The broad goal of this project is to contribute to understanding the cellular and molecular mechanisms that govern podocyte cytoskeletal dynamics with the expectation that understanding these processes will expose potential disease mechanisms and therapeutic targets. We have been guided by the hypothesis that precisely regulated cytoskeletal dynamics are essential during normal podocyte maturation, in maintenance of the differentiated podocyte phenotype, and in determining podocyte behavior in response to glomerular disease. While a decade of work has made this hypothesis dogma, a detailed understanding of mechanisms that govern podocyte cytoskeletal dynamics remains incomplete and therapeutic targets based on these mechanisms do not exist. Because integrated podocyte intercellular junction and foot process actin cytoskeletal remodeling are nearly always encountered in glomerular disease, we have focused on investigating the functions of the intercellular junction Nephrin-Neph1-Podocin complex because mutation or loss of any one of these receptor components causes proteinuria and podocyte cytoskeletal remodeling. The present proposal is based on our recent observations that podocyte foot process spreading is regulated by mechanisms that parallel those used by cultured cells to induce lamellipodial protrusion. Nephrin ligation induces lamellipodial activation by a focal adhesion kinase-p130Cas-Crk1/2-dependent pathway. In mice, podocyte-specific deletion of Crk1/2 blocks injury-induced foot process effacement. Phosphorylation of FAK and Cas are induced in podocytes of patients with human minimal change disease and in membranous nephropathy relative to normal tissue. These results provide compelling initial evidence that Crk-dependent signaling represents a therapeutic target that might be useful in blocking foot process effacement in human glomerular disease. Given these results and additional preliminary data suggesting a molecular mechanism by which Crk-dependent signaling is necessary in foot process spreading, this project will address the hypothesis that Nephrin-FAK-Cas-Crk signaling is necessary for foot process effacement and can be targeted in both acute and chronic glomerular disease processes.
Two specific aims are proposed.
In aim 1, we will examine in mechanistic detail the hypothesis that Nephrin signaling and function requires endocytosis and endocytic recycling and subsequent signaling from a Nephrin signaling endosome, a process that requires Crk1/2. In completed work, we have demonstrated that targeting Crk attenuates the glomerular disease phenotype in acute murine glomerular disease models.
In aim 2, we will extend this pre-clinical work by testing the hypothesis that targeting Crk-dependent signaling attenuates the disease phenotype associated with chronic glomerular disease models that more closely mimic progressive human chronic glomerulopathy.
Diseases resulting in the nephrotic syndrome such as diabetes mellitus and other diseases of the glomerulus frequently cause progressive kidney damage resulting in irreversible loss of renal function and account for nearly 60% of end-stage renal disease (ESRD) in the United States. While dialysis and kidney transplantation are effective treatments for ESRD, they are expensive and imperfect. Indeed, ESRD is associated with a mortality rate of 60% at five years. Moreover, the economic burden of ESRD treatment in the United States is approaching $30 billion dollars per year. Because little is understood about the biology of diseases of the glomerulus effective therapies are lacking. The goal here is to contribute to understanding the cellular and molecular mechanisms that govern the dynamics of podocyte structure and filter integrity with the expectation that providing a mechanistic understanding of these processes will expose potential disease mechanisms and therapeutic targets.
|Arif, Ehtesham; Rathore, Yogendra S; Kumari, Babita et al. (2014) Slit diaphragm protein Neph1 and its signaling: a novel therapeutic target for protection of podocytes against glomerular injury. J Biol Chem 289:9502-18|
|George, Britta; Fan, Qingfeng; Dlugos, Christopher P et al. (2014) Crk1/2 and CrkL form a hetero-oligomer and functionally complement each other during podocyte morphogenesis. Kidney Int 85:1382-94|
|George, Britta; Verma, Rakesh; Soofi, Abdulsalam A et al. (2012) Crk1/2-dependent signaling is necessary for podocyte foot process spreading in mouse models of glomerular disease. J Clin Invest 122:674-92|
|Mallik, Leena; Arif, Ehtesham; Sharma, Pankaj et al. (2012) Solution structure analysis of cytoplasmic domain of podocyte protein Neph1 using small/wide angle x-ray scattering (SWAXS). J Biol Chem 287:9441-53|
|Garg, Puneet; Holzman, Lawrence B (2012) Podocytes: gaining a foothold. Exp Cell Res 318:955-63|
|Holzman, Lawrence B; Garg, Puneet (2009) Initial insight on the determinants of podocyte polarity. J Am Soc Nephrol 20:683-5|