Characterization Of A Novel TRPC6 Mutant Identified In FSGS Patients

Abstract

TRPC6 is a Ca2+-permeable non-selective cation channel. Gain of function mutations of TRPC6 have been shown to cause focal segmental glomerulosclerosis (FSGS). Among six mutants of TRPC6 indentified in FSGS, three of them cause increase in Ca2+ influx. It appears that the enhanced Ca2+ influx underlies TRPC6 mutation associated FSGS. However, it is unclear how different mutations lead to gain of channel function and increase in Ca2+ influx. Here we report a novel TRPC6 mutant, M132T, which causes early-onset FSGS. Whole cell patch clamp experiments showed that current amplitude of M132T was 3- to 5-fold larger (476.9 ± 55.9 pA/pF) than that of wild-type (wt) TRPC6. Interestingly, while the wt TRPC6 exhibited apparent time-dependent inactivation, M132T did not show inactivation or only minor time-dependent decline of inward current. Inward Ca2+ current of M132T measured in 10 mM Ca2+ external solution was 10-fold larger than that of wt TRPC6. Moreover, Ca2+ influx of M132T was also significantly bigger than wt TRPC6. To understanding the mechanism of slow inactivation kinetics of M132T, we applied various intracellular Ca2+ concentrations and compared inactivation processes of M132T and wt TRPC6. We found that higher Ca2+ concentration was required to induce M132T inactivation in comparison with wt TRPC6, suggesting that M132T is less sensitive to intracellular Ca2+ induced inactivation. Taken together, our results indicate that the lack of inactivation may confer the enhanced Ca2+ influx in M132T. Further investigation is required to understand the mechanism of enhanced channel functions of TRPC6 mutants in FSGS.