A mutation in gamma-tubulin alters microtubule dynamics and organization and is synthetically lethal with the kinesin-like protein pkl1p.
Mitotic segregation of chromosomes requires spindle pole functions for microtubule nucleation, minus end organization, and regulation of dynamics. gamma-Tubulin is essential for nucleation, and we now extend its role to these latter processes. We have characterized a mutation in gamma-tubulin that results in cold-sensitive mitotic arrest with an elongated bipolar spindle but impaired anaphase A. At 30 degrees C cytoplasmic microtubule arrays are abnormal and bundle into single larger arrays. Three-dimensional time-lapse video microscopy reveals that microtubule dynamics are altered. Localization of the mutant gamma-tubulin is like the wild-type protein. Prediction of gamma-tubulin structure indicates that non-alpha/beta-tubulin protein-protein interactions could be affected. The kinesin-like protein (klp) Pkl1p localizes to the spindle poles and spindle and is essential for viability of the gamma-tubulin mutant and in multicopy for normal cell morphology at 30 degrees C. Localization and function of Pkl1p in the mutant appear unaltered, consistent with a redundant function for this protein in wild type. Our data indicate a broader role for gamma-tubulin at spindle poles in regulating aspects of microtubule dynamics and organization. We propose that Pkl1p rescues an impaired function of gamma-tubulin that involves non-tubulin protein-protein interactions, presumably with a second motor, MAP, or MTOC component.
Paluh
J L
JL
Department of Molecular Biology, University of California, Berkeley, California 94720-3200, USA. jlpaluh@socrates.berkeley.edu
Nogales
E
E
Oakley
B R
BR
McDonald
K
K
Pidoux
A L
AL
Cande
W Z
WZ
D000595:Amino Acid Sequence; D020090:Chromosome Segregation; D005455:Fluorescent Antibody Technique; D005656:Fungal Proteins / Q000378:metabolism*; D016547:Kinesins / Q000378:metabolism*; D008854:Microscopy, Electron; D018715:Microscopy, Video; D008870:Microtubules / Q000378:metabolism*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009154:Mutation; D017434:Protein Structure, Tertiary; D012568:Schizosaccharomyces / Q000166:cytology / Q000235:genetics / Q000378:metabolism*; D029702:Schizosaccharomyces pombe Proteins; D016415:Sequence Alignment; D014404:Tubulin / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
D005656:Fungal Proteins; C104160:PKL1 protein, S pombe; D029702:Schizosaccharomyces pombe Proteins; D014404:Tubulin; D016547:Kinesins