Cik1 targets the minus-end kinesin depolymerase kar3 to microtubule plus ends.
Kar3, a Saccharomyces cerevisiae Kinesin-14, is essential for karyogamy and meiosis I but also has specific functions during vegetative growth. For its various roles, Kar3 forms a heterodimer with either Cik1 or Vik1, both of which are noncatalytic polypeptides. Here, we present the first biochemical characterization of Kar3Cik1, the kinesin motor that is essential for karyogamy. Kar3Cik1 depolymerizes microtubules from the plus end and promotes robust minus-end-directed microtubule gliding. Immunolocalization studies show that Kar3Cik1 binds preferentially to one end of the microtubule, whereas the Kar3 motor domain, in the absence of Cik1, exhibits significantly higher microtubule lattice binding. Kar3Cik1-promoted microtubule depolymerization requires ATP turnover, and the kinetics fit a single exponential function. The disassembly mechanism is not microtubule catastrophe like that induced by the MCAK Kinesin-13s. Soluble tubulin does not activate the ATPase activity of Kar3Cik1, and there is no evidence of Kar3Cik1(.)tubulin complex formation as observed for MCAK. These results reveal a novel mechanism to regulate microtubule depolymerization. We propose that Cik1 targets Kar3 to the microtubule plus end. Kar3Cik1 then uses its minus-end-directed force to depolymerize microtubules from the plus end, with each tubulin-subunit release event tightly coupled to one ATP turnover.
Sproul
Lisa R
LR
Department of Biological Sciences, University of Pittsburgh, Pittsburgh, Pennsylvania 15260, USA.
Anderson
Daniel J
DJ
Mackey
Andrew T
AT
Saunders
William S
WS
Gilbert
Susan P
SP
D000255:Adenosine Triphosphate / Q000378:metabolism; D002467:Cell Nucleus / Q000502:physiology*; D002850:Chromatography, Gel; D017931:DNA Primers; D016547:Kinesins / Q000378:metabolism*; D008868:Microtubule Proteins / Q000378:metabolism*; D008869:Microtubule-Associated Proteins / Q000378:metabolism*; D008870:Microtubules / Q000378:metabolism*; D010957:Plasmids / Q000235:genetics; D012441:Saccharomyces cerevisiae; D029701:Saccharomyces cerevisiae Proteins / Q000378:metabolism*
C076025:CIK1 protein, S cerevisiae; D017931:DNA Primers; C063504:KAR3 protein, S cerevisiae; D008868:Microtubule Proteins; D008869:Microtubule-Associated Proteins; D029701:Saccharomyces cerevisiae Proteins; D000255:Adenosine Triphosphate; D016547:Kinesins