Kinesin-8 from fission yeast: a heterodimeric, plus-end-directed motor that can couple microtubule depolymerization to cargo movement.
Fission yeast expresses two kinesin-8s, previously identified and characterized as products of the klp5(+) and klp6(+) genes. These polypeptides colocalize throughout the vegetative cell cycle as they bind cytoplasmic microtubules during interphase, spindle microtubules, and/or kinetochores during early mitosis, and the interpolar spindle as it elongates in anaphase B. Here, we describe in vitro properties of these motor proteins and some truncated versions expressed in either bacteria or Sf9 cells. The motor-plus-neck domain of Klp6p formed soluble dimers that cross-linked microtubules and showed both microtubule-activated ATPase and plus-end-directed motor activities. Full-length Klp5p and Klp6p, coexpressed in Sf9 cells, formed soluble heterodimers with the same activities. The latter recombinant protein could also couple microbeads to the ends of shortening microtubules and use energy from tubulin depolymerization to pull a load in the minus end direction. These results, together with the spindle localizations of these proteins in vivo and their requirement for cell viability in the absence of the Dam1/DASH kinetochore complex, support the hypothesis that fission yeast kinesin-8 contributes both to chromosome congression to the metaphase plate and to the coupling of spindle microtubules to kinetochores during anaphase A.
Grissom
Paula M
PM
Department of Molecular, Cellular, and Developmental Biology, University of Colorado, Boulder, CO 80309-0347, USA.
Fiedler
Thomas
T
Grishchuk
Ekaterina L
EL
Nicastro
Daniela
D
West
Robert R
RR
McIntosh
J Richard
JR
D000251:Adenosine Triphosphatases / Q000378:metabolism; D000483:Alleles; D001692:Biological Transport / Q000187:drug effects; D003432:Cross-Linking Reagents / Q000378:metabolism; D005800:Genes, Fungal; D006160:Guanosine Triphosphate / Q000031:analogs & derivatives / Q000494:pharmacology; D016547:Kinesins / Q000378:metabolism*; D008863:Microspheres; D008869:Microtubule-Associated Proteins / Q000378:metabolism*; D008870:Microtubules / Q000187:drug effects / Q000378:metabolism* / Q000648:ultrastructure; D020409:Molecular Motor Proteins / Q000378:metabolism*; D050505:Mutant Proteins / Q000378:metabolism; D017239:Paclitaxel / Q000494:pharmacology; D011485:Protein Binding / Q000187:drug effects; D055503:Protein Multimerization / Q000187:drug effects; D012399:Rotation; D012568:Schizosaccharomyces / Q000201:enzymology / Q000235:genetics / Q000378:metabolism* / Q000648:ultrastructure; D029702:Schizosaccharomyces pombe Proteins / Q000378:metabolism*
D003432:Cross-Linking Reagents; D008869:Microtubule-Associated Proteins; D020409:Molecular Motor Proteins; D050505:Mutant Proteins; D029702:Schizosaccharomyces pombe Proteins; C004805:5'-guanylylmethylenebisphosphonate; D006160:Guanosine Triphosphate; D000251:Adenosine Triphosphatases; C469734:Klp5 protein, S pombe; C459102:Klp6 protein, S pombe; D016547:Kinesins; D017239:Paclitaxel