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A putative AP‐2 binding site in the 5' flanking region of the mouse POMC gene
Author(s) -
Bishop John F.,
Rinaudo Mario S.,
Ritter Joseph K.,
Chang Annie C.-Y.,
Conant Katherine,
Gehlert Donald R.
Publication year - 1990
Publication title -
febs letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.593
H-Index - 257
eISSN - 1873-3468
pISSN - 0014-5793
DOI - 10.1016/0014-5793(90)80781-d
Subject(s) - microbiology and biotechnology , binding site , gata6 , dna binding site , biology , 5' flanking region , transcription factor , activator (genetics) , gene , electrophoretic mobility shift assay , consensus sequence , exonuclease , peptide sequence , biochemistry , promoter , gene expression , dna polymerase
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 aetivator 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/NFl‐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.