Downregulation of constitutive and heavy metal-induced metallothionein-I expression by nuclear factor I.
Although the existence of repressor protein(s) involved in the regulation of highly inducible metallothionein-I (MT-I) gene expression has been postulated, none has been identified to date. We considered nuclear factor I (NFL) protein as a potential repressor, as three half-sites for NFI binding are present on MT-I promoter and NFI is known to downregulate several cellular gene promoters. Overexpression of all four isoforms of mouse NFI protein (NFI-A, -B, -C, and -X) suppressed both constitutive and heavy metal-induced activation of the MT-I promoter in HepG2 cells. However, unlike other target genes of NFI, direct interaction of NFI with MT-I promoter is not necessary to mediate its repression. Point mutation of the NFI binding sites within the MT-I promoter that abrogates NFI binding in vitro could not alleviate the repression. Similarly, NFI proteins also repress activity of minimal MT-I promoter deficient in the NFI binding sites. Further, an NFI-C deletion mutant lacking the DNA binding domain continued to repress MT-I promoter. Overexpression of MTF-1, the key trails-acting factor involved in MT-I gene transcription, surmounted NFI-mediated repression of the basal and zinc-induced MT-I promoter activity. These data demonstrate that NFI is a repressor of MT-I expression, where its DNA binding activity is not essential to downregulate the MT-I promoter. Interaction of NFI with another protein(s), probably MTF-I, may be involved in this repression.
Majumder
S
S
Department of Molecular and Cellular Biochemistry, College of Medicine, The Ohio State University, Columbus, 43210, USA.
Ghoshal
K
K
Gronostajski
R M
RM
Jacob
S T
ST
D000818:Animals; D001483:Base Sequence; D022762:CCAAT-Enhancer-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D002104:Cadmium / Q000494:pharmacology; D002478:Cells, Cultured; D004247:DNA / Q000235:genetics / Q000378:metabolism; D004268:DNA-Binding Proteins; D015536:Down-Regulation / Q000187:drug effects; D024202:Electrophoretic Mobility Shift Assay; D008156:Luciferases / Q000235:genetics / Q000378:metabolism; D008668:Metallothionein / Q000235:genetics*; D019216:Metals, Heavy / Q000494:pharmacology*; D051379:Mice; D009154:Mutation / Q000235:genetics; D051839:NFI Transcription Factors; D009687:Nuclear Proteins; D011401:Promoter Regions, Genetic / Q000235:genetics; D051381:Rats; D012097:Repressor Proteins / Q000378:metabolism; D020218:Response Elements / Q000235:genetics; D014157:Transcription Factors / Q000378:metabolism; D014162:Transfection; D051840:Y-Box-Binding Protein 1; D000097809:Transcription Factor MTF-1
D022762:CCAAT-Enhancer-Binding Proteins; D002104:Cadmium; D004247:DNA; D004268:DNA-Binding Proteins; D008156:Luciferases; D008668:Metallothionein; D019216:Metals, Heavy; D051839:NFI Transcription Factors; D009687:Nuclear Proteins; D012097:Repressor Proteins; D014157:Transcription Factors; D051840:Y-Box-Binding Protein 1; D000097809:Transcription Factor MTF-1; C497646:Nfia protein, mouse; C497648:Nfic protein, mouse; C497649:Nfic protein, rat; C070856:YBX1 protein, human