Functional domains of the heavy metal-responsive transcription regulator MTF-1.
Metallothioneins (MTs) constitute a class of low molecular weight, cysteine-rich, metal binding proteins which are regulated at the level of gene transcription in response to heavy metals and other adverse treatments. We have previously cloned a zinc finger factor (MTF-1) that binds specifically to heavy metal-responsive DNA sequence elements in metallothionein promoters and shown that this factor is essential for basal and heavy metal-induced transcription. Here we report that the C-terminal part of MTF-1 downstream of the DNA binding zinc fingers harbours three different transactivation domains, namely an acidic domain, a proline-rich domain and a domain rich in serine and threonine. When fused to the heterologous DNA binding domain of the yeast factor GAL4 these activation domains function constitutively, i.e. transcription of a GAL4-driven reporter gene is not induced by heavy metals. In search of the region(s) responsible for metal induction, external and internal deletion mutations of mouse and human MTF-1 and chimeric variants thereof were tested with a reporter gene driven by a metal-responsive promoter. The N-terminal part of MTF-1 containing the zinc fingers, which are dependent on zinc for efficient DNA binding, can indeed confer a limited (3- to 4-fold) zinc-responsive transcription when fused to the heterologous activation domain of the viral VP16 protein. Another region containing the acidic and proline-rich activation domains also contributes to metal inducibility, but only in the context of intact MTF-1. This indicates that the activity of MTF-1 results from a complex interplay of different functional domains.
Radtke
F
F
Institut für Molekularbiologie II der Universität Zürich, Switzerland.
Georgiev
O
O
Müller
H P
HP
Brugnera
E
E
Schaffner
W
W
D001483:Base Sequence; D001665:Binding Sites; D004247:DNA / Q000378:metabolism*; D004268:DNA-Binding Proteins; D005656:Fungal Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D017353:Gene Deletion; D006367:HeLa Cells; D017405:Herpes Simplex Virus Protein Vmw65 / Q000737:chemistry / Q000235:genetics; D006801:Humans; D006863:Hydrogen-Ion Concentration; D008668:Metallothionein / Q000235:genetics*; D008670:Metals / Q000494:pharmacology*; D008969:Molecular Sequence Data; D016296:Mutagenesis; D010957:Plasmids; D011392:Proline / Q000032:analysis; D011401:Promoter Regions, Genetic; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins; D012694:Serine / Q000032:analysis; D013912:Threonine / Q000032:analysis; D014157:Transcription Factors / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D015533:Transcriptional Activation; D016335:Zinc Fingers; D000097809:Transcription Factor MTF-1
D004247:DNA; D004268:DNA-Binding Proteins; D005656:Fungal Proteins; D017405:Herpes Simplex Virus Protein Vmw65; D008668:Metallothionein; D008670:Metals; D011392:Proline; D011993:Recombinant Fusion Proteins; D029701:Saccharomyces cerevisiae Proteins; D012694:Serine; D013912:Threonine; D014157:Transcription Factors; D000097809:Transcription Factor MTF-1; C052130:GAL4 protein, S cerevisiae