Genetic evidence for interaction between components of the yeast 26S proteasome: combination of a mutation in RPN12 (a lid component gene) with mutations in RPT1 (an ATPase gene) causes synthetic lethality.
The 19S regulatory particle of the yeast 26S proteasome consists of six related ATPases (Rpt proteins) and at least 11 non-ATPase proteins (Rpn proteins). RPN12 (formerly NIN1) encodes an Rpn component of the 19S regulatory particle and is essential for growth. To determine which subunit(s) of the 26S proteasome interact(s) with Rpn12, we attempted to screen for mutations that cause synthetic lethality in the presence of the rpn12-1 (formerly nin1-1) mutation. Among the candidates recovered was a new allele of RPT1 (formerly CIM5). This mutant allele was designated rpt1-2; on its own this mutation caused no phenotypic change, whereas the rpn12-1 rpt1-2 double mutant was lethal, suggesting a strong interaction between Rpn12 and Rpt1. The site of the rpt1-2 mutation was determined by DNA sequencing of the RPT1 locus retrieved from the mutant, and a single nucleotide alteration was found. This changes amino acid 446 of the RPT1 product from alanine to valine. The alanine residue is conserved in all Rpt proteins, except Rpt5, but no function has yet been assigned to the region that contains it. We propose that this region is necessary for Rpt1 to interact with Rpn12. The terminal phenotype of the rpn12-1 rpt1-2 double mutant was not cell cycle specific, suggesting that in the double mutant cells the function of the 26S proteasome is completely eliminated, thereby inducing multiple defects in cellular functions.
Takeuchi
J
J
Department of Biological Sciences, Graduate School of Science, University of Tokyo, Hongo, Japan.
Toh-e
A
A
D000251:Adenosine Triphosphatases / Q000235:genetics / Q000378:metabolism*; D000595:Amino Acid Sequence; D017124:Conserved Sequence; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D016368:Frameshift Mutation; D005656:Fungal Proteins / Q000235:genetics / Q000378:metabolism*; D005800:Genes, Fungal; D005804:Genes, Lethal; D015698:Genomic Library; D008969:Molecular Sequence Data; D010447:Peptide Hydrolases / Q000235:genetics / Q000378:metabolism*; D046988:Proteasome Endopeptidase Complex; D012441:Saccharomyces cerevisiae / Q000166:cytology / Q000235:genetics*; D029701:Saccharomyces cerevisiae Proteins; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D013489:Suppression, Genetic
D004268:DNA-Binding Proteins; D005656:Fungal Proteins; C074096:RPN12 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D010447:Peptide Hydrolases; D046988:Proteasome Endopeptidase Complex; C061553:ATP dependent 26S protease; D000251:Adenosine Triphosphatases; C084493:RPT1 protein, S cerevisiae