Mechanism and regulation of kinesin‐5, an essential motor for the mitotic spindle

Waitzman, Joshua S.; Rice, Sarah E.

doi:10.1111/boc.201300054

Cited by 63 publications

(67 citation statements)

References 85 publications

(117 reference statements)

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“…This is in contrast to Kif15, which was inhibited only when the full-length protein was added. The reason for these differences is not clear; one possibility is that Eg5 is more susceptible to inhibition because of differences in the neck linker and stalk, which are unique to Eg5 (Waitzman and Rice, 2014). …”

Section: Discussionmentioning

confidence: 99%

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

Mann

Balchand

Wadsworth

2017

MBoC

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Section: Discussionmentioning

confidence: 99%

“…, 2012; Sturgill and Ohi, 2013). The existence of two mitotic motors that can each power bipolar spindle formation may contribute to the lack of efficacy of Eg5 inhibitors in clinical trials, and understanding how these motors are regulated therefore may be of clinical significance (Waitzman and Rice, 2014). …”

Section: Introductionmentioning

confidence: 99%

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

Mann

Balchand

Wadsworth

2017

MBoC

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“…Given the critical mitotic role of kinesin-5 motors and their corresponding importance as targets for antimitotic agents (21), it is important to understand how they switch direction. We have studied Cut7.…”

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confidence: 99%

Schizosaccharomyces pombe kinesin-5 switches direction using a steric blocking mechanism

Britto

Goulet

Rizvi

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

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Cut7, the sole kinesin-5 in Schizosaccharomyces pombe, is essential for mitosis. Like other yeast kinesin-5 motors, Cut7 can reverse its stepping direction, by mechanisms that are currently unclear. Here we show that for full-length Cut7, the key determinant of stepping direction is the degree of motor crowding on the microtubule lattice, with greater crowding converting the motor from minus enddirected to plus end-directed stepping. To explain how high Cut7 occupancy causes this reversal, we postulate a simple proximity sensing mechanism that operates via steric blocking. We propose that the minus end-directed stepping action of Cut7 is selectively inhibited by collisions with neighbors under crowded conditions, whereas its plus end-directed action, being less space-hungry, is not. In support of this idea, we show that the direction of Cut7-driven microtubule sliding can be reversed by crowding it with non-Cut7 proteins. Thus, crowding by either dynein microtubule binding domain or Klp2, a kinesin-14, converts Cut7 from net minus end-directed to net plus end-directed stepping. Biochemical assays confirm that the Cut7 N terminus increases Cut7 occupancy by binding directly to microtubules. Direct observation by cryoEM reveals that this occupancyenhancing N-terminal domain is partially ordered. Overall, our data point to a steric blocking mechanism for directional reversal through which collisions of Cut7 motor domains with their neighbors inhibit their minus end-directed stepping action, but not their plus enddirected stepping action. Our model can potentially reconcile a number of previous, apparently conflicting, observations and proposals for the reversal mechanism of yeast kinesins-5.Cut7 | kinesin-5 | bidirectional kinesin | mitotic kinesin | kinesin crowding K inesin-5 is essential for mitosis in many eukaryotes, from yeast to humans. Its main function appears to be to establish spindle bipolarity by driving the spindle poles apart during mitotic prophase (1). All kinesins-5 studied to date are homotetramers that crosslink and slide microtubules (MTs), and until recently, all were thought to be plus end-directed (2-8), meaning that their stepping action drives antiparallel MTs to slide slowly apart, with minus ends out. However, it was recently shown that a yeast kinesin-5, Saccharomyces cerevisae Cin8, not only can step toward MT plus ends, but also can move rapidly and processively toward MT minus ends (9, 10). The mechanism and biological significance of this bidirectionality are unknown.Several factors have already been shown to influence kinesin-5 directional switching. Cin8 is a tetramer (11) that steps toward plus ends when linking antiparallel MTs but toward minus ends when moving as a single molecule on un-crosslinked MTs. Crosslinking of two MTs is required for plus end-directed stepping (5), such that the engagement of only one MT turns off plus end-directed motility but supports minus end-directed motility (10, 12). The MT sliding direction of Cin8 also has been shown to reverse according to th...

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“…Stabilising the apo state might be expected to slow down ATP binding, as is observed [ 19 ], and potentially might drive MgADP release. Waitzman and Rice [ 20 ] and Muretta et al [ 18 ] both discuss a role of L5 in accelerating MgADP release in the absence of inhibitors.…”

Section: Role Of Loop 5 In Allosteric Inhibitionmentioning

confidence: 99%

Mechanisms of Action of Eg5 Inhibitors

Cross

2015

Kinesins and Cancer

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The kinetic mechanism of Eg5 is broadly similar to the widely studied kinesin-1 mechanism, but with some distinct features (Waitzman and Rice 2013 ). As with all molecular motors, the most stable state (the ground state) is the apo state, in which the active site is empty and the motor binds strongly (stably) and stereospecifi cally to its microtubule (MT) track. The subsequent steps of MgATP turnover progressively destabilise MT binding. The initial binding of MgATP or MgAMPPNP (a non-hydrolysable analog) into the empty active site drives the motor into a new conformational state, but this new motor·MgADP state binds to MTs only slightly less stably than the apo (empty) state, so that the motor head can still hold substantial force. The subsequent catalytic steps of hydrolysis and Pi release generate a motor·MgADP state that tends to detach rapidly from the MT. Rebinding of this motor·MgADP state to MTs triggers MgADP release, regenerating the apo state. In the motor·MgADP state, spontaneous release of MgADP from the active site is extremely slow, ~0.005 s −1 , and rate-limiting for ATP turnover in the absence of MTs. The weak binding motor·MgADP state is a "slip-state" that slides along the MT almost without friction if force is applied [ 1 ]. Nonetheless, this motor·MgADP slip-state is crucial because it provides access to a transient strong binding motor·MgADP state from which MgADP release is very fast: around three orders of magnitude faster than in the absence of MTs. The MT binding surface of the motor head is on the opposite side of the central beta sheet backbone to the active site, so that much of the structural mechanism of MT activation is necessarily allosteric. As I discuss below, the binding of allosteric inhibitors to the Eg5 motor head adds further stability to the motor·MgADP state and thereby inhibits MT-activated MgADP release. The crucial point therefore is that inhibitors and MTs have

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Mechanism and regulation of kinesin‐5, an essential motor for the mitotic spindle

Cited by 63 publications

References 85 publications

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

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

Schizosaccharomyces pombe kinesin-5 switches direction using a steric blocking mechanism

Mechanisms of Action of Eg5 Inhibitors

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