Signaling Pathways in Renal Fibrogenesis

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DESCRIPTION (provided by applicant): The signaling mechanisms underlying kidney fibrosis continue to be of significant concern and interest. This proposal presents a strategy for extending our previous studies that examined how TGF-2 stimulates accumulation of type I collagen in a cellular model of renal fibrogenesis. In the previous grant period, we demonstrated that numerous signaling pathways interact with TGF-2/Smad signaling, focusing particularly on the role of the extracellular signal-regulated kinase (ERK) MAP kinase. We determined that the Smad3 and ERK pathways together promote TGF-2-stimulated collagen expression in renal mesangial and tubular epithelial cells. ERK serves a critical role in supporting Smad3-mediated collagen gene transcription, but does not activate the COL1A2 promoter in the absence of Smad activation; it is thus necessary but not sufficient for collagen expression. ERK phosphorylates serines and threonines in the linker region (LR) domain of Smad3. Additional Preliminary Studies indicate that blocking 1v23 integrin or PI3-kinase activity inhibits ERK activation and the collagen response. For this renewal, we propose the HYPOTHESIS that an adhesion-dependent signaling pathway from 1v23-integrin to ERK promotes renal cell type I collagen expression through phosphorylation of linker-region serine and/or threonine residues of Smad3; and PI3K, which acts at several points along the pathway leading to ERK, is a promising target for blunting the collagen response to TGF-2 both in vitro and in vivo. To test this hypothesis, we will pursue the following AIMS: First, we will determine the molecular mechanisms by which 1v23 integrin promotes TGF-2-stimulated renal cell fibrogenesis, establishing the roles of Rac1 and PI3K in this process. Second, we will determine the mechanism(s) through which ERK and Smad3 synergize to promote TGF-2-stimulated collagen expression by examining how specific phosphorylation events affect Smad3 signaling of collagen I expression and which of these phosphorylations are dependent upon PI3K and/or ERK. Third, we will determine how 1v23, PI3K and ERK interact to modulate the Smad3 LR in the renal fibrosis models of adriamycin nephropathy and selective podocyte ablation. These studies should provide new insights into integrin regulation of Smad signaling, address unsettled issues regarding the role of the Smad3 LR, and offer potential approaches to interrupting the fibrogenic process in kidneys. SIGNIFICANCE: Despite extensive research, little is known about how TGF-2 signaling is regulated to cause renal fibrosis. Our studies of non-canonical signaling pathways and the role of Smad3-LR will address key issues regarding the specificity of signals in fibrogenesis. By elucidating the signaling events underlying kidney fibrosis we hope to better understand fibrogenic mechanisms, how to interrupt them and, ultimately, how to treat them.