title: Zinc-regulated DNA binding of the yeast Zap1 zinc-responsive activator.
abstract: The Zap1 transcription factor of Saccharomyces cerevisiae plays a central role in zinc homeostasis by controlling the expression of genes involved in zinc metabolism. Zap1 is active in zinc-limited cells and repressed in replete cells. At the transcriptional level, Zap1 controls its own expression via positive autoregulation. In addition, Zap1's two activation domains are regulated independently of each other by zinc binding directly to those regions and repressing activation function. In this report, we show that Zap1 DNA binding is also inhibited by zinc. DMS footprinting showed that Zap1 target gene promoter occupancy is regulated with or without transcriptional autoregulation. These results were confirmed using chromatin immunoprecipitation. Zinc regulation of DNA binding activity mapped to the DNA binding domain indicating other parts of Zap1 are unnecessary for this control. Overexpression of Zap1 overrode DNA binding regulation and resulted in constitutive promoter occupancy. Under these conditions of constitutive binding, both the zinc dose response of Zap1 activity and cellular zinc accumulation were altered suggesting the importance of DNA binding control to zinc homeostasis. Thus, our results indicated that zinc regulates Zap1 activity post-translationally via three independent mechanisms, all of which contribute to the overall zinc responsiveness of Zap1.
authors.affiliation: Department of Nutritional Sciences, University of Wisconsin-Madison, Madison, Wisconsin, United States of America.
authors.forename: Avery G
authors.initials: AG
authors.lastname: Frey
authors.forename: Amanda J
authors.initials: AJ
authors.lastname: Bird
authors.forename: Marguerite V
authors.initials: MV
authors.lastname: Evans-Galea
authors.forename: Elizabeth
authors.initials: E
authors.lastname: Blankman
authors.forename: Dennis R
authors.initials: DR
authors.lastname: Winge
authors.forename: David J
authors.initials: DJ
authors.lastname: Eide
mesh_terms: D021581:Active Transport, Cell Nucleus; D001483:Base Sequence; D002467:Cell Nucleus / Q000378:metabolism; D004247:DNA / Q000235:genetics / Q000378:metabolism*; D006706:Homeostasis; D008969:Molecular Sequence Data; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D020218:Response Elements / Q000235:genetics; D012441:Saccharomyces cerevisiae / Q000166:cytology / Q000235:genetics / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D014157:Transcription Factors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D014158:Transcription, Genetic; D015032:Zinc / Q000378:metabolism*
chemical_list: D029701:Saccharomyces cerevisiae Proteins; D014157:Transcription Factors; C103264:ZAP1 protein, S cerevisiae; D004247:DNA; D015032:Zinc