How does taxol stabilize microtubules?

Arnal, Isabelle; Wade, Richard H.

doi:10.1016/s0960-9822(95)00180-1

Cited by 223 publications

(186 citation statements)

References 33 publications

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“…Although one difference between our studies was in the ionic strength of the buffers, we also noted that a potentially important difference between the two studies was that we used GMPCPPstabilized microtubules to analyze acetylation patterns on preformed microtubules, whereas Szyk et al (18) used Taxol-stabilized microtubules. Both the timing and method of Taxol introduction have been shown to affect the structure of Taxol microtubules (34). Our work has demonstrated that microtubule structure, and particularly the presence of defects and openings in the microtubule lattice, would have a substantial impact on the efficiency of αTAT1 in accessing and acetylating luminal acetylation sites.…”

Section: Discussionmentioning

confidence: 79%

Mechanism of microtubule lumen entry for the α-tubulin acetyltransferase enzyme αTAT1

Coombes

Yamamoto

McClellan

et al. 2016

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

100

113

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Microtubules are structural polymers inside of cells that are subject to posttranslational modifications. These posttranslational modifications create functionally distinct subsets of microtubule networks in the cell, and acetylation is the only modification that takes place in the hollow lumen of the microtubule. Although it is known that the α-tubulin acetyltransferase (αTAT1) is the primary enzyme responsible for microtubule acetylation, the mechanism for how αTAT1 enters the microtubule lumen to access its acetylation sites is not well understood. By performing biochemical assays, fluorescence and electron microscopy experiments, and computational simulations, we found that αTAT1 enters the microtubule lumen through the microtubule ends, and through bends or breaks in the lattice. Thus, microtubule structure is an important determinant in the acetylation process. In addition, once αTAT1 enters the microtubule lumen, the mobility of αTAT1 within the lumen is controlled by the affinity of αTAT1 for its acetylation sites, due to the rapid rebinding of αTAT1 onto highly concentrated α-tubulin acetylation sites. These results have important implications for how acetylation could gradually accumulate on stable subsets of microtubules inside of the cell.microtubule | acetylation | biophysics | microscopy | modeling

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Section: Discussionmentioning

confidence: 79%

Mechanism of microtubule lumen entry for the α-tubulin acetyltransferase enzyme αTAT1

Coombes

Yamamoto

McClellan

et al. 2016

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

100

113

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“…These consist of (i) an inward displacement in Taxol microtubules of the maxima of the J 0 Bessel function (corresponding to the low resolution transform of the excess electron density of the microtubule hollow cylinders), which serves to measure the different mean diameter (protofilament number) of each microtubule population and (ii) a small inward displacement of the J n and J n-3 Bessel function maxima in both types of taxoid induced microtubules with respect to glycerol microtubules (27,28). The latter is compatible with an increase in the spacing between tubulin monomers along the microtubule protofilament (38,39).…”

Section: Fig 4 Changes In Microtubule Protofilament Number Induced mentioning

confidence: 87%

“…The number of protofilaments corresponding to each image in this work were classified from the type of fringe pattern (27,51). This method has been widely employed (28,39,52,53). In addition, the orientation of the moiré fringe patterns in cryoelectron micrographs allows determination of microtubule polarity (54).…”

Section: Methodsmentioning

confidence: 99%

“…Will the protofilament number and the axial spacing slowly relax to the Taxol values? Arnal and Wade (39) have indicated that Taxol addition modifies only slightly the structure of assembled microtubules in comparison to assembly with Taxol, however, they did not report quantification of the Taxol binding in their study. On the other hand, during our studies of taxoid-induced assembly (27)(28)(29) we acquired preliminary synchrotron x-ray solution scattering data indicating that Taxol addition to preassembled microtubules decreases their diameter.…”

mentioning

confidence: 88%

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Changes in Microtubule Protofilament Number Induced by Taxol Binding to an Easily Accessible Site

Dı́az¹,

Valpuesta²,

Chacón³

et al. 1998

Journal of Biological Chemistry

112

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We have investigated the accessibility of the Taxolbinding site and the effects of Taxol binding on the structures of assembled microtubules. Taxol and docetaxel readily bind to and dissociate from microtubules, reaching 95% ligand exchange equilibrium in less than 3 min under our solution conditions (microtubules were previously assembled from GTP-tubulin, GTP-tubulin and microtubule-associated proteins, or GDP-tubulin and taxoid). Microtubules assembled from purified tubulin with Taxol are known to have typically one protofilament less than with the analogue docetaxel and control microtubules. Surprisingly, Taxol binding and exchange induce changes in the structure of preformed microtubules in a relatively short time scale. Cryoelectron microscopy shows changes toward the protofilament number distribution characteristic of Taxol or docetaxel, with a half-time of ϳ0.5 min, employing GDP-tubulintaxoid microtubules. Correspondingly, synchrotron xray solution scattering shows a reduction in the mean microtubule diameter upon Taxol binding to microtubules assembled from GTP-tubulin in glycerol-containing buffer, with a structural relaxation half-time of ϳ1 min. These results imply that microtubules can exchange protofilaments upon Taxol binding, due to internal dynamics along the microtubule wall. The simplest interpretation of the relatively fast taxoid exchange observed and labeling of cellular microtubules with fluorescent taxoids, is that the Taxol-binding site is at the outer microtubule surface. On the contrary, if Taxol binds at the microtubule lumen in agreement with the electron crystallographic structure of tubulin dimers, our results suggest that the inside of microtubules is easily accessible from the outer solution. Large pores or moving lattice defects in microtubules might facilitate the binding of taxoids, as well as of possible endogenous cellular ligands of the inner microtubule wall.

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“…A recent study revealed that there were soluble pools of tubulin sequestered and maintained in a temporary 'oligomeric' state, which retained assembling and disassembling potential (Liu & Lessman 2007). Upon exposure to microtubule promoting or destabilizing factors (such as g-tubulin, paclitaxel, or colchicines) (Arnal & Wade 1995;Amos & Löwe 1999;Casalou et al 2001;Raynaud-Messina & Merdes 2007), microtubules were observed to begin assembling or disassembling. It was also determined that after extraction and denaturation in boiling water, the a/b-tubulin dimer was depolymerized and a-tubulin was observed only in the supernatant.…”

Section: Biochemical Characterization Of A- G- and D-tubulinsmentioning

confidence: 99%

Localization of α-, γ-, and δ-tubulin in the hypotrich ciliateStylonychia pustulata(Hyportrichida, Ciliophora)

Yin

Sheng

Gao

et al. 2013

Animal Cells and Systems

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Localization of α-, γ-, and δ-tubulin in the hypotrich ciliate Stylonychiapustulata (Hyportrichida, Ciliophora), Animal Cells and Systems, 17:4, 259-268, DOI: 10.1080/19768354.2013.828654 To link to this article: https://doi.org/10. 1080/19768354.2013 Ciliature microtubular organelles, which organize into ciliary units, are highly complex microtubular systems. To investigate the formation of ciliature microtubular organelles and the role that different tubulins play in this process, we examined the distribution and function of a-, g-, and d-tubulins in the hypotrich ciliate Stylonychia pustulata. Immunofluorescence analysis demonstrated that a-tubulin localized to all microtubular organelles, while g-and dtubulins colocalized at basal bodies. Immunogold labeling revealed that g-and d-tubulins colocalize to the proximal part of basal bodies, suggesting that g-and d-tubulins probably interact and may function together. In addition, the permanent localization of g-tubulin to the adoral zone of membranelles suggests that g-tubulin play a role in the maintenance of oral ciliature base-associated microtubules, but does not play a role in the biogenesis of these ciliature. We observe that d-tubulin is restricted to the basal body and the connective fibers of the basal body, suggesting this protein may be required for the extension and stability of the microtubules in these structures. Dramatically different concentrations of these three tubulins were observed in the cytoskeleton and the insoluble and soluble fractions of the cell, suggesting that these highly related proteins perform different functions in S. pustulata.

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How does taxol stabilize microtubules?

Cited by 223 publications

References 33 publications

Mechanism of microtubule lumen entry for the α-tubulin acetyltransferase enzyme αTAT1

Mechanism of microtubule lumen entry for the α-tubulin acetyltransferase enzyme αTAT1

Changes in Microtubule Protofilament Number Induced by Taxol Binding to an Easily Accessible Site

Localization of α-, γ-, and δ-tubulin in the hypotrich ciliateStylonychia pustulata(Hyportrichida, Ciliophora)

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