Regulation of Cardiac Calcium Channels by an Autoinhibitory Signaling Complex

描述（由申请人提供）：由Cav1.2通道传导的l型钙电流负责钙进入，引发心肌收缩，因此这些通道是许多不同效应器（如神经递质、激素和药物及其受体）调节钙信号和收缩力的最终共同途径。l型钙电流的改变在心血管疾病和治疗中至关重要。l型钙电流的失调会导致高血压，钙通道拮抗剂药物是一种重要的治疗模式。缺血性心脏病常伴有心绞痛，也可用钙拮抗剂治疗。心律失常可由l型Ca电流的调节改变和时间不适当的Ca瞬态产生早期和延迟的去极化后产生，Ca拮抗剂药物在治疗心房心律失常中很重要，l型Ca电流的肾上腺素能调节在心力衰竭中发生改变。令人惊讶的是，尽管Cav1.2通道在心血管生理学和病理生理学中很重要，但心肌细胞中Cav1.2通道的调控尚不清楚。由于它们在收缩调节中的关键作用，许多细胞内调节剂和第二信使聚集在这些Ca通道上并调节其功能，包括Mg、cAMP、Ca和钙调蛋白。我们的工作表明，这些第二信使的作用位点在细胞内的大c端结构域，约占al亚基质量的30%。此外，c端结构域受到蛋白水解的影响，从而调节了其功能。因此，c端结构域整合了多种细胞调控信号，共同形成了一个控制Ca通道活性的完整胞内信号网络。本项目拟确定Cav1.2通道自抑制c端结构域的分子机制和生理意义，明确Cav1.2通道c端蛋白水解加工的机制和生理意义，确定(-肾上腺素能受体途径通过PKA与通道远端c端结构域结合的AKAP15，调控Cav1.2通道的分子机制。我们的实验结果对于理解心肌细胞中Ca和cAMP信号的调节及其在心血管疾病中的功能障碍至关重要。这些信息将为旨在预防和治疗心血管疾病的转化研究提供必要的基础科学背景。

DESCRIPTION (provided by applicant): L-type Ca currents conducted by Cav1.2 channels are responsible for Ca entry that initiates contraction in cardiac muscle, and these channels are therefore the final common pathway for regulation of Ca signaling and contractile force by many different effectors such as neurotransmitters, hormones and drugs, and their receptors. Alterations in L-type Ca currents are crucially involved in cardiovascular disease and therapy. Misregulation of L-type Ca currents contributes to hypertension, and Ca channel antagonist drugs are an important mode of therapy. Ischemic heart disease is often accompanied by angina pectoris, which is also treated with Ca antagonist drugs. Arrhythmias can be generated by altered regulation of L-type Ca currents and by inappropriately timed Ca transients generating early and delayed afterdepolarizations, and Ca antagonist drugs are important in treatment of atrial arrhythmias, (-adrenergic regulation of L-type Ca currents is altered in heart failure. Surprisingly, despite their importance in cardiovascular physiology and pathophysiology, regulation of Cav1.2 channels in cardiac myocytes is not well understood. Because of their key role in regulation of contraction, many intracellular regulators and second messengers converge on these Ca channels and regulate their function, including Mg, cAMP, Ca, and calmodulin. Our work has shown that the sites of action of these second messengers are in the large intracellular C-terminal domain, which represents approximately 30% of the mass of the al subunit. In addition, the C-terminal domain is subject to proteolytic processing, which modulates its function. Thus, the C-terminal domain integrates many kinds of cellular regulatory signals, which together form an integrated intracellular signaling network controlling Ca channel activity. In this project we propose to determine the molecular mechanism and physiological significance of Cav1.2 channel autoinhibition the C-terminal domain, define the mechanism and physiological significance of proteolytic processing of the C-terminal of Cav1.2 channels, and determine the molecular mechanism of regulation of the Cav1.2 channel by the (-adrenergic receptor pathway acting through PKA bound to the channel's distal C-terminal domain by AKAP15. The results of our experiments will be crucial for understanding regulation of Ca and cAMP signaling in the cardiac myocyte and its dysfunction in cardiovascular disease. This information will provide the essential basic science background for translational research aimed at preventing and treating cardiovascular disease.