PKCα Mediates β-Arrestin2-dependent Nephrin Endocytosis in Hyperglycemia

Nephrin, the key molecule of the glomerular slit diaphragm, is expressed on the surface of podocytes and is critical in preventing albuminuria. In diabetes, hyperglycemia leads to the loss of surface expression of nephrin and causes albuminuria. Here, we report a mechanism that can explain this phenomenon: hyperglycemia directly enhances the rate of nephrin endocytosis via regulation of the beta-arrestin2-nephrin interaction by PKC alpha. We identified PKC alpha and protein interacting with c kinase-1 (PICK1) as nephrin-binding proteins. Hyperglycemia induced up-regulation of PKC alpha and led to the formation of a complex of nephrin, PKC alpha, PICK1, and beta-arrestin2 in vitro and in vivo. Binding of beta-arrestin2 to the nephrin intracellular domain depended on phosphorylation of nephrin threonine residues 1120 and 1125 by PKC alpha. Further, cellular knockdown of PKC alpha and/or PICK1 attenuated the nephrin beta-arrestin2 interaction and abrogated the amplifying effect of high blood glucose on nephrin endocytosis. In C57BL/6 mice, hyperglycemia over 24 h caused a significant increase in urinary albumin excretion, supporting the concept of the rapid impact of hyperglycemia on glomerular permselectivity. In summary, we have provided a molecular model of hyperglycemia-induced nephrin endocytosis and subsequent proteinuria and highlighted PKC alpha and PICK1 as promising therapeutic targets for diabetic nephropathy.