The V260I mutation in fission yeast alpha-tubulin Atb2 affects microtubule dynamics and EB1-Mal3 localization and activates the Bub1 branch of the spindle checkpoint.
We have identified a novel temperature-sensitive mutant of fission yeast alpha-tubulin Atb2 (atb2-983) that contains a single amino acid substitution (V260I). Atb2-983 is incorporated into the microtubules, and their overall structures are not altered noticeably, but microtubule dynamics is compromised during interphase. atb2-983 displays a high rate of chromosome missegregation and is synthetically lethal with deletions in a subset of spindle checkpoint genes including bub1, bub3, and mph1, but not with mad1, mad2, and mad3. During early mitosis in this mutant, Bub1, but not Mad2, remains for a prolonged period in the kinetochores that are situated in proximity to one of the two SPBs (spindle pole bodies). High dosage mal3(+), encoding EB1 homologue, rescues atb2-983, suggesting that Mal3 function is compromised. Consistently, Mal3 localization and binding between Mal3 and Atb2-983 are impaired significantly, and a mal3 single mutant, such as atb2-983, displays prolonged Bub1 kinetochore localization. Furthermore in atb2-983 back-and-forth centromere oscillation during prometaphase is abolished. Intriguingly, this oscillation still occurs in the mal3 mutant, indicating that there is another defect independent of Mal3. These results show that microtubule dynamics is important for coordinated execution of mitotic events, in which Mal3 plays a vital role.
Asakawa
Kazuhide
K
Laboratory of Cell Regulation, Cancer Research UK, London Research Institute, Lincoln's Inn Fields Laboratories, London WC2A 3PX, UK.
Kume
Kazunori
K
Kanai
Muneyoshi
M
Goshima
Tetsuya
T
Miyahara
Kohji
K
Dhut
Susheela
S
Tee
Wee Wei
WW
Hirata
Dai
D
Toda
Takashi
T
D000483:Alleles; D000595:Amino Acid Sequence; D018797:Cell Cycle Proteins / Q000378:metabolism; D002842:Chromatids / Q000235:genetics; D020090:Chromosome Segregation / Q000235:genetics; D018628:Gene Dosage / Q000235:genetics; D018386:Kinetochores / Q000378:metabolism; D064246:Mad2 Proteins; D008869:Microtubule-Associated Proteins / Q000378:metabolism*; D008870:Microtubules / Q000378:metabolism*; D008969:Molecular Sequence Data; D020125:Mutation, Missense / Q000235:genetics; D009687:Nuclear Proteins / Q000378:metabolism; D049468:Prometaphase; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D021381:Protein Transport; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D012568:Schizosaccharomyces / Q000166:cytology / Q000378:metabolism*; D029702:Schizosaccharomyces pombe Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008941:Spindle Apparatus / Q000502:physiology*; D013489:Suppression, Genetic / Q000235:genetics; D014404:Tubulin / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
C510980:Atb2 protein, S pombe; D018797:Cell Cycle Proteins; C405804:EB1 microtubule binding proteins; D064246:Mad2 Proteins; C108851:Mal3 protein, S pombe; D008869:Microtubule-Associated Proteins; D009687:Nuclear Proteins; D011993:Recombinant Fusion Proteins; D029702:Schizosaccharomyces pombe Proteins; D014404:Tubulin; C577696:mad2 protein, S pombe; C495594:Bub1 protein, S pombe; D017346:Protein Serine-Threonine Kinases