Identification of COUP-TF as a transcriptional repressor of the c-mos proto-oncogene.
The c-mos proto-oncogene is specifically expressed in the male and female germ cells of the mouse and other vertebrates. We previously identified a 15-base pair sequence element (B2) as the binding site of a candidate repressor of c-mos transcription in somatic cells. In the present study, we used the yeast one-hybrid system to isolate HeLa cell cDNAs encoding proteins that specifically bound to the c-mos B2 element. Nucleotide sequencing identified several of the clones isolated in this screen as the orphan nuclear receptors COUP-TFI and COUP-TFII. A COUP-TF-binding site was then identified within the B2 sequence. Complexes formed between purified COUP-TFs and the c-mos B2 probe comigrated in electrophoretic mobility shift assays with those formed using whole nuclear extracts of NIH 3T3 or HeLa cells. Moreover, the complexes formed with NIH 3T3 nuclear extracts and B2 probe were supershifted with antibody against COUP-TF, identifying COUP-TF as the candidate repressor previously detected in these somatic cell extracts. Substitution of a consensus COUP-TF-binding site for the c-mos negative regulatory element suppressed expression from the c-mos promoter in transfected somatic cells, demonstrating the functional activity of COUP-TF as a repressor of c-mos transcription.
Lin
H B
HB
Department of Biology, Boston University, Boston, Massachusetts 02215, USA.
Jurk
M
M
Gulick
T
T
Cooper
G M
GM
D016475:3T3 Cells; D000818:Animals; D001483:Base Sequence; D051838:COUP Transcription Factor I; D051837:COUP Transcription Factor II; D051836:COUP Transcription Factors; D004268:DNA-Binding Proteins / Q000235:genetics*; D005260:Female; D015972:Gene Expression Regulation, Neoplastic; D016258:Genes, mos / Q000235:genetics*; D006367:HeLa Cells; D006801:Humans; D008297:Male; D051379:Mice; D008969:Molecular Sequence Data; D000090063:Proto-Oncogene Mas; D011987:Receptors, Steroid; D012097:Repressor Proteins / Q000235:genetics; D014157:Transcription Factors / Q000235:genetics*; D014158:Transcription, Genetic
D051838:COUP Transcription Factor I; D051837:COUP Transcription Factor II; D051836:COUP Transcription Factors; D004268:DNA-Binding Proteins; D000090063:Proto-Oncogene Mas; D011987:Receptors, Steroid; D012097:Repressor Proteins; D014157:Transcription Factors; C000715470:MAS1 protein, human; C497589:NR2F1 protein, human; C497590:Nr2f1 protein, mouse; C497587:Nr2f2 protein, mouse; C095569:NR2F2 protein, human