Characterization of the human glucocorticoid receptor promoter.
To elucidate the functional elements that are involved in the regulation of the human glucocorticoid receptor (hGR) gene, transient expression, DNase I footprinting, and gel mobility shift analyses were conducted. We found that the hGR promoter region between -700 and +38 bp contained 11 footprinted sites. Deletion of the -374 to -183 bp region, which is highly conserved between human and mouse (93%), induced a 5-24-fold reduction in promoter activity in HeLa, NIH3T3, CV1, and HepG2 cells. Three footprints, FP5, FP6, and FP7, were shown to map to this region. In particular, the FP7 site was found to be within the -374 to -347 bp region. Deletion of this region triggered a significant decline in promoter activity in HeLa and NIH3T3 cells but not in HepG2 cells. AP2 was found to bind FP7. In HepG2 cells AP2 elicited transactivation of the hGR promoter activity. Transfection data revealed that the upstream GC box-rich fragment between -700 and -375 bp induced a 4-7-fold activation of the heterologous tk promoter in an orientation-independent manner. Our studies demonstrate that several transcription factors are involved in regulating GR expression and that AP2 could function as an important positive regulator of GR promoter activity.
Nobukuni
Y
Y
Clinical Neurogenetics Branch, National Institute of Mental Health, National Institutes of Health, Bethesda, Maryland 20892, USA.
Smith
C L
CL
Hager
G L
GL
Detera-Wadleigh
S D
SD
D016475:3T3 Cells; D033962:Adaptor Protein Complex 2; D033965:Adaptor Protein Complex alpha Subunits; D033942:Adaptor Proteins, Vesicular Transport; D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D004247:DNA / Q000737:chemistry / Q000378:metabolism; D003850:Deoxyribonuclease I; D017353:Gene Deletion; D005786:Gene Expression Regulation; D006367:HeLa Cells; D006801:Humans; D008565:Membrane Proteins / Q000494:pharmacology; D051379:Mice; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000378:metabolism; D015345:Oligonucleotide Probes; D011401:Promoter Regions, Genetic; D011965:Receptors, Glucocorticoid / Q000235:genetics*; D011993:Recombinant Fusion Proteins / Q000737:chemistry; D014162:Transfection
D033962:Adaptor Protein Complex 2; D033965:Adaptor Protein Complex alpha Subunits; D033942:Adaptor Proteins, Vesicular Transport; D008565:Membrane Proteins; D009687:Nuclear Proteins; D015345:Oligonucleotide Probes; D011965:Receptors, Glucocorticoid; D011993:Recombinant Fusion Proteins; D004247:DNA; D003850:Deoxyribonuclease I