Loss of function of hNa_v1.5 by a ZASP1 mutation associated with intraventricular conduction disturbances in left ventricular noncompaction

Background - Defects of cytoarchitectural proteins can cause left ventricular noncompaction, which is often associated with conduction system diseases. We have previously identified a p.D117N mutation in the LIM domain-binding protein 3-encoding Z-band alternatively spliced PDZ motif gene (ZASP) in a patient with left ventricular noncompaction and conduction disturbances. We sought to investigate the role of p.D117N mutation in the LBD3 NM-001080114.1 isoform (ZASP1-D117N) for the regulation of cardiac sodium channel (Na_v1.5) that plays an important role in the cardiac conduction system. Methods and Results - Effects of ZASP1-wild-type and ZASP1-D117N on Nav1.5 were studied in human embryonic kidney-293 cells and neonatal rat cardiomyocytes. Patch-clamp study demonstrated that ZASP1-D117N significantly attenuated I_Na by 27% in human embryonic kidney-293 cells and by 32% in neonatal rat cardiomyocytes. In addition, ZASP1-D117N rightward shifted the voltage-dependent activation and inactivation in both systems. In silico simulation using Luo-Rudy phase 1 model demonstrated that altered Na_v1.5 function can reduce cardiac conduction velocity by 28% compared with control. Pull-down assays showed that both wild-type and ZASP1-D117N can complex with Na_v1.5 and telethonin/T-Cap, which required intact PDZ domains. Immunohistochemical staining in neonatal rat cardiomyocytes demonstrates that ZASP1-D117N did not significantly disturb the Z-line structure. Disruption of cytoskeletal networks with 5-iodonaphthalene-1- sulfonyl homopiperazine and cytochalasin D abolished the effects of ZASP1-D117N on Na_v1.5. Conclusions - ZASP1 can form protein complex with telethonin/T-Cap and Na_v1.5. The left ventricular noncompaction-specific ZASP1 mutation can cause loss of function of Na _v1.5, without significant alteration of the cytoskeletal protein complex. Our study suggests that electric remodeling can occur in left ventricular noncompaction subject because of a direct effect of mutant ZASP on Na_v1.5.