A mutational analysis identifies three functional regions of the spindle pole component Spc110p in Saccharomyces cerevisiae.
The central coiled coil of the essential spindle pole component Spc110p spans the distance between the central and inner plaques of the Saccharomyces cerevisiae spindle pole body (SPB). The carboxy terminus of Spc110p, which binds calmodulin, resides at the central plaque, and the amino terminus resides at the inner plaque from which nuclear microtubules originate. To dissect the functions of Spc110p, we created temperature-sensitive mutations in the amino and carboxy termini. Analysis of the temperature-sensitive spc110 mutations and intragenic complementation analysis of the spc110 alleles defined three functional regions of Spc110p. Region I is located at the amino terminus. Region II is located at the carboxy-terminal end of the coiled coil, and region III is the previously defined calmodulin-binding site. Overexpression of SPC98 suppresses the temperature sensitivity conferred by mutations in region I but not the phenotypes conferred by mutations in the other two regions, suggesting that the amino terminus of Spc110p is involved in an interaction with the gamma-tubulin complex composed of Spc97p, Spc98p, and Tub4p. Mutations in region II lead to loss of SPB integrity during mitosis, suggesting that this region is required for the stable attachment of Spc110p to the central plaque. Our results strongly argue that Spc110p links the gamma-tubulin complex to the central plaque of the SPB.
Sundberg
H A
HA
Department of Biochemistry, University of Washington, Seattle, Washington 98195-7350, USA.
Davis
T N
TN
D000483:Alleles; D001665:Binding Sites; D002147:Calmodulin / Q000378:metabolism; D002148:Calmodulin-Binding Proteins; D002453:Cell Cycle / Q000187:drug effects; D003598:Cytoskeletal Proteins; D005656:Fungal Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D018628:Gene Dosage; D020043:Genes, Essential / Q000235:genetics; D005800:Genes, Fungal / Q000235:genetics; D016153:Genes, Suppressor / Q000235:genetics; D005816:Genetic Complementation Test; D006918:Hydroxyurea / Q000494:pharmacology; D008854:Microscopy, Electron; D009154:Mutation / Q000235:genetics*; D009687:Nuclear Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D017434:Protein Structure, Tertiary; D012441:Saccharomyces cerevisiae / Q000166:cytology / Q000187:drug effects / Q000235:genetics* / Q000254:growth & development; D029701:Saccharomyces cerevisiae Proteins; D008941:Spindle Apparatus / Q000187:drug effects / Q000235:genetics / Q000648:ultrastructure; D013696:Temperature; D014404:Tubulin / Q000235:genetics / Q000378:metabolism
D002147:Calmodulin; D002148:Calmodulin-Binding Proteins; D003598:Cytoskeletal Proteins; D005656:Fungal Proteins; D009687:Nuclear Proteins; C073277:SPC110 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D014404:Tubulin; D006918:Hydroxyurea