Activation of metallothionein gene expression by hypoxia involves metal response elements and metal transcription factor-1

• Published in: Cancer research (Chicago, Ill.) Vol. 59; no. 6; pp. 1315 - 1322
• Main Authors: MURPHY, B. J, ANDREWS, G. K, WEBSTER, K. A, BITTEL, D, DISCHER, D. J, MCCUE, J, GREEN, C. J, YANOVSKY, M, GIACCIA, A, SUTHERLAND, R. M, LADEROUTE, K. R
• Format: Journal Article
• Language: English
• Published: Philadelphia, PA American Association for Cancer Research 15.03.1999
• Subjects: 3T3 Cells; Animals; Binding, Competitive; Biological and medical sciences; Cell Hypoxia - genetics; Cell metabolism, cell oxidation; Cell physiology; DNA-Binding Proteins; Fibroblasts - metabolism; Fundamental and applied biological sciences. Psychology; Gene Expression Regulation, Neoplastic; HT29 Cells; Humans; Metallothionein - biosynthesis; Metallothionein - genetics; Metals - metabolism; Mice; Molecular and cellular biology; Oxidation-Reduction; Oxygen - metabolism; Promoter Regions, Genetic; RNA, Messenger - biosynthesis; Transcription Factor MTF-1; Transcription Factors - genetics; Transcription Factors - metabolism; Tumor Cells, Cultured; Human; Oxidative stress; Carcinoma; Gene product; Rodentia; Malignant tumor; Gene expression; Response element; In vitro; Signal transduction; Vertebrata; Mammalia; Mouse; Animal; Established cell line; Hypoxia; Transcription factor; Mechanism of action; Tumor cell; Metallothionein
• ISSN: 0008-5472; 1538-7445

Metallothioneins (MTs) are a family of stress-induced proteins with diverse physiological functions, including protection against metal toxicity and oxidants. They may also contribute to the regulation of cellular proliferation, apoptosis, and malignant progression. We reported previously that the human (h)MT-IIA isoform is induced in carcinoma cells (A431, SiHa, and HT29) exposed to low oxygen, conditions commonly found in solid tumors. The present study demonstrates that the genes for hMT-IIA and mouse (m)MT-I are transcriptionally activated by hypoxia through metal response elements (MREs) in their proximal promoter regions. These elements bind metal transcription factor-1 (MTF-1). Deletion and mutational analyses of the hMT-IIA promoter indicated that the hMRE-a element is essential for basal promoter activity and for induction by hypoxia, but that other elements contribute to the full transcriptional response. Functional studies of the mMT-I promoter demonstrated that at least two other MREs (mMRE-d and mMRE-c) are responsive to hypoxia. Multiple copies of either hMRE-a or mMRE-d conferred hypoxia responsiveness to a minimal MT promoter. Mouse MT-I gene transcripts in fibroblasts with targeted deletions of both MTF-1 alleles (MTF-1(-/-); dko7 cells) were not induced by zinc and showed low responsiveness to hypoxia. A transiently transfected MT promoter was unresponsive to hypoxia or zinc in dko7 cells, but inductions were restored by cotransfecting a mouse MTF-1 expression vector. Electrophoretic mobility shift assays detected a specific protein-DNA complex containing MTF-1 in nuclear extracts from hypoxic cells. Together, these results demonstrate that hypoxia activates MT gene expression through MREs and that this activation involves MTF-1.