Genetic interactions between CDC31 and KAR1, two genes required for duplication of the microtubule organizing center in Saccharomyces cerevisiae.
KAR1 encodes an essential component of the yeast spindle pole body (SPB) that is required for karyogamy and SPB duplication. A temperature-sensitive mutation, kar1-delta 17, mapped to a region required for SPB duplication and for localization to the SPB. To identify interacting SPB proteins, we isolated 13 dominant mutations and 3 high copy number plasmids that suppressed the temperature sensitivity of kar1-delta 17. Eleven extragenic suppressor mutations mapped to two linkage groups, DSK1 and DSK2. The extragenic suppressors were specific for SPB duplication and did not suppress karyogamy-defective alleles. The major class, DSK1, consisted of mutations in CDC31. CDC31 is required for SPB duplication and encodes a calmodulin-like protein that is most closely related to caltractin/centrin, a protein associated with the Chlamydomonas basal body. The high copy number suppressor plasmids contained the wild-type CDC31 gene. One CDC31 suppressor allele conferred a temperature-sensitive defect in SPB duplication, which was counter-suppressed by recessive mutations in KAR1. In spite of the evidence for a direct interaction, the strongest CDC31 alleles, as well as both DSK2 alleles, suppressed a complete deletion of KAR1. However, the CDC31 alleles also made the cell supersensitive to KAR1 gene dosage, arguing against a simple bypass mechanism of suppression. We propose a model in which Kar1p helps localize Cdc31p to the SPB and that Cdc31p then initiates SPB duplication via interaction with a downstream effector.
Vallen
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Department of Molecular Biology, Princeton University, New Jersey 08544-1014.
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D000483:Alleles; D000595:Amino Acid Sequence; D000818:Animals; D002135:Calcium-Binding Proteins / Q000235:genetics*; D018797:Cell Cycle Proteins; D002696:Chlamydomonas / Q000235:genetics; D002868:Chromosomal Proteins, Non-Histone; D004271:DNA, Fungal / Q000235:genetics / Q000378:metabolism; D005455:Fluorescent Antibody Technique; D005656:Fungal Proteins / Q000235:genetics*; D017353:Gene Deletion; D005800:Genes, Fungal; D005838:Genotype; D008854:Microscopy, Electron; D008870:Microtubules / Q000502:physiology / Q000648:ultrastructure*; D008969:Molecular Sequence Data; D009154:Mutation; D009687:Nuclear Proteins / Q000235:genetics*; D010957:Plasmids; D015183:Restriction Mapping; D012441:Saccharomyces cerevisiae / Q000235:genetics* / Q000254:growth & development / Q000648:ultrastructure; D029701:Saccharomyces cerevisiae Proteins; D017386:Sequence Homology, Amino Acid; D013489:Suppression, Genetic; D013696:Temperature
C083769:CDC31 protein, S cerevisiae; D002135:Calcium-Binding Proteins; D018797:Cell Cycle Proteins; D002868:Chromosomal Proteins, Non-Histone; D004271:DNA, Fungal; D005656:Fungal Proteins; C074559:KAR1 protein, S cerevisiae; D009687:Nuclear Proteins; D029701:Saccharomyces cerevisiae Proteins; C056608:caltractin