Minus-end-directed motor Ncd exhibits processive movement that is enhanced by microtubule bundling in vitro.
Drosophila Ncd, a kinesin-14A family member, is essential for meiosis and mitosis. Ncd is a minus-end-directed motor protein that has an ATP-independent microtubule binding site in the tail region, which enables it to act as a dynamic crosslinker of microtubules to assemble and maintain the spindle. Although a tailless Ncd has been shown to be nonprocessive, the role of the Ncd tail in single-molecule motility is unknown. Here, we show that individual Ncd dimers containing the tail region can move processively along microtubules at very low ionic strength, which provides the first evidence of processivity for minus-end-directed kinesins. The movement of GFP-Ncd consists of both a unidirectional and a diffusive element, and it was sensitive to ionic strength. Motility of a truncation series of Ncd and removal of the tubulin tail suggested that the Ncd tail serves as an electrostatic tether to microtubules. Under higher ionic conditions, Ncd showed only a small bias in diffusion along "single" microtubules, whereas it exhibited processive movement along "bundled" microtubules. This property may allow Ncd to accumulate preferentially in the vicinity of focused microtubules and then to crosslink and slide microtubules, possibly contributing to dynamic spindle self-organization.
Furuta
Ken'ya
K
Department of Life Sciences, Graduate School of Arts and Sciences, The University of Tokyo, Komaba 3-8-1, Meguro-ku, Tokyo 153-8902, Japan.
Toyoshima
Yoko Yano
YY
D000251:Adenosine Triphosphatases / Q000378:metabolism; D000818:Animals; D004330:Drosophila; D029721:Drosophila Proteins / Q000378:metabolism*; D049452:Green Fluorescent Proteins / Q000378:metabolism; D016547:Kinesins / Q000378:metabolism*; D008870:Microtubules / Q000378:metabolism*; D020836:Protein Structure, Quaternary; D055672:Static Electricity
D029721:Drosophila Proteins; C066696:ncd protein, Drosophila; D049452:Green Fluorescent Proteins; D000251:Adenosine Triphosphatases; D016547:Kinesins