Depolymerizing Kinesins Kip3 and MCAK Shape Cellular Microtubule Architecture by Differential Control of Catastrophe

Abstract

Microtubules are dynamic filaments whose ends alternate between periods of slow growth and rapid shortening as they explore intracellular space and move organelles. A key question is how regulatory proteins modulate catastrophe, the conversion from growth to shortening. To study this process, we reconstituted microtubule dynamics in the absence and presence of the kinesin-8 Kip3 and the kinesin-13 MCAK. Surprisingly, we found that, even in the absence of the kinesins, the microtubule catastrophe frequency depends on the age of the microtubule, indicating that catastrophe is a multistep process. Kip3 slowed microtubule growth in a length-dependent manner and increased the rate of aging. In contrast, MCAK eliminated the aging process. Thus, both kinesins are catastrophe factors; Kip3 mediates fine control of microtubule length by narrowing the distribution of maximum lengths prior to catastrophe, whereas MCAK promotes rapid restructuring of the microtubule cytoskeleton by making catastrophe a first-order random process.

Theoretical Analysis of Microtubule Dynamics at All Times

“…We associated the shrinking phase of the microtubules with polymer configurations that have at least It is found that the experimental observations can be well fit by N = 3, W T = 37.7 s −1 , r = 0.0036 s −1 , with other parameters as listed in Table 1, and with the free tubulin concentration C T = 12µM as was used in experiments. 19 Our calculations show that N < 3 cases cannot describe experimental cumulative distributions. The fact that fitted values are close to experimentally measured values of relevant parameters can be viewed as an additional argument that supports our theoretical picture.…”

“…It agrees with recent in vitro experimental observations. 19 In these experiments microtubules were growing from the seeds composed of GMPCPP-tubulins, which are essentially the non-hydrolyzable analogs of GTP-tubulins. As we predict, the large protective cap of unhydrolyzed subunits makes f c (t) much smaller at early times.…”

“…In reality, the hydrolyzed monomers detach much faster (see Table 1). In addition, in vitro experiments 19 were done under conditions when no rescues are observed, and the catastrophe times were measured as the the times before the first catastrophe occurs. To make a quantitative test of our theoretical model, we performed Monte Carlo computer simulations for the same experimental conditions with realistic values of the Tsubunit attachment rate constants k on and the detachment rate for D-subunits W D as given in Table 1.…”

“…However, these theoretical views have been challenged recently by high-resolution measurements of filament length and lifetime distributions. [17][18][19][20] In these experiments, deviations from exponential distributions expected for the single-transition picture have been observed. 19,20 In addition, it was found that dynamic properties of microtubules change with time.…”

“…[17][18][19][20] In these experiments, deviations from exponential distributions expected for the single-transition picture have been observed. 19,20 In addition, it was found that dynamic properties of microtubules change with time. 17,19,20 Furthermore, experiments show that various types of kinesin motor proteins bound to microtubules might influence these aging phenomena differently.…”