Glomerulosclerosis is the most serious sequela of progressive renal disease. Glomerulosclerosis results from an abnormal accumulation of proteins, some of which, that normally make up the glomerular basement membrane (GBM) and mesangial matrix (MM). Laminin is one of the components of GBM and MM that accumulates within glomeruli during the progression of glomerular disease. A number of growth factors, including transforming growth factor-beta (TGF-beta), activate glomerular cells resulting in abnormal accumulation of laminin. Although laminin is known to play a key role in the development of glomerulosclerosis, the mechanisms that regulate expression of laminin chains are incompletely understood. Treatment of glomerular cells with TGF-beta and other growth factors increases mRNA levels of laminin gamma1 chain. The human and rodent laminin gamma1 chain (LAMC1) gene promoters contain the critical bcn-1 element that is activated by transcription factor uE3 (TFE3). In glomerular mesangial cells, the TFE3-mediated activation of the bcn-1 element is greatly augmented by Smad proteins, acting through the LAMC1 promoter's Smad binding elements (SBE), and by the TGF-beta-signaling pathways. The bcn-1-element-dependent activation of the LAMC1 promoter by the synergistic action of TFE3 and Smad proteins provides insight into how TGF-beta mediates activation of the endogenous LAMC1 gene in these cells. Expression of the transcriptional co-activator, CREB binding protein (CBP), increased the activity of the LAMC1 promoter in mesangial cells. The goal of the current proposal is to define the molecular mechanisms responsible for the TFE3- and Smad-dependent activation of LAMC 1 gene transcription that is induced by TGF-beta in glomerular cells. The following specific aims are proposed to explore these mechanisms. We will define the role of CBP and other co-activators in mediating the TFE3-Smad-dependent TGF-beta-induced LAMC1 gene expression. We will identify components of the basal transcriptional machinery that interact with TFE3 protein and define their role in mediating TGF-beta-induced gene expression. We will define the molecular mechanisms responsible for the TFE3-Smad3 synergistic activation of the LAMC1 gene in response to TGF-beta. These studies will provide insight into transcriptional mechanisms utilized by TGF-beta to direct TFE3- and Smad-dependent LAMC1 gene transcription in glomerular cells. The results of these studies are anticipated to have a broad impact on understanding the basic mechanisms by which TGF-beta triggers transcription of genes encoding components of extracellular matrix and of other genes expressed in glomerular cells.
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