Mitotic Spindle Positioning in Saccharomyces cerevisiae Is Accomplished by Antagonistically Acting Microtubule Motor Proteins

Abstract

Proper positioning of the mitotic spindle is often essential for cell division and differentiation processes. The asymmetric cell division characteristic of budding yeast, Saccharomyces cerevisiae, requires that the spindle be positioned at the mother–bud neck and oriented along the mother–bud axis. The single dynein motor encoded by the S. cerevisiae genome performs an important but nonessential spindle-positioning role. We demonstrate that kinesin-related Kip3p makes a major contribution to spindle positioning in the absence of dynein. The elimination of Kip3p function in dyn1Δ cells severely compromised spindle movement to the mother–bud neck. In dyn1Δ cells that had completed positioning, elimination of Kip3p function caused spindles to mislocalize to distal positions in mother cell bodies. We also demonstrate that the spindle-positioning defects exhibited by dyn1 kip3 cells are caused, to a large extent, by the actions of kinesin- related Kip2p. Microtubules in kip2Δ cells were shorter and more sensitive to benomyl than wild-type, in contrast to the longer and benomyl-resistant microtubules found in dyn1Δ and kip3Δ cells. Most significantly, the deletion of KIP2 greatly suppressed the spindle localization defect and slow growth exhibited by dyn1 kip3 cells. Likewise, induced expression of KIP2 caused spindles to mislocalize in cells deficient for dynein and Kip3p. Our findings indicate that Kip2p participates in normal spindle positioning but antagonizes a positioning mechanism acting in dyn1 kip3 cells. The observation that deletion of KIP2 could also suppress the inviability of dyn1Δ kar3Δ cells suggests that kinesin-related Kar3p also contributes to spindle positioning.

Two Related Kinesins,klp5+andklp6+, Foster Microtubule Disassembly and Are Required for Meiosis in Fission Yeast

“…Kip3p, Klp5p, and Klp6p also all localize to cytoplasmic and spindle microtubules (Figure 7) (DeZwaan et al, 1997). Furthermore, Klp5p and Klp6p share with Kip3p an activity that fosters microtubule disassembly, as evidenced by the unusually long and/or robust microtubules found in null mutants (Figures 3-6) (Cottingham and Hoyt, 1997;DeZwaan et al, 1997;Miller et al, 1998). Microtubule-disassembling activity has also been described for the KinI kinesin subfamily (XKCM1, Walczak et al, 1996;MCAK, Maney et al, 1998;Desai et al, 1999) and for Kar3p (Endow et al, 1994).…”

“…However, no such functions have been reported for KIP3 in budding yeast. Finally, klp5 ϩ and klp6 ϩ are both essential for meiosis, as discussed below, whereas KIP3 is not (Cottingham and Hoyt, 1997;Miller et al, 1998).…”

“…Of particular interest for our studies of klp5 ϩ and klp6 ϩ are the antagonistic interactions between budding yeast KIP3 and KIP2 (Cottingham and Hoyt, 1997;Miller et al, 1998), and the aspects of overlapping function between KIP3 and KAR3 (DeZwaan et al, 1997;Cottingham et al, 1999). Similar interactions in fission yeast have been investigated by constructing double, triple, and quadruple mutants between the klp⌬ strains and the relevant fission yeast homologs.…”

“…These promote microtubule disassembly and, together with dynein, are essential for meiosis (Troxell et al, 2001). S. pombe also expresses tea2 ϩ , a member of the KIP2 subfamily, which promotes microtubule growth (Cottingham and Hoyt, 1997). Deletion of tea2 ϩ leads to unusually short microtubules and the mislocalization of the tip-specific marker Tea1p (Browning et al, 2000).…”

Functional Analysis of Human Microtubule-based Motor Proteins, the Kinesins and Dyneins, in Mitosis/Cytokinesis Using RNA Interference

“…Database analysis indicates that Kif18 shares significant sequence homology with the Kip3 subfamily of N-terminal positioned motor kinesins, Kip3 (S. cerevisiae), Klp5/6 (Schizosaccharomyces pombe), KipB (Aspergillus nidulans), and Klp67A (Drosophila) (unpublished data and Cottingham and Hoyt, 1997;Garcia et al, 2002;West et al, 2002;Gandhi et al, 2004;Rischitor et al, 2004;Savoian et al, 2004). Klp5/6, KipB, and Klp67A are involved in spindle formation and chromosome segregation during meiosis and/or mitosis (Garcia et al, 2002;West et al, 2002;Gandhi et al, 2004;Rischitor et al, 2004;Savoian et al, 2004).…”