Perturbations in Microtubule Mechanics from Tubulin Preparation

Hawkins, Taviare L.; Mirigian, Matthew; Li, Jingqiang; Yaşar, Mahmut; Sackett, Dan L.; Sept, David; Ross, Jennifer L.

doi:10.1007/s12195-012-0229-8

Cited by 28 publications

(59 citation statements)

References 37 publications

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“…1B), and the distribution of individually measured microtubule persistence lengths is wide, with values ranging over an order of magnitude ( Fig. 1C), consistent with prior studies of microtubule mechanics [Gittes et al, 1993;Kurachi et al, 1995;Kikumoto et al, 2006;Hawkins et al, 2012Hawkins et al, , 2013Valdman et al, 2012;Yu et al, 2012]. In comparing the distributions of microtubules assembled under various conditions, the statistical significance of differences was determined using the Kolmogorov-Smirnov test (Table I).…”

Section: Microtubule Stiffness Depends On Nucleotide Type and Presencsupporting

confidence: 84%

“…However, when we add paclitaxel during polymerization, the microtubules are significantly more compliant, suggesting that the importance of a particular stabilizer on microtubule mechanics may be driven by its ability to act during polymerization, when it can influence the overall protofilament number and the nature of the tubulin‐tubulin interactions [Matesanz et al, ; Alushin et al, ]. We also note that although we find similar trends, our measured values of persistence length are systematically larger than those reported by Hawkins et al It is possible that differences in tubulin source or purification method lead to systematic variations in stiffness [Hawkins et al, ] or that the differences arise from our different approaches to spectral analysis [Valdman et al, ]. Our method uses global fitting routines to express the filament contour shape in terms of a series of Chebyshev polynomials, rather than local fitting of discrete points to a Fourier series.…”

Section: Discussionsupporting

confidence: 70%

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Mechanical effects of EB1 on microtubules depend on GTP hydrolysis state and presence of paclitaxel

López

Valentine

2014

Cytoskeleton

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Using the nonhydrolyzable GTP analog GMPCPP and the slowly hydrolyzable GTPγS, we polymerize microtubules that recapitulate the end binding behavior of the plus end interacting protein (+TIP) EB1 along their entire length, and use these to investigate the impact of EB1 binding on microtubule mechanics. To measure the stiffness of single filaments, we use a spectral analysis method to determine the ensemble of shapes adopted by a freely diffusing, fluorescently labeled microtubule. We find that the presence of EB1 can stiffen microtubules in a manner that depends on the hydrolysis state of the tubulin-bound nucleotide, as well as the presence of the small-molecule stabilizer paclitaxel. We find that the magnitude of the EB1-induced stiffening is not proportional to the EB1-microtubule binding affinity, suggesting that the stiffening effect does not arise purely from an increase in the total amount of bound EB1. Additionally, we find that EB1 binds cooperatively to microtubules in manner that depends on tubulin-bound nucleotide state.

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Section: Microtubule Stiffness Depends On Nucleotide Type and Presencsupporting

confidence: 84%

Section: Discussionsupporting

confidence: 70%

Mechanical effects of EB1 on microtubules depend on GTP hydrolysis state and presence of paclitaxel

López

Valentine

2014

Cytoskeleton

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“…Consistent with prior studies, we find the overall distribution of MT stiffness to be broad and non-Gaussian, so we characterize the filament stiffness at each condition by the median value and indicate the percentile differences in the distributions using a box and whiskers plot format [Hawkins et al, 2012]. As shown in Fig.…”

Section: Effects Of Small-molecule Stabilizers and Tau On Mt Stiffnesssupporting

confidence: 75%

Mechanical and functional properties of epothilone‐stabilized microtubules

Pessino

Kuei

et al. 2012

Cytoskeleton

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Using a suite of biophysical tools, we assess the mechanical, structural, and functional properties of microtubules (MTs) stabilized by the chemotherapeutic compounds epothilone-A, epothilone-B, and taxol in vitro. We demonstrate that MTs stabilized by epothilone-A or epothilone-B are competent to bind tau proteins and support kinesin translocation. Kinesin speed is sensitive not only to the type of small molecule stabilizer used but also to the presence of the essential MT-associated protein tau. Epothilone-stabilized MTs are substantially less stiff than taxol-stabilized MTs. The addition of tau proteins to MTs stabilized by either epothilone compound or taxol further reduces stiffness. Taken together, these results suggest that small molecule stabilizers do not simply stabilize a "native" MT structure, but rather they modulate the structure, function, and mechanics of the MTs they bind. This may have important consequences to the therapeutic use of these agents in cancer chemotherapies.

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“…For a particular MT preparation, the experimental values of stiffness reported, even by a single laboratory, show a broad and non-Gaussian distribution [115,120], which some groups have described as log-normal [112]. The distribution is typically wide, with as much as an order of magnitude variation from maximal to minimal values, much larger than the errors expected from the image processing and spectral analysis steps, which has been estimated to be less than 10% [44].…”

Section: The Challenges Of Measuring Mt Stiffnessmentioning

confidence: 98%

“…It is clear that microtubule stiffness is sensitive to a large number of parameters: tubulin source [112], polymerization rate [113], presence of GTP or GTP analog [114,115], small molecule stabilizers [115][116][117][118][119][120], and microtubule associating proteins (MAPs) [96,114,115,[121][122][123][124]. While there appear to be consistent stiffness values reported within a particular research group for a fixed condition, the values vary significantly among different laboratories, even under seemingly similar MT preparation conditions.…”

Section: The Challenges Of Measuring Mt Stiffnessmentioning

confidence: 99%

Molecular control of stress transmission in the microtubule cytoskeleton

López

Valentine

2015

Biochimica et Biophysica Acta (BBA) - Molecular Cell Research

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In this article, we will summarize recent progress in understanding the mechanical origins of rigidity, strength, resiliency and stress transmission in the MT cytoskeleton using reconstituted networks formed from purified components. We focus on the role of network architecture, crosslinker compliance and dynamics, and molecular determinants of single filament elasticity, while highlighting open questions and future directions for this work.

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Perturbations in Microtubule Mechanics from Tubulin Preparation

Cited by 28 publications

References 37 publications

Mechanical effects of EB1 on microtubules depend on GTP hydrolysis state and presence of paclitaxel

Mechanical effects of EB1 on microtubules depend on GTP hydrolysis state and presence of paclitaxel

Mechanical and functional properties of epothilone‐stabilized microtubules

Molecular control of stress transmission in the microtubule cytoskeleton

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