Roles of Polymerization Dynamics, Opposed Motors, and a Tensile Element in Governing the Length ofXenopusExtract Meiotic Spindles

Abstract

Metaphase spindles assemble to a steady state in length by mechanisms that involve microtubule dynamics and motor proteins, but they are incompletely understood. We found that Xenopus extract spindles recapitulate the length of egg meiosis II spindles, by using mechanisms intrinsic to the spindle. To probe these mechanisms, we perturbed microtubule polymerization dynamics and opposed motor proteins and measured effects on spindle morphology and dynamics. Microtubules were stabilized by hexylene glycol and inhibition of the catastrophe factor mitotic centromere-associated kinesin (MCAK) (a kinesin 13, previously called XKCM) and destabilized by depolymerizing drugs. The opposed motors Eg5 and dynein were inhibited separately and together. Our results are consistent with important roles for polymerization dynamics in regulating spindle length, and for opposed motors in regulating the relative stability of bipolar versus monopolar organization. The response to microtubule destabilization suggests that an unidentified tensile element acts in parallel with these conventional factors, generating spindle shortening force.

Cooperation Between Kinesin Motors Promotes Spindle Symmetry and Chromosome Organization in Oocytes

“…ASP has previously been linked to spindle positioning and asymmetric cell division in Drosophila (Rujano et al 2013) and mammals (Fish et al 2006;Gai et al 2016), but this represents a novel function for NCD and KLP54D. However, the observation that loss of kinesin-5 is rescued by loss of dynein at the poles (Miyamoto et al 2004;Mitchison et al 2005), may be mechanistically similar to the rescue we see by NCD and ASP. The relationship between kinesin-5 and kinesin-12 may appear as antagonism or cooperation, depending on the situation, which may reflect different effects on parallel and antiparallel MT bundles (Tanenbaum et al 2009;Vanneste et al 2009;Sturgill and Ohi 2013;Drechsler and McAinsh 2016).…”

Mechanical impulses can control metaphase progression in a mammalian cell

“…The time-lapse images of chromosome dynamics were then taken at 5-second intervals, either until the chromosomes were sufficiently separated (t TAO perturbations modulating the tension between sister chromatids have also been used to indirectly study the relationship between the persistent loss of tension and chromosome dynamics during mitosis. However, it is difficult to produce reversible transient perturbations on the order of 1-100 msec by biochemical techniques (e.g., even when using caged microtubule-destabilizing drugs) (31). Here, using precisely controlled mechanical perturbations by a pair of cantilevers, we made it possible to investigate how a human mitotic cell mechanochemically responds to an external mechanical impulse.…”

Kinesin-14 Family Proteins HSET/XCTK2 Control Spindle Length by Cross-Linking and Sliding Microtubules

“…Although different cell types form spindles with varying morphologies, for a certain cell type the spindle morphology remains steady (Mitchison et al, 2005). It is therefore critical to understand how cells organize and maintain the spindle machinery of a certain size and shape so that we can understand how spindle organization affects chromosome behavior.…”

“…The coordinated regulation of the dynamics and organization of these three classes of MTs is needed to form a morphologically normal spindle. Molecular perturbation of factors that regulate dynamics of MTs in the spindle or the addition of anti-MT drugs affects spindle organization and changes spindle length (Severin et al, 2001;Goshima et al, 2005b;Mitchison et al, 2005;Ohi et al, 2007). However, perturbation of molecular motors involved in MT sliding has given contradictory results on their roles in spindle length control (Hoyt et al, 1993;Sharp et al, 1999;Sharp et al, 2000a;Troxell et al, 2001;Goshima et al, 2005b;Burbank et al, 2007), raising the question of what are the principle mechanisms that control spindle length.…”