Kinesin-12 Kif15 Targets Kinetochore Fibers through an Intrinsic Two-Step Mechanism

Abstract

SUMMARY Proteins that recognize and act on specific subsets of microtubules (MTs) enable the varied functions of the MT cytoskeleton. We recently discovered that Kif15 localizes exclusively to kinetochore-fibers (K-fibers) [1, 2], or bundles of kinetochore-MTs within the mitotic spindle. It is currently speculated that the MT-associated protein TPX2 loads Kif15 onto spindle MTs [3–5], but this model has not been rigorously tested. Here, we show that Kif15 accumulates on MT bundles as a consequence of two inherent biochemical properties. First, Kif15 is self-repressed by its C-terminus. Second, Kif15 harbors a non-motor MT-binding site, enabling dimeric Kif15 to crosslink and slide MTs. Two-MT-binding activates Kif15, resulting in its accumulation on and motility within MT bundles but not on individual MTs. We propose that Kif15 targets K-fibers via an intrinsic two-step mechanism involving molecular unfolding and two-MT-binding. This work challenges the current model of Kif15 regulation and provides the first account of a kinesin that specifically recognizes a higher order MT array.

Regulation of Kif15 localization and motility by the C-terminus of TPX2 and microtubule dynamics

“…, 2014; Sturgill et al. , 2014) or motors in mammalian cell extracts (Dreschler and McAinsh, 2016; Sturgill et al.…”

Kinesin-12 motors cooperate to suppress microtubule catastrophes and drive the formation of parallel microtubule bundles

“…S8A). Parallel monopolar tetramers could cross-link microtubules by a second ATP-independent microtubule-binding site, which has been proposed recently (13). In this case, the velocity differential would be generated by unequal distribution of motor domains between the cross-linked microtubules (Fig.…”

“…One could argue that Kif15 tetramers do not cross-link microtubules by their two motor domain pairs but by one motor domain pair and the recently reported putative second microtubule-binding site within the Kif15 stalk (13). In that case, motor-driven movement along the substrate microtubule would passively drag the diffusive second microtubule-binding domain along the substrate microtubule.…”

“…Kif15 tetramers can also mediate the cross-linking of two microtubules, but give rise to very little microtubule sliding, except for episodes of short duration transport of short microtubules (12). A second study recently reported that a dimeric Kif15 missing the carboxyl-terminal half of the protein is also able to drive microtubule transport (13). One limitation of both studies is that they use stabilized microtubules rather than dynamic microtubules.…”

Resistance is not futile: Surviving Eg5 inhibition

“…Thus, Eg5 is converted in these cells from a motile force generator to a static MT crosslinker. By excessively bundling spindle MTs, the preferred substrate of Kif15, 6 the Eg5 rigor variant effectively upregulates Kif15 activity to rescue spindle assembly (Fig. 1).…”