Regulation of nephrin gene by the Ets transcription factor, GA-binding protein

Abstract

Transcription factor GA-binding protein (GABP) is suggested to be involved in the formation of the neuromuscular junctions by regulating the transcription of synapse genes. Interestingly, neurons and podocytes share molecular and functional similarities that led us to investigate the expression and function of GABP in podocytes and its role in transcriptional regulation of nephrin, the key molecule of the podocyte slit diaphragm that is essential for normal glomerular ultrafiltration. The expression and localization of GABP in the rat and human kidney as well as in human embryonic kidney A293 cells and undifferentiated and differentiated human podocytes were analysed by immunoblotting and immunostaining. The role of GABP in activating the nephrin promoter was investigated by reporter gene assay and site-directed mutagenesis of the GABP-binding elements, and the interaction of GABP with the nephrin promoter was analysed by chromatin immunoprecipitation. The function of GABP in podocytes was studied by knocking down GABPα in differentiated human podocytes using lentiviral shRNA targeting GABPα. GABP is expressed in the nuclei in rat and human glomeruli. In addition, in A293 cells and undifferentiated and differentiated human podocytes, GABP highly enriches in the nucleus. GABP activates and binds nephrin proximal promoter and Ets sites are essential for this activity. Knock-down of GABPα stimulates apoptosis in cultured podocytes. The results show that GABP is expressed in podocytes and is involved in the regulation of nephrin gene expression. Furthermore, GABP may be important in the maintenance of podocyte function by regulating apoptosis.