Skip to main content
PMC home page
Nucleic Acids Research logoLink to Nucleic Acids Research
. 1998 Jun 1;26(11):2702–2707. doi: 10.1093/nar/26.11.2702

DNA binding and transcription activation specificity of hepatocyte nuclear factor 4.

J D Fraser 1, V Martinez 1, R Straney 1, M R Briggs 1
PMCID: PMC147588  PMID: 9592157

Abstract

Hepatocyte nuclear factor 4 (HNF-4) is an orphan intracellular receptor that appears to be a key factor in the regulation of energy metabolism. In order to gain greater understanding of the binding and activation requirements of HNF-4, we performed genetic analysis of the apoCIII promoter, a promoter that has previously been shown to be highly sensitive to HNF-4-induced transcription. We identified two elements within the apoCIII promoter that bind HNF-4, either of which are sufficient to confer promoter induction in response to HNF-4. These two elements are both direct repeat-like in nature, but they differ significantly in motif sequence and the repeats are separated by either 1 or 2 nt. Therefore, to better define the DNA sequence recognition requirements of HNF-4, we utilized PCR-based binding site selection. HNF-4 selected for direct repeat-like elements with either 1 or 2 nt between the repeats. Surprisingly, the strongest selection was for a core motif that included the nucleotides between the repeats. Mutation of the nucleotide between the repeats resulted in a 6-fold reduction in affinity, indicating that the interaction between HNF-4 and the intervening nucleotide(s) is critical for high affinity binding.

Full Text

The Full Text of this article is available as a PDF (199.4 KB).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Bisaha J. G., Simon T. C., Gordon J. I., Breslow J. L. Characterization of an enhancer element in the human apolipoprotein C-III gene that regulates human apolipoprotein A-I gene expression in the intestinal epithelium. J Biol Chem. 1995 Aug 25;270(34):19979–19988. doi: 10.1074/jbc.270.34.19979. [DOI] [PubMed] [Google Scholar]
  2. Bradford M. M. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem. 1976 May 7;72:248–254. doi: 10.1006/abio.1976.9999. [DOI] [PubMed] [Google Scholar]
  3. Fraser J. D., Keller D., Martinez V., Santiso-Mere D., Straney R., Briggs M. R. Utilization of recombinant adenovirus and dominant negative mutants to characterize hepatocyte nuclear factor 4-regulated apolipoprotein AI and CIII expression. J Biol Chem. 1997 May 23;272(21):13892–13898. doi: 10.1074/jbc.272.21.13892. [DOI] [PubMed] [Google Scholar]
  4. Freedman L. P. Anatomy of the steroid receptor zinc finger region. Endocr Rev. 1992 May;13(2):129–145. doi: 10.1210/edrv-13-2-129. [DOI] [PubMed] [Google Scholar]
  5. Giguère V., Tini M., Flock G., Ong E., Evans R. M., Otulakowski G. Isoform-specific amino-terminal domains dictate DNA-binding properties of ROR alpha, a novel family of orphan hormone nuclear receptors. Genes Dev. 1994 Mar 1;8(5):538–553. doi: 10.1101/gad.8.5.538. [DOI] [PubMed] [Google Scholar]
  6. Ginsburg G. S., Ozer J., Karathanasis S. K. Intestinal apolipoprotein AI gene transcription is regulated by multiple distinct DNA elements and is synergistically activated by the orphan nuclear receptor, hepatocyte nuclear factor 4. J Clin Invest. 1995 Jul;96(1):528–538. doi: 10.1172/JCI118065. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Hadzopoulou-Cladaras M., Kistanova E., Evagelopoulou C., Zeng S., Cladaras C., Ladias J. A. Functional domains of the nuclear receptor hepatocyte nuclear factor 4. J Biol Chem. 1997 Jan 3;272(1):539–550. doi: 10.1074/jbc.272.1.539. [DOI] [PubMed] [Google Scholar]
  8. Ho S. N., Hunt H. D., Horton R. M., Pullen J. K., Pease L. R. Site-directed mutagenesis by overlap extension using the polymerase chain reaction. Gene. 1989 Apr 15;77(1):51–59. doi: 10.1016/0378-1119(89)90358-2. [DOI] [PubMed] [Google Scholar]
  9. Jiang G., Nepomuceno L., Hopkins K., Sladek F. M. Exclusive homodimerization of the orphan receptor hepatocyte nuclear factor 4 defines a new subclass of nuclear receptors. Mol Cell Biol. 1995 Sep;15(9):5131–5143. doi: 10.1128/mcb.15.9.5131. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Kastner P., Mark M., Chambon P. Nonsteroid nuclear receptors: what are genetic studies telling us about their role in real life? Cell. 1995 Dec 15;83(6):859–869. doi: 10.1016/0092-8674(95)90202-3. [DOI] [PubMed] [Google Scholar]
  11. Katz R. W., Koenig R. J. Nonbiased identification of DNA sequences that bind thyroid hormone receptor alpha 1 with high affinity. J Biol Chem. 1993 Sep 15;268(26):19392–19397. [PubMed] [Google Scholar]
  12. Ktistaki E., Lacorte J. M., Katrakili N., Zannis V. I., Talianidis I. Transcriptional regulation of the apolipoprotein A-IV gene involves synergism between a proximal orphan receptor response element and a distant enhancer located in the upstream promoter region of the apolipoprotein C-III gene. Nucleic Acids Res. 1994 Nov 11;22(22):4689–4696. doi: 10.1093/nar/22.22.4689. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Kuo C. J., Conley P. B., Chen L., Sladek F. M., Darnell J. E., Jr, Crabtree G. R. A transcriptional hierarchy involved in mammalian cell-type specification. Nature. 1992 Jan 30;355(6359):457–461. doi: 10.1038/355457a0. [DOI] [PubMed] [Google Scholar]
  14. Kurokawa R., Yu V. C., När A., Kyakumoto S., Han Z., Silverman S., Rosenfeld M. G., Glass C. K. Differential orientations of the DNA-binding domain and carboxy-terminal dimerization interface regulate binding site selection by nuclear receptor heterodimers. Genes Dev. 1993 Jul;7(7B):1423–1435. doi: 10.1101/gad.7.7b.1423. [DOI] [PubMed] [Google Scholar]
  15. Ladias J. A., Hadzopoulou-Cladaras M., Kardassis D., Cardot P., Cheng J., Zannis V., Cladaras C. Transcriptional regulation of human apolipoprotein genes ApoB, ApoCIII, and ApoAII by members of the steroid hormone receptor superfamily HNF-4, ARP-1, EAR-2, and EAR-3. J Biol Chem. 1992 Aug 5;267(22):15849–15860. [PubMed] [Google Scholar]
  16. Mangelsdorf D. J., Evans R. M. The RXR heterodimers and orphan receptors. Cell. 1995 Dec 15;83(6):841–850. doi: 10.1016/0092-8674(95)90200-7. [DOI] [PubMed] [Google Scholar]
  17. Metzger S., Halaas J. L., Breslow J. L., Sladek F. M. Orphan receptor HNF-4 and bZip protein C/EBP alpha bind to overlapping regions of the apolipoprotein B gene promoter and synergistically activate transcription. J Biol Chem. 1993 Aug 5;268(22):16831–16838. [PubMed] [Google Scholar]
  18. Mietus-Snyder M., Sladek F. M., Ginsburg G. S., Kuo C. F., Ladias J. A., Darnell J. E., Jr, Karathanasis S. K. Antagonism between apolipoprotein AI regulatory protein 1, Ear3/COUP-TF, and hepatocyte nuclear factor 4 modulates apolipoprotein CIII gene expression in liver and intestinal cells. Mol Cell Biol. 1992 Apr;12(4):1708–1718. doi: 10.1128/mcb.12.4.1708. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Miquerol L., Lopez S., Cartier N., Tulliez M., Raymondjean M., Kahn A. Expression of the L-type pyruvate kinase gene and the hepatocyte nuclear factor 4 transcription factor in exocrine and endocrine pancreas. J Biol Chem. 1994 Mar 25;269(12):8944–8951. [PubMed] [Google Scholar]
  20. Sladek F. M., Zhong W. M., Lai E., Darnell J. E., Jr Liver-enriched transcription factor HNF-4 is a novel member of the steroid hormone receptor superfamily. Genes Dev. 1990 Dec;4(12B):2353–2365. doi: 10.1101/gad.4.12b.2353. [DOI] [PubMed] [Google Scholar]
  21. Stoffel M., Duncan S. A. The maturity-onset diabetes of the young (MODY1) transcription factor HNF4alpha regulates expression of genes required for glucose transport and metabolism. Proc Natl Acad Sci U S A. 1997 Nov 25;94(24):13209–13214. doi: 10.1073/pnas.94.24.13209. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Tsai M. J., O'Malley B. W. Molecular mechanisms of action of steroid/thyroid receptor superfamily members. Annu Rev Biochem. 1994;63:451–486. doi: 10.1146/annurev.bi.63.070194.002315. [DOI] [PubMed] [Google Scholar]
  23. Yamagata K., Furuta H., Oda N., Kaisaki P. J., Menzel S., Cox N. J., Fajans S. S., Signorini S., Stoffel M., Bell G. I. Mutations in the hepatocyte nuclear factor-4alpha gene in maturity-onset diabetes of the young (MODY1) Nature. 1996 Dec 5;384(6608):458–460. doi: 10.1038/384458a0. [DOI] [PubMed] [Google Scholar]

Articles from Nucleic Acids Research are provided here courtesy of Oxford University Press

RESOURCES