A putative AP-2 binding site in the 5' flanking region of the mouse POMC gene.
Using extracts of AtT-20 cell nuclei, protein binding sites on the POMC gene 5'-flanking region were examined with an exonuclease protection approach. One such binding site, located from -119 to -106 bp upstream from the mouse POMC gene transcription initiation site, which exhibited a close homology to the activator protein-2 (AP-2) site [1]. A double-stranded oligonucleotide containing this site was subsequently used in gel shift assays to demonstrate AP-2 consensus sequence binding activity in extracts of AtT-20 cell nuclei. Gel shift competition experiments using both homologous and heterologous competitor DNA sequences revealed that the AP-2 like factor(s) exhibited specific binding to the mouse AP-2 consensus sequence. Furthermore, AP-2 factor binding was also modulated by a CTF/NF1-like factor. Pretreatment of AtT-20 cell nuclear extracts with alkaline phosphatase prior to inclusion in gel shift assays led to a reduction in the intensities of AP-2 factor-specific bands, indicating a potential involvement of protein phosphorylation in AP-2 factor binding in AtT-20 cells.
Bishop
J F
JF
Experimental Therapeutics Branch, NINDS, NIH, Bethesda, MD 20892.
Rinaudo
M S
MS
Ritter
J K
JK
Chang
A C
AC
Conant
K
K
Gehlert
D R
DR
D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism*; D005796:Genes; D005809:Genes, Regulator; D051379:Mice; D008969:Molecular Sequence Data; D015345:Oligonucleotide Probes; D010911:Pituitary Neoplasms; D011333:Pro-Opiomelanocortin / Q000235:genetics*; D050656:Transcription Factor AP-2; D014157:Transcription Factors / Q000378:metabolism*; D014158:Transcription, Genetic
D004268:DNA-Binding Proteins; D015345:Oligonucleotide Probes; D050656:Transcription Factor AP-2; D014157:Transcription Factors; D011333:Pro-Opiomelanocortin