Nucleic Acids Research, 1990, Vol. 18, No. 17 5097-5106
© 1990
MOLECULAR BIOLOGY |
Identification of an estrogen response element upstream of the human c-fos gene that binds the estrogen receptor and the AP-1 transcription factor
Laboratoire de Génétique Moléculaire des Eucaryotes du CNRS, Unité 184 de Biologie Moléculaire et de Génie Génétique de I'INSERM, Institut de Chimie Biologique, Faculté de Médecine Strasbourg, France 1Instituto di Patologia Generale e Oncologia, Prima Facoltà di Medicina e Chirurgia, Università di Napoli Naples l-80138, Italy
*To whom correspondence should be addressed
Received July 8, 1990. Revised July 20, 1990. Accepted July 20, 1990.
Transcription of the proto-oncogene c-fos is stimulated by 17ß-estradiol in estrogen responsive human and rat cells. To understand the molecular mechanisms of estrogen regulation of c-fos gene transcription, the human c-fos gene promoter, with 2.25 Kb of 5'-flanklng DNA, was cloned upstream of the bacterial CAT gene and tested for estrogen regulation by transient transfection in HeLa cells. When an expression vector coding for the human estrogen receptor was cotransfected with the fos -CAT reporter, the promoter was found to respond to 17ß-estradiol. An element responsible for estrogen induction was mapped in a 240 bp region localized 1060 to 1300 bases upstream of the startsite of transcription of the gene. Sequence analysis revealed, clustered in a 19 bp sub-region, a sequence corresponding to an imperfectly palindromic ERE: CGGCAGCGTGACC and two sequences: CTGAG and GTGAC, homologous to the core sequence of AP-1 transcription factor binding sites. A synthetic oligonucleotide reproducing this sub-region binds ‘in vitro’ both the estrogen receptor and AP-1 factor(s) and confers estrogen-responsivity to the HSV-tk gene promoter. Transcriptional activation by the estrogen receptor is prevented by mutations in the fos ERE that hamper binding of the receptor in vitro. Activation of the c-fos gene promoter in HeLa cells requires the DNA binding domain of the estrogen receptor, and can be achieved independently by the TAF-1 and the TAF-2 transcriptional activation functions of this molecule. A receptor mutant lacking the hormone binding domain can activate the c-fos gene promoter in the absence of estrogen.
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