AP-2beta represses D(1A) dopamine receptor gene transcription in neuro2a cells.
Expression of the D(1A) dopamine receptor in brain is restricted to specific neuronal populations. To investigate the mechanism of this selective expression, we localized a silencer upstream of the human D(1A) gene and identified its binding transcription factor in the D(1A)-negative neural cell line Neuro2a. Using deletion CAT analysis, we narrowed this silencer to the region between nucleotides -561 and -532 relative to the CAP site. This 30-bp region, designated D1AS1, contains a sequence homologous to the AP-2 binding site and binds to a factor that also interacts with the AP-2 consensus sequence. In gel supershift assays, this factor is recognized by anti-AP-2beta antibody. Co-transfection of Neuro2a cells with an AP-2beta expression vector repressed the basal CAT activity of D(1A) promoter-reporter plasmids in a D1AS1-dependent manner. RT-PCR analysis indicated that, among AP-2 family members, Neuro2a cells express only AP-2beta. Furthermore, co-transfection of these cells with decoy oligonucleotides corresponding to the D1AS1 sequence de-repressed the D(1A) gene promoter. Unlike in Neuro2a cells, AP-2beta could not repress the D(1A) promoter in the D(1A)-positive neural cell line, NS20Y. In addition, the expression of AP-2beta in different brain regions does not inversely correlate with that of D(1A) dopamine receptor. These observations taken together indicate that AP-2beta is a repressive transcription factor that acts on the D1AS1 silencer of the D(1A) dopamine receptor gene via some cell-specific mechanism(s) in Neuro2a.
Takeuchi
S
S
Department of Neurology, Graduate School of Medicine, University of Tokyo, 7-3-1, Hongo, Bunkyo-ku, Tokyo 113, Japan.
Imafuku
I
I
Waragai
M
M
Roth
C
C
Kanazawa
I
I
Buettner
R
R
Mouradian
M M
MM
Okazawa
H
H
D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D001921:Brain / Q000378:metabolism; D015500:Chloramphenicol O-Acetyltransferase / Q000235:genetics / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D005786:Gene Expression Regulation; D006801:Humans; D051379:Mice; D009687:Nuclear Proteins / Q000378:metabolism; D009841:Oligonucleotides / Q000235:genetics; D011401:Promoter Regions, Genetic / Q000235:genetics; D011485:Protein Binding; D017447:Receptors, Dopamine D1 / Q000235:genetics* / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D012045:Regulatory Sequences, Nucleic Acid / Q000235:genetics; D050656:Transcription Factor AP-2; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D014158:Transcription, Genetic; D014162:Transfection; D014407:Tumor Cells, Cultured; D015854:Up-Regulation
D004268:DNA-Binding Proteins; D009687:Nuclear Proteins; D009841:Oligonucleotides; D017447:Receptors, Dopamine D1; D011993:Recombinant Fusion Proteins; C493677:TFAP2B protein, human; C493669:Tfap2b protein, mouse; D050656:Transcription Factor AP-2; D014157:Transcription Factors; C098593:dopamine D1A receptor; D015500:Chloramphenicol O-Acetyltransferase