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Signalling cross-talk between hepatocyte nuclear factor 4alpha and growth-hormone-activated STAT5b.

ABSTRACT: In the present study, we have characterized signalling cross-talk between STAT5b (signal transducer and activator of transcription 5b) and HNF4alpha (hepatocyte nuclear factor 4alpha), two major regulators of sex-dependent gene expression in the liver. In a HepG2 liver cell model, HNF4alpha strongly inhibited beta-casein and ntcp (Na+/taurocholate cotransporting polypeptide) promoter activity stimulated by GH (growth hormone)-activated STAT5b, but had no effect on interferon-gamma-stimulated STAT1 transcriptional activity. By contrast, STAT5b synergistically enhanced the transcriptional activity of HNF4alpha towards the ApoCIII (apolipoprotein CIII) promoter. The inhibitory effect of HNF4alpha on STAT5b transcription was associated with the inhibition of GH-stimulated STAT5b tyrosine phosphorylation and nuclear translocation. The short-chain fatty acid, butyrate, reversed STAT5b transcriptional inhibition by HNF4alpha, but did not reverse the inhibition of STAT5b tyrosine phosphorylation. HNF4alpha inhibition of STAT5b tyrosine phosphorylation was not reversed by pervanadate or by dominant-negative phosphotyrosine phosphatase 1B, suggesting that it does not result from an increase in STAT5b dephosphorylation. Rather, HNF4alpha blocked GH-stimulated tyrosine phosphorylation of JAK2 (Janus kinase 2), a STAT5b tyrosine kinase. Thus STAT5b and HNF4alpha exhibit bi-directional cross-talk that may augment HNF4alpha-dependent gene transcription while inhibiting STAT5b transcriptional activity via the inhibitory effects of HNF4alpha on JAK2 phosphorylation, which leads to inhibition of STAT5b signalling initiated by the GH receptor at the cell surface.

SUBMITTER: Park SH

PROVIDER: S-EPMC1479742 | biostudies-other | 2006 Jul

REPOSITORIES: biostudies-other

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Regulatory mechanisms controlling human hepatocyte nuclear factor 4alpha gene expression.

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Hepatocyte nuclear factor-4alpha is a central transactivator of the mouse Ntcp gene.

Project description:Sodium taurocholate cotransporting polypeptide (Ntcp) is the major uptake system for conjugated bile acids. Deletions of hepatocyte nuclear factor (HNF)-1alpha and retinoid X receptor-alpha:retinoic acid receptor-alpha binding sites in the mouse 5'-flanking region corresponding to putatively central regulatory elements of rat Ntcp do not significantly reduce promoter activity. We hypothesized that HNF-4alpha, which is increasingly recognized as a central regulator of hepatocyte function, may directly transactivate mouse (mNtcp). A 1.1-kb 5'-upstream region including the mouse Ntcp promoter was cloned and compared with the rat promoter. In contrast to a moderate 3.5-fold activation of mNtcp by HNF-1alpha, HNF-4alpha cotransfection led to a robust 20-fold activation. Deletion analysis of mouse and rat Ntcp promoters mapped a conserved HNF-4alpha consensus site at -345/-326 and -335/-316 bp, respectively. p-475bpmNtcpLUC is not transactivated by HNF-1alpha but shows a 50-fold enhanced activity upon cotransfection with HNF-4alpha. Gel mobility shift assays demonstrated a complex of the HNF-4alpha-element formed with liver nuclear extracts that was blocked by an HNF-4alpha specific antibody. HNF-4alpha binding was confirmed by chromatin immunoprecipitation. Using Hepa 1-6 cells, HNF-4alpha-knockdown resulted in a significant 95% reduction in NTCP mRNA. In conclusion, mouse Ntcp is regulated by HNF-4alpha via a conserved distal cis-element independently of HNF-1alpha.

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