Antagonistic activities of Klp10A and Orbit regulate spindle length, bipolarity and function in vivo.
The metaphase-spindle steady-state length occurs as spindle microtubules ;flux', incorporating new subunits at their plus ends, while simultaneously losing subunits from their minus ends. Orbit/Mast/CLASP is required for tubulin subunit addition at kinetochores, and several kinesins regulate spindle morphology and/or flux by serving as microtubule depolymerases. Here, we use RNA interference in S2 cells to examine the relationship between Orbit and the four predicted kinesin-type depolymerases encoded by the Drosophila genome (Klp10A, Klp59C, Klp59D and Klp67A). Single depletion of Orbit results in monopolar spindles, mitotic arrest and a subsequent increase in apoptotic cells. These phenotypes are rescued by co-depleting Klp10A but none of the other three depolymerases. Spindle bipolarity is restored by preventing the spindle collapse seen in cells that lack Orbit, leading to functional spindles that are similar to controls in shape and length. We conclude that Klp10A exclusively antagonises Orbit in the regulation of bipolar spindle formation and maintenance.
Laycock
Joseph E
JE
Cancer Research UK Cell Cycle Genetics Group, University of Cambridge, Department of Genetics.
Savoian
Matthew S
MS
Glover
David M
DM
D000705:Anaphase / Q000502:physiology; D000818:Animals; D018797:Cell Cycle Proteins / Q000378:metabolism; D002460:Cell Line; D016764:Cell Polarity / Q000502:physiology*; D002478:Cells, Cultured; D002503:Centromere / Q000378:metabolism; D004330:Drosophila; D029721:Drosophila Proteins / Q000172:deficiency / Q000235:genetics / Q000378:metabolism*; D016547:Kinesins / Q000235:genetics / Q000378:metabolism*; D008869:Microtubule-Associated Proteins / Q000172:deficiency / Q000235:genetics / Q000378:metabolism*; D008870:Microtubules / Q000378:metabolism; D008938:Mitosis; D034622:RNA Interference; D012330:RNA, Double-Stranded / Q000502:physiology; D008941:Spindle Apparatus / Q000502:physiology*; D013997:Time Factors
C506005:BubR1 protein, Drosophila; D018797:Cell Cycle Proteins; D029721:Drosophila Proteins; D008869:Microtubule-Associated Proteins; D012330:RNA, Double-Stranded; C407307:chb protein, Drosophila; C481339:KLP10A protein, Drosophila; D016547:Kinesins