Eg5 causes elongation of meiotic spindles when flux-associated microtubule depolymerization is blocked.
In higher eukaryotes, microtubules (MT) in both halves of the mitotic spindle translocate continuously away from the midzone in a phenomenon called poleward microtubule flux. Because the spindle maintains constant length and microtubule density, this microtubule translocation must somehow be coupled to net MT depolymerization at spindle poles. The molecular mechanisms underlying both flux-associated translocation and flux-associated depolymerization are not well understood, but it can be predicted that blocking pole-based destabilization will increase spindle length, an idea that has not been tested in meiotic spindles. Here, we show that simultaneous addition of two pole-disrupting reagents p50/dynamitin and a truncated version of Xklp2 results in continuous spindle elongation in Xenopus egg extracts, and we quantitatively correlate this elongation rate with the poleward translocation of stabilized microtubules. We further use this system to demonstrate that this poleward translocation requires the activity of the kinesin-related protein Eg5. These results suggest that Eg5 is responsible for flux-associated MT translocation and that dynein and Xklp2 regulate flux-associated microtubule depolymerization at spindle poles.
Shirasu-Hiza
Mimi
M
Department of Microbiology and Immunology, Stanford University, Stanford, CA 94305, USA. mshirasu@stanford.edu
Perlman
Zachary E
ZE
Wittmann
Torsten
T
Karsenti
Eric
E
Mitchison
Timothy J
TJ
D000818:Animals; D018797:Cell Cycle Proteins / Q000494:pharmacology; D002457:Cell Extracts; D000072159:Dynactin Complex; D005453:Fluorescence; D016547:Kinesins / Q000378:metabolism* / Q000494:pharmacology; D008863:Microspheres; D008869:Microtubule-Associated Proteins / Q000494:pharmacology; D008870:Microtubules / Q000187:drug effects* / Q000378:metabolism; D010063:Ovum / Q000378:metabolism; D008941:Spindle Apparatus / Q000187:drug effects* / Q000378:metabolism*; D014404:Tubulin / Q000378:metabolism; D014981:Xenopus; D029867:Xenopus Proteins / Q000378:metabolism* / Q000494:pharmacology
D018797:Cell Cycle Proteins; D002457:Cell Extracts; D000072159:Dynactin Complex; C088335:KIF11 protein, Xenopus; C097723:KIF15 protein, Xenopus; D008869:Microtubule-Associated Proteins; D014404:Tubulin; D029867:Xenopus Proteins; D016547:Kinesins