Regulation of podocyte BKCa channels by synaptopodin, Rho, and actin microfilaments

Kim EY, Suh JM, Chiu YH, Dryer SE. Regulation of podocyte BKCa channels by synaptopodin, Rho, and actin microfilaments. Am J Physiol Renal Physiol 299: F594-F604, 2010. First published July 14, 2010; doi:10.1152/ajprenal.00206.2010.-Mechanosensitive large-conductance Ca2+-activated K+ channels encoded by the Slo1 gene (BKCa channels) are expressed in podocytes. Here we show that BKCa channels reciprocally coimmunoprecipitate with synaptopodin (Synpo) in mouse glomeruli, in mouse podocytes, and in a heterologous expression system (HEK293T cells) in which these proteins are transiently expressed. Synpo and Slo1 colocalize along the surface of the glomerular basement membrane in mouse glomeruli. Synpo interacts with BKCa channels at COOH-terminal domains that overlap with an actin-binding domain on the channel molecule that is necessary for trafficking of BKCa channels to the cell surface. Moreover, addition of exogenous beta-actin to mouse podocyte lysates reduces BKCa-Synpo interactions. Coexpression of Synpo increases steady-state surface expression of BKCa channels in HEK293T cells. However, Synpo does not affect the stability of cell surface BKCa channels, suggesting a primary effect on the rate of forward trafficking, and Synpo coexpression does not affect BKCa gating. Conversely, stable knockdown of Synpo expression in mouse podocyte cell lines reduces steady-state surface expression of BKCa channels but does not affect total expression of BKCa channels or their gating. The effects of Synpo on surface expression of BKCa are blocked by inhibition of Rho signaling in HEK293T cells and in podocytes. Functional cell surface BKCa channels in podocytes are also reduced by sustained (2 h) but not acute (15 min) depolymerization of actin with cytochalasin D. Synpo may regulate BKCa channels through its effects on actin dynamics and by modulating interactions between BKCa channels and regulatory proteins of the podocyte slit diaphragm.