Human Bin1 Isoforms Maintain, Regenerate And Elicit Functional Ec-Coupling And Couplons In Adult Rat And Human Induced Pluripotent Stem Cell-Derived Cardiomyocytes

In ventricular myocytes, membrane invaginations (T-tubules) are instrumental for excitation-contraction (EC) coupling and an aberrant T-tubular system is a hallmark of many cardiac diseases. The Bridging integrator 1 (BIN1) protein is a key contributor to the biogenesis of membrane invaginations. We set out to study the expression pattern of BIN1 isoforms in human heart tissue and study their efficiency in generating and maintaining T-tubules and functional EC-coupling sites, so called couplons. For this, we investigated the expression pattern of BIN1 isoforms in human heart samples and identified 5 BIN1 splice variants of which the isoform 8 was formerly exclusively attributed to skeletal muscle. We characterized these 5 BIN1 isoforms in adult rat ventricular myocytes and human induced pluripotent stem cell-derived cardiac myocytes (hiPSC-CMs) by using viral gene transfer, realtime confocal microscopy and functional couplon analysis using our CaCLEAN approach published recently (Tian et al., 2017). We found that all five BIN1 splice variants induced de novo generation of T-tubules in both cell types. Isoforms with the phosphoinositide binding motif (PI) were most potent in maintenance and regeneration of T-tubules and functional EC-coupling in adult myocytes. In hiPSC-CMs, BIN1 induced de novo generation of T-tubules, formation of EC-coupling sites and enhanced calcium handling. Functional couplon sites were investigated by the CaCLEAN approach and by a genetically encoded calcium sensor directed to couplon sites (junctin-GCaMP6). From these data we conclude that BIN1 plays key roles in the maintenance, regeneration, and de novo generation of functional T-tubules, especially the isoforms with the PI motif. These newly generated T-tubules evoke the formation of functional EC couplons contributing to enhanced calcium handling.The work was funded in parts by the DFB/SFB894 and DFB/SFB TR 152.