Xin proteins and cardiac rhythms

描述（由申请人提供）：心力衰竭是唯一一种全球发病率不断上升的心血管疾病1。在美国，大约有500万人患有心力衰竭，每年有超过550，000人首次被诊断出。在心脏病中，心律失常导致的衰竭在2009年造成约37，000人死亡，2006年心律失常的总估计医疗费用总计31亿2。心律失常的分子基础尚不清楚，但Xin已被确定为与心脏病相关3 - 12。人类心肌病相关基因（分别为CMYA 1和CMYA 3）9的小鼠直系同源物（mXina和mXinb）编码定位于闰盘（ICD）的蛋白质。缺乏mXina的小鼠心脏导致成人迟发性心肌病，伴有传导缺陷并上调mXinb，尽管年轻时ICD外观正常7。另一方面，mXinb的完全丧失导致ICD形成失败、舒张功能障碍和出生后早期致死4。mXinb无效心脏表现出mXina的错误定位，因此可能导致传导缺陷和/或心律失常。我们的长期目标是确定mXin蛋白影响心律过程的分子机制。我们发现，mXina-空心肌细胞具有降低的瞬时外向钾（Ito）电流密度。类似于Kv4.2（Ito的通道形成亚基），mXina也与Kv通道相互作用蛋白2（KChIP 2，Ito的辅助亚基）和细丝蛋白（肌动蛋白交联蛋白）相互作用。通过这些相互作用，mXina可以促进Ito通道的表面表达。我们的工作假设是，在一个功能层次中，mXinb在控制mXina的定位中起着至关重要的作用，而mXina又通过与KChIP 2和细丝蛋白的相互作用调节Ito通道的表面表达。在目的1中，我们将建立mXina调节Ito通道的表面表达和功能的作用和机制。我们将定义KChIP 2和细丝蛋白结合域的mXina，产生非偶联突变体，并测试其对通道表面表达的影响。目的2是确定mXinb在出生后早期和成人心脏中mXina和其他ICD组分的ICD定位中的作用。我们预计mXinb，mXina和ICD组件之间的功能层次结构，在他们的ICD本地化的行动。我们将使用可诱导的心脏特异性mXinb缺失小鼠来测试mXinb是否是维持成人心脏中ICD组装所必需的。在目标3中，我们将确定mXinb调节表面通道表达、动作电位时程和心律的作用和机制。mXina和mXinb都代表了相对未开发的领域，尽管它们在ICD形成和离子通道表面表达中起着重要作用。通过这些研究，我们将进一步了解心律失常和充血性心力衰竭的疾病过程中的机制，从而希望找到新的，有效的治疗靶点。

DESCRIPTION (provided by applicant): Heart failure is the only cardiovascular disease with an increasing worldwide incidence1. Approximately 5 million people in the United States have heart failure, with over 550,000 diagnosed for the first time each year. Among heart diseases, failure due to arrhythmias is causing about 37,000 deaths in 2009 and the total estimated health care costs for arrhythmias in 2006 totaled 3.1 billion2. The molecular basis of arrhythmias is unclear, but Xin has been identified as being related to heart diseases3- 12. The mouse orthologs (mXina and mXinb) of human cardiomyopathy-associated genes (CMYA1 and CMYA3, respectively)9 encode proteins localized to the intercalated discs (ICDs). Mouse hearts deficient in mXina lead to adult late-onset cardiomyopathy with conduction defects and up-regulate mXinb, despite a normal appearance of ICD at young ages7. On the other hand, complete loss of mXinb results in failure of forming ICD, diastolic dysfunction, and early postnatal lethality4. The mXinb-null hearts exhibit mis-localization of mXina, and thus could lead to conduction defects and/or arrhythmias. Our long-term goal is to determine the molecular mechanisms by which mXin proteins influence the process of cardiac rhythms. We found that the mXina-null cardiomyocytes had reduced transient outward potassium (Ito) current density. Similar to Kv4.2 (a channel-forming subunit of Ito), mXina also interacted with Kv channel interacting protein 2 (KChIP2, an auxiliary subunit of Ito) and filamin (an actin crosslinking protein). Through these interactions, mXina may promote the surface expression of the Ito channel. Our working hypothesis is that in a functional hierarchy, mXinb plays essential role in controlling the localization of mXina, which, in turn, regulates the surface expression of Ito channel via its interactions with KChIP2 and filamin. In the Aim 1, we will establish the roles and the mechanisms by which mXina regulates the surface expression and functioning of Ito channels. We will define KChIP2 and filamin binding domains on mXina, generate uncoupled mutants, and test their effects on channel surface expression. The Aim 2 is to establish the roles of mXinb in the ICD localizations of mXina and other ICD components in early postnatal and adult heart. We anticipate a functional hierarchy among mXinb, mXina, and ICD components, in their ICD localized actions. We will use inducible, cardiac-specific mXinb-null mice to test if mXinb is required for maintaining ICD assembly in adult heart. In the Aim 3, we will determine the role and the mechanisms by which mXinb regulates surface channel expression, action potential duration and then cardiac rhythm. Both mXina and mXinb represent relatively unexplored territories, despite their essential roles in the ICD formation and ion channel surface expression. Through these studies, we will advance our understanding of the mechanisms in the disease processes of arrhythmias and congestive heart failure and thus hope to identify novel, effective therapeutic targets.