A mutation uncouples the tubulin conformational and GTPase cycles, revealing allosteric control of microtubule dynamics

Geyer, Elisabeth A.; Burns, Alexander S.; Lalonde, Beth A.; Ye, Xuecheng; Piedra, Felipe Andrés; Huffaker, Tim C.; Rice, Luke M.

doi:10.7554/elife.10113

Cited by 88 publications

(148 citation statements)

References 55 publications

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“…In particular, these mutants suppress catastrophe frequency by magnitudes similar to what is achieved by regulatory proteins or microtubule stabilizing drugs (Mohan et al, 2013; Wieczorek et al, 2015). Structural and mutagenesis studies have indicated that conformational changes in tubulin’s intermediate domain are coupled to GTP hydrolysis upon microtubule polymerization (Geyer et al, 2015; Nogales et al, 1998; Ravelli et al, 2004). In contrast, the effects of mutating residues located in tubulin’s kinesin binding site on microtubule dynamics have not been explored.…”

Section: Discussionmentioning

confidence: 99%

Mutations in Human Tubulin Proximal to the Kinesin-Binding Site Alter Dynamic Instability at Microtubule Plus- and Minus-Ends

Pamula

Howes

et al. 2016

Developmental Cell

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The assembly of microtubule-based cellular structures depends on regulated tubulin polymerization and directional transport. Here, we purify and characterize tubulin heterodimers that have human β-tubulin isotype III (TUBB3), and heterodimers with one of two β-tubulin mutations (D417H or R262H). Both point mutations are proximal to the kinesin binding site and have been linked to an ocular motility disorder in humans. Compared to wild-type, microtubules with these mutations have decreased catastrophe frequencies and increased average lifetimes of plus- and minus-end stabilizing caps. Importantly, the D417H mutation does not alter microtubule lattice structure or Mal3 binding to growing filaments. Instead, this mutation reduces the affinity of tubulin for TOG domains and colchicine, suggesting that the distribution of tubulin heterodimer conformations is changed. Together, our findings reveal how residues on the surface of microtubules, distal from the GTP-hydrolysis site and inter-subunit contacts, can alter polymerization dynamics at the plus- and minus-ends of microtubules.

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Section: Discussionmentioning

confidence: 99%

Mutations in Human Tubulin Proximal to the Kinesin-Binding Site Alter Dynamic Instability at Microtubule Plus- and Minus-Ends

Pamula

Howes

et al. 2016

Developmental Cell

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“…Typical microtubule lifetimes at steady state are several hundreds of seconds, much longer than the maturation time (24,25,(45)(46)(47)(48)(49); how these two timescales are linked is not clear. Currently, there is no exact agreement on the criterion for inducing catastrophe.…”

Section: Discussionmentioning

confidence: 99%

Steady-state EB cap size fluctuations are determined by stochastic microtubule growth and maturation

Rickman

Duellberg

Cade

et al. 2017

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

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Growing microtubules are protected from depolymerization by the presence of a GTP or GDP/Pi cap. End-binding proteins of the EB1 family bind to the stabilizing cap, allowing monitoring of its size in real time. The cap size has been shown to correlate with instantaneous microtubule stability. Here we have quantitatively characterized the properties of cap size fluctuations during steadystate growth and have developed a theory predicting their timescale and amplitude from the kinetics of microtubule growth and cap maturation. In contrast to growth speed fluctuations, cap size fluctuations show a characteristic timescale, which is defined by the lifetime of the cap sites. Growth fluctuations affect the amplitude of cap size fluctuations; however, cap size does not affect growth speed, indicating that microtubules are far from instability during most of their time of growth. Our theory provides the basis for a quantitative understanding of microtubule stability fluctuations during steady-state growth.microtubules | dynamic instability | GTP cap | EB1 | biochemical network

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“…, 2010), were introduced into the flow chambers and imaged at 30°C by total internal reflection fluorescence microscopy as described previously (Geyer et al. , 2015).…”

Section: Methodsmentioning

confidence: 99%

GDP-to-GTP exchange on the microtubule end can contribute to the frequency of catastrophe

Piedra

Kim

Garza

et al. 2016

MBoC

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A mutation uncouples the tubulin conformational and GTPase cycles, revealing allosteric control of microtubule dynamics

Cited by 88 publications

References 55 publications

Mutations in Human Tubulin Proximal to the Kinesin-Binding Site Alter Dynamic Instability at Microtubule Plus- and Minus-Ends

Mutations in Human Tubulin Proximal to the Kinesin-Binding Site Alter Dynamic Instability at Microtubule Plus- and Minus-Ends

Steady-state EB cap size fluctuations are determined by stochastic microtubule growth and maturation

GDP-to-GTP exchange on the microtubule end can contribute to the frequency of catastrophe

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