Isolation and characterization of the promoter of the human GABAA receptor alpha 1 subunit gene.
The GABAA receptor, as assessed by ligand binding and chloride flux measurement in vivo and in vitro, is down-regulated in response to chronic benzodiazepine exposure. The mRNA levels of the alpha 1 and gamma 2 subunits of the receptor are also reduced. We have isolated the promoter of the gene encoding the alpha 1 subunit of the GABAA receptor to elucidate the regulatory mechanism of its expression. A DNA segment 650 bp long has been isolated that includes 151 bp of untranslated 5' end of the cDNA sequence and 500 bp of potential promoter-enhancer region. The transcriptional activity of this DNA segment linked to the firefly luciferase gene showed a strong orientation specificity. The promoter activity was localized to a 60-bp segment by deletion mapping. Mobility shift binding assay results suggest that this segment may interact with one or more factors in HeLa cell nuclear extracts to form a transcriptional complex. Primary cultures of embryonic chick cortical cells transfected with the promoter-luciferase construct were treated chronically with lorazepam. Transcriptional activity of this promoter construct was strongly repressed by chronic administration of lorazepam.
Kang
I
I
Department of Pharmacology and Experimental Therapeutics, Tufts University School of Medicine, Boston, Massachusetts 02111.
Lindquist
D G
DG
Kinane
T B
TB
Ercolani
L
L
Pritchard
G A
GA
Miller
L G
LG
D000818:Animals; D001483:Base Sequence; D002540:Cerebral Cortex / Q000378:metabolism; D002642:Chick Embryo; D002998:Clonazepam / Q000494:pharmacology; D004247:DNA / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification*; D018076:DNA, Complementary / Q000737:chemistry; D004742:Enhancer Elements, Genetic; D006367:HeLa Cells; D006801:Humans; D008156:Luciferases / Q000235:genetics; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic; D018079:Receptors, GABA / Q000235:genetics*; D012045:Regulatory Sequences, Nucleic Acid; D014158:Transcription, Genetic; D014162:Transfection
D018076:DNA, Complementary; D018079:Receptors, GABA; D002998:Clonazepam; D004247:DNA; D008156:Luciferases