A dosage-dependent suppressor of a temperature-sensitive calmodulin mutant encodes a protein related to the fork head family of DNA-binding proteins.
The cmd1-1 mutation of calmodulin causes temperature-sensitive growth in Saccharomyces cerevisiae. We have isolated a dosage-dependent suppressor of cmd1-1, designated HCM1. Twentyfold overexpression of HCM1 permits strains carrying cmd1-1 to grow at temperatures up to and including 34 degrees C but does not suppress the lethality of either cmd1-1 at higher temperatures or the deletion of CMD1. Thus, overexpression of HCM1 does not bypass the requirement for calmodulin but enhances the ability of the mutant calmodulin to function. HCM1 is not essential for growth, but deletion of HCM1 exacerbates the phenotype of a strain carrying cmd1-1. HCM1 is located on chromosome III, which was recently sequenced. Our results correct errors in the published DNA sequence. The putative polypeptide encoded by HCM1 is 564 amino acids long and has a predicted molecular weight of 63,622. Antisera prepared against Hcm1p detect a protein that is overproduced in yeast strains overexpressing HCM1 and has an apparent molecular mass of 65 kDa. Eighty-six amino acid residues in the N terminus of Hcm1p show 50% identity with a DNA-binding region of the fork head family of DNA-binding proteins. When fused to the DNA-binding domain of Gal4p, residues 139 to 511 of Hcm1p can act as a strong activator of transcription. However, overexpression of HCM1 does not affect the expression of calmodulin. Furthermore, Hcm1p does not bind to calmodulin in a gel overlay assay. Thus, overexpression of HCM1 enhances calmodulin function by an apparently indirect mechanism.
Zhu
G
G
Department of Biochemistry, University of Washington, Seattle 98195.
Muller
E G
EG
Amacher
S L
SL
Northrop
J L
JL
Davis
T N
TN
D000595:Amino Acid Sequence; D001483:Base Sequence; D002147:Calmodulin / Q000235:genetics* / Q000378:metabolism; D002455:Cell Division; D004252:DNA Mutational Analysis; D004268:DNA-Binding Proteins / Q000096:biosynthesis / Q000235:genetics*; D051858:Forkhead Transcription Factors; D016153:Genes, Suppressor / Q000235:genetics*; D051541:Hepatocyte Nuclear Factor 3-alpha; D051542:Hepatocyte Nuclear Factor 3-beta; D051543:Hepatocyte Nuclear Factor 3-gamma; D006358:Hot Temperature; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000235:genetics; D012441:Saccharomyces cerevisiae / Q000235:genetics* / Q000254:growth & development; D029701:Saccharomyces cerevisiae Proteins; D017386:Sequence Homology, Amino Acid; D014157:Transcription Factors / Q000235:genetics; D014158:Transcription, Genetic
D002147:Calmodulin; D004268:DNA-Binding Proteins; D051858:Forkhead Transcription Factors; C079874:HCM1 protein, S cerevisiae; D051541:Hepatocyte Nuclear Factor 3-alpha; D009687:Nuclear Proteins; D029701:Saccharomyces cerevisiae Proteins; D014157:Transcription Factors; D051543:Hepatocyte Nuclear Factor 3-gamma; D051542:Hepatocyte Nuclear Factor 3-beta