The role of Cofilin-1 in regulating podocyte actin dynamics and maintaining foot process morphology
Final Report Abstract
The slit diaphragm protein Nephrin is necessary for establishing the morphology of the podocyte in development by transducing from the podocyte intercellular junction phosphorylationmediated signals that regulate cytoskeletal dynamics. While investigating the mechanisms that mediate this function, we found that activation of Nephrin induced lamellipodia formation in cell culture. We also found that the cytoskeletal adaptor protein Crk1/2 interacted with Nephrin in a tyrosine phosphorylation-dependent fashion following Nephrin activation. Crk1/2 was necessary for Nephrin-induced lamellipodia dynamics in cultured podocytes since RNAi-mediated knockdown of Crk1/2 attenuated cell spreading. In other systems, Crk 1/2 functions in complex with Cas within the focal adhesion complex to mediate lamellipodial-dynamics. Similarly, in the podocyte, tyrosine-phosphorylated Cas was recruited to Nephrin following its activation in a pI3- and Src kinase-dependent manner. Recruitment of Crk to the Nephrin-Cas complex did not occur in Cas null mouse embryonic fibroblasts, but did occur upon rescue of Cas expression. Like Crk1/2, Cas was necessary for Nephrin-induced lamellipodia dynamics since cell spreading was attenuated in Cas-deficient fibroblasts. In developing mouse glomeruli we verified that phospho-Cas was present at the podocyte precursor adherens junction. To test the relevance of Crk1/2 signaling, mice deleted of Crk1/2 in a podocyte-specific fashion were bred and found to develop and age normally. Surprisingly, podocytes in these mice were protected from protamine sulfate (PS)-induced foot process spreading. Furthermore, Crk complex proteins were activated in podocytes in human proteinuric kidney disease such as minimal change disease and membranous nephropathy compared to healthy controls as shown by tissue immunofluorescence analysis. We conclude that Crk interacts with Nephrin indirectly, through Cas, and plays an essential role in Nephrin-induced lamellipodia formation. We speculate that podocyte effacement observed in the PS model results from induction of lamellipodial-like Crk-dependent actin dynamics in the foot process. Altogether, the Crk protein complex impresses as a promising drug targetable site in proteinuric kidney disease.
Publications
- (2010). Crk1/2 Is Necessary for Nephrin-Induced Lamellipodia Formation in Podocyte Culture and Kidney Podocyte Foot Process Spreading in Mice. Mol. Biol. Cell 21 (suppl), 1881
George B, Verma R, Soofi AA, and Holzman LB
- Actin-depolymerizing factor cofilin-1 is necessary in maintaining mature podocyte architecture. J Biol Chem. 2010 Jul 16;285(29):22676-88
Garg P, Verma R, Cook L, Soofi A, Venkatareddy M, George B, Mizuno K, Gurniak C, Witke W, Holzman LB
- Crk1/2 forms a Complex with Nephrin and p130Cas and Is Essential for Nephrin-Induced Cell Spreading. Renal Week, 2010 Nov 16-21, Denver, CO, F-PO1872
George B, Verma R, Soofi AA, and Holzman LB
- Podocyte-Specific Deletion of MYH9 Predisposes Mice to Nephropathy. Renal Week, 2010 Nov 16-21, Denver, CO, SA-FC396
Johnstone DB, Zhang J, George B, Holzman LB
- Motor protein My1c is a podocyte protein that facilitates the transport of slit diaphragm protein Neph1 to podocyte membrane. Mol Cell Biol. 2011 Mar 14. (Epub ahead of print)
Arif E, Wagner MC, Johnstone DB, Wong HN, George B, Pruthi PA, Lazarra MJ, Nihalani D
- Podocyte-specific deletion of Myh9 encoding non-muscle myosin heavy chain 2A predisposes mice to glomerulopathy. Mol Cell Biol. 2011 Mar 14. (Epub ahead of print)
Johnstone DB, Zhang J, George B, Leon C, Gachet C, Wong H, Parekh R, Holzman LB
