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. 1988 Apr;8(4):1398–1407. doi: 10.1128/mcb.8.4.1398

Enhancer and promoter elements directing activation and glucocorticoid repression of the alpha 1-fetoprotein gene in hepatocytes.

M Guertin 1, H LaRue 1, D Bernier 1, O Wrange 1, M Chevrette 1, M C Gingras 1, L Bélanger 1
PMCID: PMC363296  PMID: 2454390

Abstract

Mutations were introduced in 7 kilobases of 5'-flanking rat alpha 1-fetoprotein (AFP) genomic DNA, linked to the chloramphenicol acetyltransferase gene. AFP promoter activity and its repression by a glucocorticoid hormone were assessed by stable and transient expression assays. Stable transfection assays were more sensitive and accurate than transient expression assays in a Morris 7777 rat hepatoma recipient (Hepa7.6), selected for its strong AFP repression by dexamethasone. The segment of DNA encompassing a hepatocyte-constitutive chromatin DNase I-hypersensitive site at -3.7 kilobases and a liver developmental stage-specific site at -2.5 kilobases contains interacting enhancer elements sufficient for high AFP promoter activity in Hepa7.6 or HepG2 cells. Deletions and point mutations define an upstream promoter domain of AFP gene activation, operating with at least three distinct promoter-activating elements, PEI at -65 base pairs, PEII at -120 base pairs, and DE at -160 base pairs. PEI and PEII share homologies with albumin promoter sequences, PEII is a near-consensus nuclear factor I recognition sequence, and DE overlaps a glucocorticoid receptor recognition sequence. An element conferring glucocorticoid repression of AFP gene activity is located in the upstream AFP promoter domain. Receptor-binding assays indicate that this element is the glucocorticoid receptor recognition sequence which overlaps with promoter-activating element DE.

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Selected References

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

  1. Abelev G. I. Alpha-fetoprotein in ontogenesis and its association with malignant tumors. Adv Cancer Res. 1971;14:295–358. doi: 10.1016/s0065-230x(08)60523-0. [DOI] [PubMed] [Google Scholar]
  2. Belanger L., Baril P., Guertin M., Gingras M. C., Gourdeau H., Anderson A., Hamel D., Boucher J. M. Oncodevelopmental and hormonal regulation of alpha 1-fetoprotein gene expression. Adv Enzyme Regul. 1983;21:73–99. doi: 10.1016/0065-2571(83)90009-2. [DOI] [PubMed] [Google Scholar]
  3. Belanger L., Hamel D., Lachance L., Dufour D., Tremblay M., Gagnon P. M. Hormonal regulation of alpha1 foetoprotein. Nature. 1975 Aug 21;256(5519):657–659. doi: 10.1038/256657a0. [DOI] [PubMed] [Google Scholar]
  4. Camper S. A., Yao Y. A., Rottman F. M. Hormonal regulation of the bovine prolactin promoter in rat pituitary tumor cells. J Biol Chem. 1985 Oct 5;260(22):12246–12251. [PubMed] [Google Scholar]
  5. Cereghini S., Raymondjean M., Carranca A. G., Herbomel P., Yaniv M. Factors involved in control of tissue-specific expression of albumin gene. Cell. 1987 Aug 14;50(4):627–638. doi: 10.1016/0092-8674(87)90036-5. [DOI] [PubMed] [Google Scholar]
  6. Chandler V. L., Maler B. A., Yamamoto K. R. DNA sequences bound specifically by glucocorticoid receptor in vitro render a heterologous promoter hormone responsive in vivo. Cell. 1983 Jun;33(2):489–499. doi: 10.1016/0092-8674(83)90430-0. [DOI] [PubMed] [Google Scholar]
  7. Chevrette M., Guertin M., Turcotte B., Bélanger L. The rat alpha 1-fetoprotein gene: characterization of the 5'-flanking region and tandem organization with the albumin gene. Nucleic Acids Res. 1987 Feb 11;15(3):1338–1339. doi: 10.1093/nar/15.3.1338. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Cordingley M. G., Riegel A. T., Hager G. L. Steroid-dependent interaction of transcription factors with the inducible promoter of mouse mammary tumor virus in vivo. Cell. 1987 Jan 30;48(2):261–270. doi: 10.1016/0092-8674(87)90429-6. [DOI] [PubMed] [Google Scholar]
  9. Eberwine J. H., Roberts J. L. Glucocorticoid regulation of pro-opiomelanocortin gene transcription in the rat pituitary. J Biol Chem. 1984 Feb 25;259(4):2166–2170. [PubMed] [Google Scholar]
  10. Eissenberg J. C., Cartwright I. L., Thomas G. H., Elgin S. C. Selected topics in chromatin structure. Annu Rev Genet. 1985;19:485–536. doi: 10.1146/annurev.ge.19.120185.002413. [DOI] [PubMed] [Google Scholar]
  11. Erdos T., Best-Belpomme M., Bessada R. A rapid assay for binding estradiol to uterine receptor(s). Anal Biochem. 1970 Oct;37(2):244–252. doi: 10.1016/0003-2697(70)90044-8. [DOI] [PubMed] [Google Scholar]
  12. Falkner F. G., Mocikat R., Zachau H. G. Sequences closely related to an immunoglobulin gene promoter/enhancer element occur also upstream of other eukaryotic and of prokaryotic genes. Nucleic Acids Res. 1986 Nov 25;14(22):8819–8827. doi: 10.1093/nar/14.22.8819. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Gagner J. P., Drouin J. Opposite regulation of pro-opiomelanocortin gene transcription by glucocorticoids and CRH. Mol Cell Endocrinol. 1985 Apr;40(1):25–32. doi: 10.1016/0303-7207(85)90154-6. [DOI] [PubMed] [Google Scholar]
  14. Gibbs P. E., Zielinski R., Boyd C., Dugaiczyk A. Structure, polymorphism, and novel repeated DNA elements revealed by a complete sequence of the human alpha-fetoprotein gene. Biochemistry. 1987 Mar 10;26(5):1332–1343. doi: 10.1021/bi00379a020. [DOI] [PubMed] [Google Scholar]
  15. Godbout R., Ingram R., Tilghman S. M. Multiple regulatory elements in the intergenic region between the alpha-fetoprotein and albumin genes. Mol Cell Biol. 1986 Feb;6(2):477–487. doi: 10.1128/mcb.6.2.477. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Gorman C. M., Moffat L. F., Howard B. H. Recombinant genomes which express chloramphenicol acetyltransferase in mammalian cells. Mol Cell Biol. 1982 Sep;2(9):1044–1051. doi: 10.1128/mcb.2.9.1044. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Gorski K., Carneiro M., Schibler U. Tissue-specific in vitro transcription from the mouse albumin promoter. Cell. 1986 Dec 5;47(5):767–776. doi: 10.1016/0092-8674(86)90519-2. [DOI] [PubMed] [Google Scholar]
  18. Guertin M., Baril P., Bartkowiak J., Anderson A., Bélanger L. Rapid suppression of alpha 1-fetoprotein gene transcription by dexamethasone in developing rat liver. Biochemistry. 1983 Aug 30;22(18):4296–4302. doi: 10.1021/bi00287a021. [DOI] [PubMed] [Google Scholar]
  19. Hammer R. E., Krumlauf R., Camper S. A., Brinster R. L., Tilghman S. M. Diversity of alpha-fetoprotein gene expression in mice is generated by a combination of separate enhancer elements. Science. 1987 Jan 2;235(4784):53–58. doi: 10.1126/science.2432657. [DOI] [PubMed] [Google Scholar]
  20. Heard J. M., Herbomel P., Ott M. O., Mottura-Rollier A., Weiss M., Yaniv M. Determinants of rat albumin promoter tissue specificity analyzed by an improved transient expression system. Mol Cell Biol. 1987 Jul;7(7):2425–2434. doi: 10.1128/mcb.7.7.2425. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Hochschild A., Ptashne M. Cooperative binding of lambda repressors to sites separated by integral turns of the DNA helix. Cell. 1986 Mar 14;44(5):681–687. doi: 10.1016/0092-8674(86)90833-0. [DOI] [PubMed] [Google Scholar]
  22. Jantzen H. M., Strähle U., Gloss B., Stewart F., Schmid W., Boshart M., Miksicek R., Schütz G. Cooperativity of glucocorticoid response elements located far upstream of the tyrosine aminotransferase gene. Cell. 1987 Apr 10;49(1):29–38. doi: 10.1016/0092-8674(87)90752-5. [DOI] [PubMed] [Google Scholar]
  23. Johnsen H. E., Madsen M. Lymphocyte subpopulations in man: no selective loss during isolation from defibrinated blood. Scand J Immunol. 1978;8(3):239–245. doi: 10.1111/j.1365-3083.1978.tb00516.x. [DOI] [PubMed] [Google Scholar]
  24. Jones K. A., Kadonaga J. T., Rosenfeld P. J., Kelly T. J., Tjian R. A cellular DNA-binding protein that activates eukaryotic transcription and DNA replication. Cell. 1987 Jan 16;48(1):79–89. doi: 10.1016/0092-8674(87)90358-8. [DOI] [PubMed] [Google Scholar]
  25. Kunkel G. R., Martinson H. G. Nucleosomes will not form on double-stranded RNa or over poly(dA).poly(dT) tracts in recombinant DNA. Nucleic Acids Res. 1981 Dec 21;9(24):6869–6888. doi: 10.1093/nar/9.24.6869. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Laimins L. A., Khoury G., Gorman C., Howard B., Gruss P. Host-specific activation of transcription by tandem repeats from simian virus 40 and Moloney murine sarcoma virus. Proc Natl Acad Sci U S A. 1982 Nov;79(21):6453–6457. doi: 10.1073/pnas.79.21.6453. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Miesfeld R., Godowski P. J., Maler B. A., Yamamoto K. R. Glucocorticoid receptor mutants that define a small region sufficient for enhancer activation. Science. 1987 Apr 24;236(4800):423–427. doi: 10.1126/science.3563519. [DOI] [PubMed] [Google Scholar]
  28. Miesfeld R., Rusconi S., Godowski P. J., Maler B. A., Okret S., Wikström A. C., Gustafsson J. A., Yamamoto K. R. Genetic complementation of a glucocorticoid receptor deficiency by expression of cloned receptor cDNA. Cell. 1986 Aug 1;46(3):389–399. doi: 10.1016/0092-8674(86)90659-8. [DOI] [PubMed] [Google Scholar]
  29. Muglia L., Rothman-Denes L. B. Cell type-specific negative regulatory element in the control region of the rat alpha-fetoprotein gene. Proc Natl Acad Sci U S A. 1986 Oct;83(20):7653–7657. doi: 10.1073/pnas.83.20.7653. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Nahon J. L., Venetianer A., Sala-Trepat J. M. Specific sets of DNase I-hypersensitive sites are associated with the potential and overt expression of the rat albumin and alpha-fetoprotein genes. Proc Natl Acad Sci U S A. 1987 Apr;84(8):2135–2139. doi: 10.1073/pnas.84.8.2135. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Pruijn G. J., van Driel W., van der Vliet P. C. Nuclear factor III, a novel sequence-specific DNA-binding protein from HeLa cells stimulating adenovirus DNA replication. Nature. 1986 Aug 14;322(6080):656–659. doi: 10.1038/322656a0. [DOI] [PubMed] [Google Scholar]
  32. Sakai M., Morinaga T., Urano Y., Watanabe K., Wegmann T. G., Tamaoki T. The human alpha-fetoprotein gene. Sequence organization and the 5' flanking region. J Biol Chem. 1985 Apr 25;260(8):5055–5060. [PubMed] [Google Scholar]
  33. Scheidereit C., Westphal H. M., Carlson C., Bosshard H., Beato M. Molecular model of the interaction between the glucocorticoid receptor and the regulatory elements of inducible genes. DNA. 1986 Oct;5(5):383–391. doi: 10.1089/dna.1986.5.383. [DOI] [PubMed] [Google Scholar]
  34. Scott R. W., Tilghman S. M. Transient expression of a mouse alpha-fetoprotein minigene: deletion analyses of promoter function. Mol Cell Biol. 1983 Jul;3(7):1295–1309. doi: 10.1128/mcb.3.7.1295. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Sell S., Morris H. P. Relationship of rat alpha 1-fetoprotein to growth rate and chromosome composition of Morris hepatomas. Cancer Res. 1974 Jun;34(6):1413–1417. [PubMed] [Google Scholar]
  36. Shuey D. J., Parker C. S. Bending of promoter DNA on binding of heat shock transcription factor. Nature. 1986 Oct 2;323(6087):459–461. doi: 10.1038/323459a0. [DOI] [PubMed] [Google Scholar]
  37. Southern P. J., Berg P. Transformation of mammalian cells to antibiotic resistance with a bacterial gene under control of the SV40 early region promoter. J Mol Appl Genet. 1982;1(4):327–341. [PubMed] [Google Scholar]
  38. Tilghman S. M. The structure and regulation of the alpha-fetoprotein and albumin genes. Oxf Surv Eukaryot Genes. 1985;2:160–206. [PubMed] [Google Scholar]
  39. Tratner I., Nahon J. L., Sala-Trepat J. M., Venetianer A. Albumin and alpha-fetoprotein gene transcription in rat hepatoma cell lines is correlated with specific DNA hypomethylation and altered chromatin structure in the 5' region. Mol Cell Biol. 1987 May;7(5):1856–1864. doi: 10.1128/mcb.7.5.1856. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Turcotte B., Guertin M., Chevrette M., Bélanger L. Rat alpha 1-fetoprotein messenger RNA: 5'-end sequence and glucocorticoid-suppressed liver transcription in an improved nuclear run-off assay. Nucleic Acids Res. 1985 Apr 11;13(7):2387–2398. doi: 10.1093/nar/13.7.2387. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Turcotte B., Guertin M., Chevrette M., LaRue H., Bélanger L. DNase I hypersensitivity and methylation of the 5'-flanking region of the alpha 1-fetoprotein gene during developmental and glucocorticoid-induced repression of its activity in rat liver. Nucleic Acids Res. 1986 Dec 22;14(24):9827–9841. doi: 10.1093/nar/14.24.9827. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Urano Y., Watanabe K., Sakai M., Tamaoki T. The human albumin gene. Characterization of the 5' and 3' flanking regions and the polymorphic gene transcripts. J Biol Chem. 1986 Mar 5;261(7):3244–3251. [PubMed] [Google Scholar]
  43. Watanabe K., Saito A., Tamaoki T. Cell-specific enhancer activity in a far upstream region of the human alpha-fetoprotein gene. J Biol Chem. 1987 Apr 5;262(10):4812–4818. [PubMed] [Google Scholar]
  44. Widen S. G., Papaconstantinou J. Liver-specific expression of the mouse alpha-fetoprotein gene is mediated by cis-acting DNA elements. Proc Natl Acad Sci U S A. 1986 Nov;83(21):8196–8200. doi: 10.1073/pnas.83.21.8196. [DOI] [PMC free article] [PubMed] [Google Scholar]
  45. Wrange O., Carlstedt-Duke J., Gustafsson J. A. Stoichiometric analysis of the specific interaction of the glucocorticoid receptor with DNA. J Biol Chem. 1986 Sep 5;261(25):11770–11778. [PubMed] [Google Scholar]
  46. Yamamoto K. R. Steroid receptor regulated transcription of specific genes and gene networks. Annu Rev Genet. 1985;19:209–252. doi: 10.1146/annurev.ge.19.120185.001233. [DOI] [PubMed] [Google Scholar]
  47. Zaret K. S., Yamamoto K. R. Reversible and persistent changes in chromatin structure accompany activation of a glucocorticoid-dependent enhancer element. Cell. 1984 Aug;38(1):29–38. doi: 10.1016/0092-8674(84)90523-3. [DOI] [PubMed] [Google Scholar]
  48. Zoller M. J., Smith M. Oligonucleotide-directed mutagenesis of DNA fragments cloned into M13 vectors. Methods Enzymol. 1983;100:468–500. doi: 10.1016/0076-6879(83)00074-9. [DOI] [PubMed] [Google Scholar]
  49. van der Eb A. J., Graham F. L. Assay of transforming activity of tumor virus DNA. Methods Enzymol. 1980;65(1):826–839. doi: 10.1016/s0076-6879(80)65077-0. [DOI] [PubMed] [Google Scholar]

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