Metal-dependent binding of a nuclear factor to the rat metallothionein-I promoter

• Published in: Nucleic acids research Vol. 18; no. 20; pp. 6049 - 6055
• Main Authors: ANDERSEN, R. D, TAPLITZ, S. J, OBERBAUER, A. M, CALAME, K. L, HERSCHMAN, H. R
• Format: Journal Article
• Language: English
• Published: Oxford Oxford University Press 25.10.1990
• Subjects: Animals; Base Sequence; Binding Sites; Biological and medical sciences; Cadmium - pharmacology; Cell Line; Cell Nucleus - metabolism; Fundamental and applied biological sciences. Psychology; Liver Neoplasms, Experimental; Metallothionein - genetics; Molecular and cellular biology; Molecular genetics; Molecular Sequence Data; Nuclear Proteins - metabolism; Oligonucleotide Probes; Promoter Regions, Genetic - drug effects; Rats; Restriction Mapping; Transcription Factors - metabolism; Transcription, Genetic; Transcription. Transcription factor. Splicing. Rna processing; Ultraviolet Rays; Cadmium; Rat; Transcription promoter; Rodentia; DNA binding protein; Molecular interaction; Crosslinking; Heavy metal; Vertebrata; Regulation(control); Mammalia; Regulatory sequence; Transcription factor; Metallothionein
• ISSN: 0305-1048; 1362-4962
• DOI: 10.1093/nar/18.20.6049

Genomic footprinting studies in vivo and experiments using synthetic metal regulatory elements (MREs) in vitro suggest protein binding to the MREs of the mouse and rat metallothionein I (MT-I) genes. Using gel-retardation assays of promoter fragments, we observe a cadmium-dependent binding factor for the rat MT-I promoter in rat hepatoma cells. This factor is present in extracts from both uninduced and cadmium-induced cells, but requires the presence of cadmium to bind to the promoter. The formation of a cadmium-dependent complex is competed by an oligonucleotide containing two MREs. This competition is lost when when one of the MREs is mutated, indicating a requirement for at least two MREs for binding of this factor. The cadmium-dependent factor dissociates more rapidly from the MT-I promoter than does a factor that binds to a consensus Sp1 site present on the same DNA fragment. UV crosslinking analysis using nuclear extracts from cadmium induced cells, in the presence of an oligonucleotide probe containing both 5-bromodeoxyuridine and 32P-deoxycytidine, identifies a 39 kDalton protein associated with the metal inducible complex.