Kinetochore targeting of fission yeast Mad and Bub proteins is essential for spindle checkpoint function but not for all chromosome segregation roles of Bub1p.
Several lines of evidence suggest that kinetochores are organizing centers for the spindle checkpoint response and the synthesis of a "wait anaphase" signal in cases of incomplete or improper kinetochore-microtubule attachment. Here we characterize Schizosaccharomyces pombe Bub3p and study the recruitment of spindle checkpoint components to kinetochores. We demonstrate by chromatin immunoprecipitation that they all interact with the central domain of centromeres, consistent with their role in monitoring kinetochore-microtubule interactions. Bub1p and Bub3p are dependent upon one another, but independent of the Mad proteins, for their kinetochore localization. We demonstrate a clear role for the highly conserved N-terminal domain of Bub1p in the robust targeting of Bub1p, Bub3p, and Mad3p to kinetochores and show that this is crucial for an efficient checkpoint response. Surprisingly, neither this domain nor kinetochore localization is required for other functions of Bub1p in chromosome segregation.
Vanoosthuyse
Vincent
V
Wellcome Trust Centre for Cell Biology, ICMB, University of Edinburgh, Edinburgh EH9 3JR, United Kingdom.
Valsdottir
Rebekka
R
Javerzat
Jean-Paul
JP
Hardwick
Kevin G
KG
D015825:Chromosomes, Fungal / Q000235:genetics / Q000378:metabolism; D004271:DNA, Fungal / Q000235:genetics; D005800:Genes, Fungal; D018386:Kinetochores / Q000378:metabolism*; D008938:Mitosis; D008954:Models, Biological; D034521:Nuclear Matrix-Associated Proteins / Q000235:genetics / Q000378:metabolism; D029543:Nucleocytoplasmic Transport Proteins / Q000235:genetics / Q000378:metabolism; D017434:Protein Structure, Tertiary; D012568:Schizosaccharomyces / Q000166:cytology / Q000235:genetics / Q000378:metabolism*; D029702:Schizosaccharomyces pombe Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008941:Spindle Apparatus / Q000378:metabolism
D004271:DNA, Fungal; D034521:Nuclear Matrix-Associated Proteins; D029543:Nucleocytoplasmic Transport Proteins; D029702:Schizosaccharomyces pombe Proteins; C092953:rae1 protein, S pombe