Involvement of the protein kinase C substrate, SSeCKS, in the actin-based stellate morphology of mesangial cells

Activation of protein kinase C is a key signal transduction event in mesang ial cell dedifferentiation and proliferation, yet little is known about dow nstream substrates or their roles in normal or diseased glomeruli, SSeCKS, a novel protein kinase C substrate originally isolated as a src-suppressed negative mitogenic regulator in fibroblasts, controls actin-based cytoskele tal architecture and scaffolds key signaling kinases such as protein kinase C and protein kinase A, Based on the morphologic similarity between SSeCKS -overexpressing fibroblasts and stellate mesangial tells, we hypothesized t hat SSeCKS might play a role in mesangial cell morphology in a protein kina se C-dependent manner. Immunoblotting, in situ staining and northern blotti ng detected abundant expression of SSeCKS in human and rodent mesangial cel ls and glomerular parietal cells but not in renal tubular epithelia, Immuno fluorescence analysis showed enrichment of SSeCKS in mesangial cell podosom es and along a cytoskeletal network distinct from F-actin, Activation of pr otein kinase C by phorbol ester resulted in a rapid serine phosphorylation of SSeCKS and its subsequent translocation to perinuclear sites, coincident with the retraction of stellate processes, These effects were blocked by c oncentrations of bis-indolylmaleimide that selectively inhibit protein kina se C, Finally, ablation of SSeCKS expression using retroviral anti-sense ve ctors induced (1) an elongated, fibroblastic cell morphology, (2) productio n of thick, longitudinal stress fibers and (3) repositioning of vinculin-as sociated focal complexes away from the cell edges, These data suggest a rol e for SSeCKS as a downstream mediator of protein kinase C-controlled, actin -based mesangial cell cytoskeletal architecture.