Transcriptional activation mediated by the yeast AP-1 protein is required for normal cadmium tolerance.
The yeast YAP1 gene encodes a transcriptional regulatory protein that utilizes a basic region-leucine zipper (bZip) DNA-binding domain to recognize its cognate DNA element. A synthetic reporter gene containing a SV40 AP-1 response element (ARE) cloned upstream of a TRP5 promoter-lacZ gene fusion shows yAP-1-dependent transactivation in vivo. Recent work has shown that changes in the gene dosage of this factor can dramatically alter the ability of a cell to tolerate a host of toxic agents including cadmium, cycloheximide, and sulfometuron methyl. We have focused on the YAP1-dependent cadmium resistance as cells that lack a functional YAP1 gene are hypersensitive to this metal. Deletion mapping experiments define two domains in the carboxyl-terminal region of the yAP-1 protein that are required for normal cadmium tolerance and ARE-TRP5-lacZ expression. Single amino acid substitutions in the bZip domain of yAP-1 indicate that this region is required for normal DNA binding and in vivo function of the protein. Replacement of a non-canonical asparagine with leucine in the yAP-1 leucine zipper leads to production of a defective protein. A substitution mutation in the basic domain converts this mutant protein into a dominant negative factor. The ability of yAP-1 to act as a positive regulator of transcription is required for its biological action.
Wemmie
J A
JA
Department of Physiology and Biophysics, University of Iowa, Iowa City 52242.
Wu
A L
AL
Harshman
K D
KD
Parker
C S
CS
Moye-Rowley
W S
WS
D001483:Base Sequence; D015153:Blotting, Western; D002104:Cadmium / Q000633:toxicity*; D003513:Cycloheximide / Q000633:toxicity; D017931:DNA Primers; D004268:DNA-Binding Proteins / Q000096:biosynthesis / Q000302:isolation & purification / Q000378:metabolism*; D004352:Drug Resistance, Microbial / Q000235:genetics; D005656:Fungal Proteins / Q000378:metabolism; D015966:Gene Expression Regulation, Fungal / Q000187:drug effects; D005800:Genes, Fungal; D008969:Molecular Sequence Data; D016296:Mutagenesis; D010641:Phenotype; D010957:Plasmids; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000302:isolation & purification / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000187:drug effects / Q000235:genetics* / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins; D017384:Sequence Deletion; D013453:Sulfonylurea Compounds / Q000633:toxicity; D014157:Transcription Factors / Q000096:biosynthesis / Q000302:isolation & purification / Q000378:metabolism*; D014158:Transcription, Genetic; D015533:Transcriptional Activation; D001616:beta-Galactosidase / Q000096:biosynthesis
D017931:DNA Primers; D004268:DNA-Binding Proteins; D005656:Fungal Proteins; D011994:Recombinant Proteins; D029701:Saccharomyces cerevisiae Proteins; D013453:Sulfonylurea Compounds; D014157:Transcription Factors; C086677:YAP1 protein, S cerevisiae; D002104:Cadmium; D003513:Cycloheximide; D001616:beta-Galactosidase; C042031:sulfometuron methyl