Nuclear factor-1 and metal transcription factor-1 synergistically activate the mouse metallothionein-1 gene in response to metal ions.
Metal activation of metallothionein (MT) gene transcription is dependent on the presence of metal regulatory elements (MREs), which are present in five non-identical copies (MREa through MREe) in the promoter of the mouse MT-1 gene and on the capacity of metal transcription factor-1 (MTF-1) to bind to the MREs in the presence of zinc. We detected a protein, distinct from MTF-1, specifically binding to the MREc region. DNA binding competition experiments using synthetic oligonucleotides and specific anti-NF1 antibodies showed that this protein binds to an NF1 site overlapping the MREc element as well as to a second site upstream of the Sp1a site and corresponds to NF1 or a related protein. Transfection experiments showed that loss of the two NF1 sites decreased metal-induced MT promoter activity by 55-70% in transiently transfected cells and almost completely abrogated metal and tert-butylhydroquinone (tBHQ) induction in stably transfected cells. Similarly, expression of an inactive NF1 protein strongly inhibited MT-1 promoter activity. Using synthetic promoters containing NF1 and MRE sites fused to a minimal MT promoter, we showed that these NF1 sites did not confer metal induction but enhanced metal-induced promoter activity. Chromatin immunoprecipitation assays confirmed that NF1 binds to the mouse MT-1 promoter in vivo and showed that NF1 binding is zinc-inducible. In addition, zinc-induced NF1 DNA binding was MTF-1-dependent. Taken together, these studies show that NF1 acts synergistically with MTF-1 to activate the mouse MT-1 promoter in response to metal ions and tert-butylhydroquinone and contributes to maximal activation of the gene.
LaRochelle
Olivier
O
Centre de Recherche en Cancérologie de l'Université Laval, CHUQ, Hôtel-Dieu de Québec, Québec, Canada.
Labbé
Simon
S
Harrisson
Jean-François
JF
Simard
Carl
C
Tremblay
Véronique
V
St-Gelais
Geneviève
G
Govindan
Manjapra V
MV
Séguin
Carl
C
D000818:Animals; D002412:Cations / Q000737:chemistry; D002460:Cell Line; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D005786:Gene Expression Regulation; D006801:Humans; D008668:Metallothionein / Q000235:genetics*; D051379:Mice; D008969:Molecular Sequence Data; D051839:NFI Transcription Factors / Q000235:genetics / Q000378:metabolism*; D011401:Promoter Regions, Genetic / Q000235:genetics; D011485:Protein Binding; D020218:Response Elements / Q000235:genetics; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism*; D015032:Zinc / Q000737:chemistry / Q000378:metabolism*; D000097809:Transcription Factor MTF-1
D002412:Cations; D004268:DNA-Binding Proteins; D008668:Metallothionein; D051839:NFI Transcription Factors; D014157:Transcription Factors; D015032:Zinc; D000097809:Transcription Factor MTF-1