Mechanisms of Long-term Cardiac Ion Channel Regulation

描述(由申请人提供)：心肌细胞中的钙调节失调导致心脏发育缺陷和老化心脏的疾病。该建议的长期目标是提供一种解释心脏L型钙通道(LTCC)如何感知和转导稳态调节心肌细胞的信号的分子机制。心肌细胞在钙信号转导到细胞核方面是一个难题。细胞内钙的幅度在每个心动周期中变化10倍，但钙的变化以某种方式被差异解码为长期转录信号。在这个资助时期，我们将测试钙通道的活动和心脏L型钙通道本身是否编码钙信号，以便进行长期调节。在过去的资助期间，我们发现RGK慢性抑制ICa，L(LTCC电流)，这种RGK抑制ICa，L导致CaV1.2mRNA的代偿性上调。这提示，ICA、L阻断可能是核内转录事件的信号传导途径。在新的研究中，我们证实并扩展了这一概念，表明LTCC-药理-阻断，而不是一般的内部钙负责扰乱心脏发育。同样，在成熟心脏中，长期阻断LTCC也会导致LTCC和ICa，L的代偿性上调。我们的驱动假设是，信号不是简单地由钙决定的，而是由活跃的钙通道决定的。LTCC的可移动片段定位于细胞核或T管，再加上最近报道该肽是一种转录因子，这一发现推动了令人兴奋的新假说，即LTCC的这一片段调节LTCC的表达。我们将从三个方面研究这方面的长期通道调节：1.我们将评估LTCC一个结构域的核转位，并确定LTCC活性与亚细胞定位之间的相互作用；2.我们将确定LTCC转录自我调节的能力；以及3.我们将确定LTCC阻断后SL Ca调节蛋白的代偿性变化。这项工作可能在LTCC功能和下游信号事件之间提供了缺失的分子链接。公共卫生相关性：这些研究表明，广泛使用的用于阻断LTCC的临床相关药物可能会通过矛盾地增加LTCC功能而无意中加剧心功能障碍。这一提议将有助于理解一种新的机制，即控制心脏电活动的离子通道也可能控制对维持心脏结构和功能至关重要的长期信号通路。

DESCRIPTION (provided by applicant): Ca dysregulation in cardiac myocytes contributes to heart development defects and diseases of the aging heart. The long-term objective of this proposal is to provide a molecular mechanism that explains how cardiac L-type Ca channels (LTCC) sense and transduce signals that homeostatically regulate cardiac myocytes. Cardiac myocytes present a conundrum with respect to Ca signaling to the nucleus. Cytosolic Ca amplitude varies >10-fold during each cardiac cycle, yet alterations of Ca somehow are differentially decoded for longer-term transcriptional signaling. In this funding period we will test whether Ca channel activity and the cardiac L-type Ca channel itself encodes Ca signaling for long-term regulation. In the past funding period we discovered that RGK chronically inhibited ICa,L (LTCC current), and this RGK inhibition of ICa,L resulted in a compensatory up-regulation of CaV1.2 mRNA. This suggests that ICa,L block may signal transcriptional events in the nucleus. In new studies we confirmed and extended this notion by showing that LTCC-pharmacological- block, but not internal Ca in general is responsible for perturbing heart development. Along the same lines, in mature heart, long-term blockade of LTCC also causes a compensatory up-regulation of LTCC and ICa,L. Our driving hypothesis is that signaling is not simply determined by Ca, but by active Ca channels. The discovery that mobile segment of LTCC is localized to the nucleus or t-tubules coupled with the recent report that this peptide is a transcription factor drives the exciting new hypothesis that this segment of the LTCC, regulates LTCC expression. We will study this aspect of long-term channel regulation in three aims: 1. We will assess nuclear translocation of a domain of the LTCC, and determine the interaction between LTCC activity and sub- cellular localization; 2. We will determine the ability of LTCC to auto-regulate itself transcriptionally; and 3. We will determine the compensatory changes of SL Ca handling proteins in response to LTCC blockade. This work may provide a missing molecular link between LTCC function and downstream signaling events. PUBLIC HEALTH RELEVANCE: These studies show that widely used clinically-relevant drugs that are used to block LTCC may inadvertently exacerbate heart dysfunction by paradoxically increasing LTCC function. This proposal will lead to understanding of a new mechanism whereby ion channels that control cardiac electrical activity also may control long-term signaling pathways that are critical for maintenance of cardiac structure and function.