The yeast protein kinase Mps1p is required for assembly of the integral spindle pole body component Spc42p

Andrea R. Castillo, Janet B. Meehl, Garry Morgan, Amy Schutz-Geschwender, Mark Winey

Research output: Contribution to journalArticle

51 Citations (Scopus)

Abstract

Saccharomyces cerevisiae MPS1 encodes an essential protein kinase that has roles in spindle pole body (SPB) duplication and the spindle checkpoint. Previously characterized MPS1 mutants fail in both functions, leading to aberrant DNA segregation with lethal consequences. Here, we report the identification of a unique conditional allele, mps1-8, that is defective in SPB duplication but not the spindle checkpoint. The mutations in mps1-8 are in the noncatalytic region of MPS1, and analysis of the mutant protein indicates that Mps1-8p has wild-type kinase activity in vitro. A screen for dosage suppressors of the mps1-8 conditional growth phenotype identified the gene encoding the integral SPB component SPC42. Additional analysis revealed that mps1-8 exhibits synthetic growth defects when combined with certain mutant alleles of SPC42. An epitope-tagged version of Mps1p (Mps1p-myc) localizes to SPBs and kinetochores by immunofluorescence microscopy and immuno-EM analysis. This is consistent with the physical interaction we detect between Mps1p and Spc42p by coimmunoprecipitation. Spc42p is a substrate for Mps1p phosphorylation in vitro, and Spc42p phosphorylation is dependent on Mps1p in vivo. Finally, Spc42p assembly is abnormal in a mps1-1 mutant strain. We conclude that Mps1p regulates assembly of the integral SPB component Spc42p during SPB duplication.

Original languageEnglish (US)
Pages (from-to)453-465
Number of pages13
JournalJournal of Cell Biology
Volume156
Issue number3
DOIs
StatePublished - Feb 4 2002
Externally publishedYes

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Spindle Pole Bodies
Fungal Proteins
Protein Kinases
Alleles
Phosphorylation
Kinetochores
Mutant Proteins
Growth
Fluorescence Microscopy
Saccharomyces cerevisiae
Epitopes
Phosphotransferases
Phenotype
Mutation
DNA
Genes

Keywords

  • Budding yeast
  • MPS1
  • Protein kinase
  • SPC42
  • Spindle pole body

ASJC Scopus subject areas

  • Cell Biology

Cite this

The yeast protein kinase Mps1p is required for assembly of the integral spindle pole body component Spc42p. / Castillo, Andrea R.; Meehl, Janet B.; Morgan, Garry; Schutz-Geschwender, Amy; Winey, Mark.

In: Journal of Cell Biology, Vol. 156, No. 3, 04.02.2002, p. 453-465.

Research output: Contribution to journalArticle

Castillo, Andrea R. ; Meehl, Janet B. ; Morgan, Garry ; Schutz-Geschwender, Amy ; Winey, Mark. / The yeast protein kinase Mps1p is required for assembly of the integral spindle pole body component Spc42p. In: Journal of Cell Biology. 2002 ; Vol. 156, No. 3. pp. 453-465.
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abstract = "Saccharomyces cerevisiae MPS1 encodes an essential protein kinase that has roles in spindle pole body (SPB) duplication and the spindle checkpoint. Previously characterized MPS1 mutants fail in both functions, leading to aberrant DNA segregation with lethal consequences. Here, we report the identification of a unique conditional allele, mps1-8, that is defective in SPB duplication but not the spindle checkpoint. The mutations in mps1-8 are in the noncatalytic region of MPS1, and analysis of the mutant protein indicates that Mps1-8p has wild-type kinase activity in vitro. A screen for dosage suppressors of the mps1-8 conditional growth phenotype identified the gene encoding the integral SPB component SPC42. Additional analysis revealed that mps1-8 exhibits synthetic growth defects when combined with certain mutant alleles of SPC42. An epitope-tagged version of Mps1p (Mps1p-myc) localizes to SPBs and kinetochores by immunofluorescence microscopy and immuno-EM analysis. This is consistent with the physical interaction we detect between Mps1p and Spc42p by coimmunoprecipitation. Spc42p is a substrate for Mps1p phosphorylation in vitro, and Spc42p phosphorylation is dependent on Mps1p in vivo. Finally, Spc42p assembly is abnormal in a mps1-1 mutant strain. We conclude that Mps1p regulates assembly of the integral SPB component Spc42p during SPB duplication.",
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AB - Saccharomyces cerevisiae MPS1 encodes an essential protein kinase that has roles in spindle pole body (SPB) duplication and the spindle checkpoint. Previously characterized MPS1 mutants fail in both functions, leading to aberrant DNA segregation with lethal consequences. Here, we report the identification of a unique conditional allele, mps1-8, that is defective in SPB duplication but not the spindle checkpoint. The mutations in mps1-8 are in the noncatalytic region of MPS1, and analysis of the mutant protein indicates that Mps1-8p has wild-type kinase activity in vitro. A screen for dosage suppressors of the mps1-8 conditional growth phenotype identified the gene encoding the integral SPB component SPC42. Additional analysis revealed that mps1-8 exhibits synthetic growth defects when combined with certain mutant alleles of SPC42. An epitope-tagged version of Mps1p (Mps1p-myc) localizes to SPBs and kinetochores by immunofluorescence microscopy and immuno-EM analysis. This is consistent with the physical interaction we detect between Mps1p and Spc42p by coimmunoprecipitation. Spc42p is a substrate for Mps1p phosphorylation in vitro, and Spc42p phosphorylation is dependent on Mps1p in vivo. Finally, Spc42p assembly is abnormal in a mps1-1 mutant strain. We conclude that Mps1p regulates assembly of the integral SPB component Spc42p during SPB duplication.

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