Inhibition of hepatocyte nuclear factor 4 transcriptional activity by the nuclear factor kappaB pathway.
HNF-4 (hepatocyte nuclear factor 4) is a key regulator of liver-specific gene expression in mammals. We have shown previously that the activity of the human APOC3 (apolipoprotein C-III) promoter is positively regulated by the anti-inflammatory cytokine TGFbeta (transforming growth factor beta) and its effectors Smad3 (similar to mothers against decapentaplegic 3) and Smad4 proteins via physical and functional interactions between Smads and HNF-4. We now show that the pro-inflammatory cytokine TNFalpha (tumour necrosis factor alpha) antagonizes TGFbeta for the regulation of APOC3 gene expression in hepatocytes. TNFalpha was a strong inhibitor of the activity of apolipoprotein promoters that harbour HNF-4 binding sites and this inhibition required HNF-4. Using specific inhibitors of TNFalpha-induced signalling pathways, it was shown that inhibition of the APOC3 promoter by TNFalpha involved NF-kappaB (nuclear factor kappaB). Latent membrane protein 1 of the Epstein-Barr virus, which is an established potent activator of NF-kappaB as well as wild-type forms of various NF-kappaB signalling mediators, also inhibited strongly the APOC3 promoter and the transactivation function of HNF-4. TNFalpha had no effect on the stability or the nuclear localization of HNF-4 in HepG2 cells, but inhibited the binding of HNF-4 to the proximal APOC3 HRE (hormone response element). Using the yeast-transactivator-GAL4 system, we showed that both AF-1 and AF-2 (activation functions 1 and 2) of HNF-4 are inhibited by TNFalpha and that this inhibition was abolished by overexpression of different HNF-4 co-activators, including PGC-1 (peroxisome-proliferator-activated-receptor-gamma co-activator 1), CBP [CREB (cAMP-response-element-binding protein) binding protein] and SRC3 (steroid receptor co-activator 3). In summary, our findings indicate that TNFalpha, or other factors that trigger an NF-kappaB response in hepatic cells, inhibit the transcriptional activity of the APOC3 and other HNF-4-dependent promoters and that this inhibition could be accounted for by a decrease in DNA binding and the down-regulation of the transactivation potential of the AF-1 and AF-2 domains of HNF-4.
Nikolaidou-Neokosmidou
Varvara
V
Department of Basic Sciences, University of Crete Medical School and Institute of Molecular Biology and Biotechnology, Foundation of Research and Technology Hellas, Heraklion 71003, Crete, Greece.
Zannis
Vassilis I
VI
Kardassis
Dimitris
D
D000818:Animals; D053305:Apolipoprotein C-III; D001056:Apolipoproteins C / Q000235:genetics / Q000378:metabolism; D002460:Cell Line; D002522:Chlorocebus aethiops; D051557:Hepatocyte Nuclear Factor 4 / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D016328:NF-kappa B / Q000378:metabolism*; D011401:Promoter Regions, Genetic; D017434:Protein Structure, Tertiary; D014158:Transcription, Genetic / Q000502:physiology*; D015533:Transcriptional Activation; D016212:Transforming Growth Factor beta / Q000378:metabolism; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism
D053305:Apolipoprotein C-III; D001056:Apolipoproteins C; D051557:Hepatocyte Nuclear Factor 4; D016328:NF-kappa B; D016212:Transforming Growth Factor beta; D014409:Tumor Necrosis Factor-alpha