title: Dual mechanisms regulate the recruitment of spindle assembly checkpoint proteins to the budding yeast kinetochore.
abstract: Recruitment of spindle assembly checkpoint (SAC) proteins by an unattached kinetochore leads to SAC activation. This recruitment is licensed by the Mps1 kinase, which phosphorylates the kinetochore protein Spc105 at one or more of its six MELT repeats. Spc105 then recruits the Bub3-Bub1 and Mad1-Mad2 complexes, which produce the inhibitory signal that arrests cell division. The strength of this signal depends, in part, on the number of Bub3-Bub1 and Mad1-Mad2 molecules that Spc105 recruits. Therefore regulation of this recruitment will influence SAC signaling. To understand this regulation, we established the physiological binding curves that describe the binding of Bub3-Bub1 and Mad1-Mad2 to the budding yeast kinetochore. We find that the binding of both follows the mass action law. Mps1 likely phosphorylates all six MELT repeats of Spc105. However, two mechanisms prevent Spc105 from recruiting six Bub3-Bub1 molecules: low Bub1 abundance and hindrance in the binding of more than one Bub3-Bub1 molecule to the same Spc105. Surprisingly, the kinetochore recruits two Mad1-Mad2 heterotetramers for every Bub3-Bub1 molecule. Finally, at least three MELT repeats per Spc105 are needed for accurate chromosome segregation. These data reveal that kinetochore-intrinsic and -extrinsic mechanisms influence the physiological operation of SAC signaling, potentially to maximize chromosome segregation accuracy.
authors.affiliation: Cell and Developmental Biology, University of Michigan Medical School, Ann Arbor, MI 48109.
authors.forename: Pavithra
authors.initials: P
authors.lastname: Aravamudhan
authors.affiliation: Cell and Developmental Biology, University of Michigan Medical School, Ann Arbor, MI 48109.
authors.forename: Renjie
authors.initials: R
authors.lastname: Chen
authors.affiliation: Cell and Developmental Biology, University of Michigan Medical School, Ann Arbor, MI 48109.
authors.forename: Babhrubahan
authors.initials: B
authors.lastname: Roy
authors.affiliation: Cell and Developmental Biology, University of Michigan Medical School, Ann Arbor, MI 48109.
authors.forename: Janice
authors.initials: J
authors.lastname: Sim
authors.affiliation: Cell and Developmental Biology, University of Michigan Medical School, Ann Arbor, MI 48109 ajitj@umich.edu.
authors.forename: Ajit P
authors.initials: AP
authors.lastname: Joglekar
mesh_terms: D059447:Cell Cycle Checkpoints; D018797:Cell Cycle Proteins / Q000378:metabolism; D002868:Chromosomal Proteins, Non-Histone / Q000378:metabolism; D020090:Chromosome Segregation; D018386:Kinetochores / Q000378:metabolism* / Q000502:physiology; D059566:M Phase Cell Cycle Checkpoints / Q000502:physiology*; D064246:Mad2 Proteins / Q000378:metabolism; D008869:Microtubule-Associated Proteins / Q000378:metabolism; D009687:Nuclear Proteins / Q000378:metabolism; D010766:Phosphorylation / Q000502:physiology; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000378:metabolism; D004718:Saccharomycetales / Q000378:metabolism; D015398:Signal Transduction; D008941:Spindle Apparatus / Q000378:metabolism* / Q000502:physiology
chemical_list: C070352:BUB3 protein, S cerevisiae; C584280:Bub1 protein, S cerevisiae; D018797:Cell Cycle Proteins; D002868:Chromosomal Proteins, Non-Histone; C501382:MAD1 protein, S cerevisiae; C577695:MAD2 protein, S cerevisiae; D064246:Mad2 Proteins; D008869:Microtubule-Associated Proteins; D009687:Nuclear Proteins; D029701:Saccharomyces cerevisiae Proteins; C546124:Spc105 protein, S cerevisiae; D017346:Protein Serine-Threonine Kinases