments to enable elementary tests of this theory. 2. To engineer the local/global Ca2+ preference of CaM/Ca channel regulation, as a higher-order test of the kinetic Ca2+ decoding theory, and as means to glean design principles for developing novel channel modulators. A principal prediction of the kinetic Ca2+ decoding theory is that the local/global Ca2+ preference of channel regulation reflects competition between channel affinities for the Ca2+-bound and Ca2+-free forms of a lobe of CaM. Aim 2 will alter these affinities and check for the predicted changes in Ca2+ preference. Aim 2 will also explore whether these modifications can inform the design of drug compounds that modulate channel regulation in new ways. 3. To experimentally determine Ca2+ concentrations and diffusion within the nanometers of the Ca channel. Crucial to the next phase of progress is the direct measurements of local and global Ca2+ concentration signals, and Ca2+ diffusion, in the actual channel 'nanodomain' environment. Fusions of a genetically-encoded Ca2+ sensor (TNL-15) to channels, combined with TIRF microscopy promise to reveal these long sought-after entities. These aims promise bold progress, with basic and applied ramifications.

描述（由申请人提供）：在过去的资助周期中，钙调蛋白 (CaM) 对 Ca 通道家族的调节的一般规则已被识别。 CaM 有两个叶，每个叶有两个 Ca2+ 结合位点。第一个一般方面是每个波瓣可以自主触发某种形式的通道调节。其次，每当 CaM 的 C 端叶 (C-lobe) 触发通道调节时，就会优先响应局部 Ca2+ 信号。相反，只要 N 端叶 (N-lobe) 启动调节，全局 Ca2+ 信号就会具有选择性。第三，有理由预期这一操作规则会推广到 Ca 通道之外，推广到 CaM 与目标分子预关联的许多复合物。由于 CaM 对 Ca 通道（和其他信号分子）的调节对于正常的神经处理至关重要，并且可能对于与疼痛、精神病和心律失常发生相关的治疗也很重要，因此回答这些一般规则如何发生是下一个研究周期的首要任务。三个目标将解决这一总体主题。 1. 开发和执行 CaM/Ca 通道复合物的动力学 Ca2+ 解码机制的基本测试。该目标制定了 CaM 解码如何发生的“动态 Ca2+ 解码”理论，并设计了新颖的电压块”实验来对该理论进行基本测试。 2. 设计 CaM/Ca 通道调节的局部/全局 Ca2+ 偏好，作为动力学 Ca2+ 解码理论的高阶测试，并作为收集开发新型通道调制器的设计原理的手段。动力学 Ca2+ 解码理论的主要预测是，通道调节的局部/全局 Ca2+ 偏好反映了 CaM 叶的 Ca2+ 结合和 Ca2+ 游离形式的通道亲和力之间的竞争。目标 2 将改变这些亲和力并检查 Ca2+ 偏好的预测变化。目标 2 还将探索这些修饰是否可以为以新方式调节通道调节的药物化合物的设计提供信息。 3. 通过实验确定 Ca2+ 浓度和 Ca 通道纳米范围内的扩散。下一阶段进展的关键是在实际通道“纳米域”环境中直接测量局部和全局 Ca2+ 浓度信号以及 Ca2+ 扩散。基因编码 Ca2+ 传感器 (TNL-15) 与通道的融合，与 TIRF 显微镜相结合，有望揭示这些长期追捧的实体。这些目标有望取得大胆进展，并产生基本和应用影响。