Interactome of splice variants of the ß2-subunit of L-type voltage-gated calcium channels in cardiac myocytes.

心肌细胞通过L型电压门控钙通道(LTCC)的钙内流的区划是正确调控心脏活动的必要条件。LTCC位于肌膜内不同的亚细胞室。Cav？是LTCCs最重要的调节亚基。在四种Cav？亚型中，Cav？2主要在心肌细胞中表达。我们最近的数据和以前的报告已经检测到在成年鼠和人的心肌细胞中共存的不同的Cav？2剪接变体。此外，除了LTCC外，该亚基还与不同的蛋白质相关联。我们的初步结果表明，Cav？2的不同亚群位于小鼠心肌细胞肌膜的不同亚细胞微区。这些不同的亚群可能对应于特定的剪接变异体，参与到LTCC附近的不同信号通路成员的招募。因此，这个项目有两个主要目标。首先，我们将专注于鉴定与小鼠心肌细胞表达的每个Cav？2剪接变异体相互作用的蛋白质。其次，我们将研究在生理性和心肌肥大条件下，每种Cav？2剪接变异体及其相关蛋白的细胞定位和蛋白表达。为了达到这些目标，我们将结合串联亲和纯化、质谱学、系统生物学分析和超分辨显微镜。

The compartmentalization of calcium influxes through L-type voltage-gated calcium channels (LTCCs) in cardiomyocytes is necessary for the correct regulation of cardiac processes. LTCCs are located in different subcellular compartments in the sarcolemma. Cavß is the most important LTCCs regulatory subunit. Of the four Cavß isoforms, Cavß2 is predominantly expressed in cardiomyocytes. Our recent data and previous reports have detected different Cavß2 splice variants coexisting in adult mouse and human cardiomyocytes. In addition, the association of this subunit to diverse proteins besides the LTCCs has also been reported. Our preliminary results show that diverse subpopulations of Cavß2 are located in different subcellular microdomains of the sarcolemma in mouse cardiomyocytes. These different subpopulations could correspond to specific splice variants being involved in the recruitment of members of distinctive signaling pathways to LTCCs vicinity. Therefore, this project has two main goals. First, we will focus on the identification of proteins interacting with each Cavß2 splice variant expressed in mouse cardiomyocytes. Secondly, we will study the cellular localization and protein expression of each Cavß2 splice variant and their associated proteins under physiological and cardiac hypertrophy conditions. To reach these objectives we will combine tandem affinity purifications, mass spectrometry, systems biology analyses and super-resolution microscopy.