p62cdc23 of Saccharomyces cerevisiae: a nuclear tetratricopeptide repeat protein with two mutable domains

Abstract

CDC23 is required in Saccharomyces cerevisiae for cell cycle progression through the G2/M transition. The CDC23 gene product contains tandem, imperfect repeats, termed tetratricopeptide repeats, (TPR) units common to a protein family that includes several other nuclear division CDC genes. In this report we have used mutagenesis to probe the functional significance of the TPR units within CDC23. Analysis of truncated derivatives indicates that the TPR block of CDC23 is necessary for the function or stability of the polypeptide. In-frame deletion of a single TPR unit within the repeat block proved sufficient to inactivate CDC23 in vivo, though this allele could rescue the temperature-sensitive defect of a cdc23 point mutant by intragenic complementation. By both in vitro and in vivo mutagenesis techniques, 17 thermolabile cdc23 alleles were produced and examined. Fourteen alleles contained single amino acid changes that were found to cluster within two distinct mutable domains, one of which encompasses the most canonical TPR unit found in CDC23. In addition, we have characterized CDC23 as a 62-kDa protein (p62cdc23) that is localized to the yeast nucleus. Our mutagenesis results suggest that TPR blocks form an essential domain within members of the TPR family.