Physical Interaction of RyR2 and VDAC2 as a Molecular Architecture for the Coupling Between Sarcoplasmic Reticulum and Mitochondria in Cardiomyocytes

Inter-organellar physical coupling between mitochondria and sarcoplasmic reticulum (SR) and the presence of high Ca2+ microdomain for Ca2+ transfer into mitochondria have been reported in cardiomyocytes (Garcia-perez et al., J. Biol. Chem., 2008). However, the molecular components involved in the physical coupling remain to be elucidated. In the present study, we identified VDAC2 as a putative binding partner for RyR2 in SR by the bacterial two-hybrid screening. The interaction of VDAC2 with RyR2 was further confirmed by GST-pull down and co-immunoprecipitation assays. As tethering structures between SR and mitochondria was detected in cardiomyocytes (Boncompagni et al., Mol. Biol. Cell., 2009), we identified a close apposition between mitochondria and SR in HL-1 cells by electron microscopic study. With the immuno-gold labeling strategy, furthermore, the co-localization of VDAC2 with RyR2 at the junction between SR and mitochondria was revealed at sub-micrometer resolution. In adult ventricular cardiomyocytes, the association of VDAC2 and RyR2 was detected at the sub-sarcolemmal region according to immunocytochemical study and differential centrifugations. To investigate the roles of VDAC2 in cardiomyocytes, we attempted to use adenovirus-based shRNA system to knock down VDAC2 gene expression. Under electrical field stimulation (1Hz), the diastolic [Ca2+] increased significantly in VDAC2 knockdown cells without alteration of Ca2+ transient amplitude and Ca2+ uptake rate. Taken together, the present study suggests that VDAC2 and RyR2 are the molecular components for the linkage between mitochondria and SR, and regulate Ca2+ signaling in cardiomyocytes. (Supported by Korea MEST NRF grant (20110002144), the 2011 GIST Systems Biology Infrastructure Establishment Grant and KISTI-KREONET)