Hepatocyte nuclear factor-4alpha mediates redox sensitivity of inducible nitric-oxide synthase gene transcription.

The underlying redox-sensitive mechanisms that regulate hepatocyte expression of inducible nitric-oxide synthase (iNOS) and its antioxidant functions are largely unknown. We have demonstrated previously that oxidative stress induced by benzenetriol-mediated superoxide production increases interleukin-1beta-induced iNOS protein synthesis, steady state iNOS mRNA expression, NO production, iNOS gene transcription, and trans-activation of the iNOS promoter in primary cultures of rat hepatocytes. In this study, we extend these studies by establishing the sequence specificity and binding of nuclear protein to the previously described 15-base cis-regulatory element of the rat hepatocyte iNOS promoter, isolating and identifying the cis-regulatory element transcription factor as hepatocyte nuclear factor-4alpha (HNF-4alpha), and confirming the functional role of HNF-4alpha in mediating redox-sensitive iNOS promoter trans-activation. In addition, we demonstrate that binding of HNF-4alpha to the transcriptional coactivator, PC4, in the presence of oxidative stress and interleukin-1beta stimulation is essential for increased iNOS promoter activity in this setting. Our results indicate that HNF-4alpha is the transcription factor that mediates redox regulation of hepatocyte iNOS gene transcription.