Regulation of Cav1.2 Trafficking by GJA1-20k and cBIN1

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In normal hearts, the transverse tubules (t-tubules) concentrate L-type calcium channels (LTCCs) to initiate the beat-to-beat intracellular calcium transients which contribute to the regulation of cardiac contraction and relaxation. An altered calcium transient is a key pathophysiological sign of failing heart. However, mechanisms of maintenance of LTCC expression at t-tubule microdomains for optimal calcium-induced-calcium-release (CICR) remain poorly understood. Understanding the dynamic regulation of calcium handling microdomains at t-tubules, and in particular LTCC regulation at the microdomains, is needed to understand the pathophysiology of failing hearts and to identify new pathways and molecules that can be targeted for heart failure therapy development. The overall objective of this MPI application is to further our mechanistic understanding of how LTCC expression at t-tubule cBIN1-microdomains is maintained and why LTCC localization is altered in heart failure. We will explore the central hypothesis that homeostatic LTCC expression at t-tubules is highly dynamic and maintained by both internal delivery and external delivery. We expect that the internal delivery is determined by intracellular actin reservoirs of LTCCs organized at Z-disks by a newly identified actin nucleator GJA1-20k (Aim 1, RMS). We also expect that there is a local extracellular reservoir of cBIN1 vesicles (Aim 2, TTH), which feed t-tubule cBIN1-microdomains from other regions of the heart to modulate LTCC expression at t-tubule surfaces. In heart failure, both sources are depleted yet could be exogenously restored. The resources of a membrane biology lab (TTH) and a trafficking lab focused on heart failure progression (RMS) are combined in this application for the proposed research plan. The rationale that underlies the proposed research is that LTCCs at t-tubule microdomains are dynamically regulated from both inside and out, are reduced in heart failure, and can be restored presenting a novel therapy for heart failure. The proposed research is innovative because it will identify new regulatory mechanisms of remodeled myocardium during heart failure progression, and lead to the establishment of molecules and pathways that can be targeted for therapy development. The proposal has a strong premise because it is based on extensive published and preliminary data from successful in vitro cell-free studies, intact cell line and primary cardiomyocyte studies, and successful genetic and in vivo animal heart failure models. The significance is high because it will lead to development of new therapeutic strategies that will allow preservation and restoration of efficient excitation-contraction coupling in diseased hearts.