title: Transcriptional control of adrenomedullin induction by phorbol ester in human monocytic leukemia cells.
abstract: Adrenomedullin is a potent vasodilator peptide that was originally identified from human pheochromocytoma. In this study, we investigated the induction of adrenomedullin gene expression in THP-1 acute monocytic leukemia cells during differentiation into macrophage-like cells by 12-O-tetradecanoylphorbol-13-acetate (TPA), and identified a cis-regulatory region of the human adrenomedullin gene responsible for TPA-induced adrenomedullin expression. Upon treatment with TPA (100 ng x mL(-1)) for 24 h, immunoreactive adrenomedullin concentrations in the culture medium and adrenomedullin mRNA levels were increased more than 10-fold, concomitant with the differentiation of THP-1 cells into macrophage-like cells. Actinomycin D abolished the TPA-induced adrenomedullin expression, indicating that the induction of ADM gene expression by TPA was regulated at the transcriptional level. Transient transfection assay revealed that a cis-acting region (positions -70 to -30) of human adrenomedullin gene was necessary for TPA-induced reporter gene expression. This region contains multiple copies of activator protein 2 (AP-2) binding sites, which are bound by purified AP-2 protein, as judged by electrophoretic mobility shift assay. The binding activity to this region was undetectable in nuclear extracts prepared from untreated THP-1 cells, but was increased in extracts prepared from TPA-treated cells. The protein binding was abolished by unlabeled oligonucleotides containing the AP-2 consensus sequence. These results indicate that the region (-70 to -30) of the human ADM gene containing multiple AP-2 binding sites is responsible for TPA-induced adrenomedullin expression in THP-1 cells.
authors.affiliation: Department of Molecular Biology and Applied Physiology, Tohoku University School of Medicine, Aoba-ku, Sendai, Miyagi, Japan.
authors.forename: M
authors.initials: M
authors.lastname: Nakayama
authors.forename: K
authors.initials: K
authors.lastname: Takahashi
authors.forename: T
authors.initials: T
authors.lastname: Kitamuro
authors.forename: O
authors.initials: O
authors.lastname: Murakami
authors.forename: K
authors.initials: K
authors.lastname: Shirato
authors.forename: S
authors.initials: S
authors.lastname: Shibahara
mesh_terms: D053607:Adrenomedullin; D001665:Binding Sites; D002454:Cell Differentiation / Q000187:drug effects; D004268:DNA-Binding Proteins / Q000378:metabolism; D003609:Dactinomycin / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004586:Electrophoresis; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D006801:Humans; D007948:Leukemia, Monocytic, Acute / Q000473:pathology; D010455:Peptides / Q000187:drug effects* / Q000235:genetics* / Q000378:metabolism; D012045:Regulatory Sequences, Nucleic Acid; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology*; D050656:Transcription Factor AP-2; D014157:Transcription Factors / Q000378:metabolism; D014158:Transcription, Genetic; D014162:Transfection; D014407:Tumor Cells, Cultured
chemical_list: D004268:DNA-Binding Proteins; D010455:Peptides; D050656:Transcription Factor AP-2; D014157:Transcription Factors; D053607:Adrenomedullin; D003609:Dactinomycin; D013755:Tetradecanoylphorbol Acetate