1. Department of Nephrology, Jiangxi Provincial People's Hospital Affiliated to Nanchang University, Nanchang 330006, Jiangxi, China.
2. Department of Critical Care Medicine, Jiangxi Provincial People's Hospital Affiliated to Nanchang University, Nanchang 330006, Jiangxi, China.
Chronic renal failure (CRF), also known as chronic kidney disease (CKD), is a common renal disorder characterized by gradual kidney dysfunction. Molecular dissection reveals that transforming growth factor beta (TGF-β) plays a central role in the pathogenesis of CRF. However, the mechanism underlying TGF-β upregulation has not been demonstrated. Here, we verified that the elevated level of TGF-β was associated with the severity of CRF stages and the activation of TGF-β-mediated signaling in 120 renal biopsies from CRF patients. By analyzing the promoter region of the TGFB1 gene, we identified one AP-1 (activator protein 1) and four NF-κB (nuclear factor kappa-light-chain-enhancer of activated B cells) binding sites. Knockdown of two AP-1 subunits (c-Jun and c-FOS) or blockage of AP-1 signaling with two inhibitors T-5224 and SR11302 could cause the downregulation of TGFB1, whereas knockdown of two NF-κB subunits (p65 and p50) or blockage of NF-κB signaling with two inhibitors TPCA1 and BOT-64 could not change the expression of TGFB1. Using mass spectrometry and coimmunoprecipitation analyses, we found that both c-Jun and c-FOS formed a complex with CtBP2 (C-terminal binding protein 2) and histone acetyltransferase p300. Our in vitro data demonstrated that induction of CtBP2 by recombinant IL-1β (interleukin-1 beta) led to the upregulation of TGFB1 and the activation of TGF-β downstream signaling, while knockdown of CtBP2 resulted in the reversed effects. Using chromatin immunoprecipitation assays, we revealed that the CtBP2-p300-AP1 complex specifically bound to the promoter of TGFB and that knockdown or blockage of CtBP2 significantly decreased the occupancies of the p300 and AP-1 subunits. Our results support a model in which the CtBP2-p300-AP1 transcriptional complex activates the expression of TGFB1, increasing its production and extracellular secretion. The secreted TGF-β binds to its receptors and initiates downstream signaling.
Keywords: TGF-β, CtBP2, p300, c-Jun, c-FOS, chronic renal failure