Regulation of podocyte Slo1 BK channels by synaptopodin

Abstract

Large‐conductance Ca2+‐activated K+ channels encoded by the Slo1 gene are expressed in podocytes, where they can interact with TRPC6 channels at slit diaphragms in close conjunction with cytoskeletal regulatory proteins such as synaptopodin. We observed that synaptopodin and Slo1 proteins can be reciprocally co‐immunoprecipitated when they are heterologously expressed in HEK293T cells. This also occurs with endogenously expressed proteins in an immortalized mouse podocyte cell line, where these proteins are closely co‐localized. GST pull‐down assays show that the both C‐ and N‐terminal domains of synaptopodin can interact with distal portions of cytoplasmic C‐termini of Slo1. Co‐expression of synaptopodin with the VEDEC isoform of Slo1 proteins, which is endogenously expressed in podocytes, increases the expression of Slo1 on the surface of HEK293T cells. Moreover, there is a reduction of Slo1 on the surface of mouse podocytes with stable knockdown of synaptopodin compared to a control cell line. Stable synaptopodin knockdown does not affect total expression of Slo1 proteins but does result in a reduction in macroscopic currents carried by these channels. These data indicate that synaptopodin regulates trafficking of functional BKCa channels in podocytes and thereby modulate responses to mechanical and chemical stimuli. Supported by R01 DK082529 (SED) and RO1 DK057683 (PM)