A negative regulatory element in the promoter region of the rat alpha 2A-adrenergic receptor gene overlaps an SP1 consensus binding site.
Three subtypes of alpha 2-adrenergic receptors (alpha 2A, alpha 2B and alpha 2C) have been described that differ in their primary sequence and tissue-specific expression and are encoded by three distinct genes. Previous work has shown that the human alpha 2A-adrenergic receptor gene promoter consists of a TATA-box (TATAAA), palindromic sequence (CCCACGTGGG) and GC-box (GGGGCGG) motif. Sequence analysis of the putative promoter region of the rat alpha 2A-adrenergic receptor gene showed that these promoter regions are conserved in their sequence and relative location. We analysed the transcriptional activity of these regions using RINm5F, a rat insulinoma cell line that expresses the endogenous alpha 2A-adrenergic receptor gene. These results showed that the region from -484 to -92 has a negative effect on transcription, as deletion of this region in alpha 2A-adrenergic receptor gene-chloramphenicol acetyltransferase reporter constructs increased reporter gene activity. This region included the GC-box sequence which is a consensus binding site for the nuclear factor SP1, which is a positive activator of transcription. Gel-mobility-shift assays and supershift assays with an antibody that recognizes SP1 showed binding of the SP1 nuclear factor as well as other nuclear factors to this GC-box region. Additional nuclear factors bind to the downstream palindromic region. We suggest that positive- and negative-acting nuclear factors contribute to the activity of the alpha 2-adrenergic receptor promoter.
D000818:Animals; D001483:Base Sequence; D001665:Binding Sites / Q000235:genetics; D022762:CCAAT-Enhancer-Binding Proteins; D003110:Colonic Neoplasms / Q000378:metabolism; D016384:Consensus Sequence; D017124:Conserved Sequence; D004268:DNA-Binding Proteins / Q000378:metabolism*; D005809:Genes, Regulator; D006801:Humans; D007340:Insulinoma / Q000378:metabolism; D008969:Molecular Sequence Data; D051839:NFI Transcription Factors; D009687:Nuclear Proteins; D010190:Pancreatic Neoplasms / Q000378:metabolism; D011401:Promoter Regions, Genetic / Q000235:genetics*; D051381:Rats; D018341:Receptors, Adrenergic, alpha-2 / Q000235:genetics* / Q000378:metabolism; D016385:TATA Box; D014157:Transcription Factors / Q000378:metabolism; D014407:Tumor Cells, Cultured; D051840:Y-Box-Binding Protein 1
D022762:CCAAT-Enhancer-Binding Proteins; D004268:DNA-Binding Proteins; D051839:NFI Transcription Factors; D009687:Nuclear Proteins; D018341:Receptors, Adrenergic, alpha-2; D014157:Transcription Factors; D051840:Y-Box-Binding Protein 1; C070856:YBX1 protein, human