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Polyamine sharing between tubulin dimers favours microtubule nucleation and elongation via facilitated diffusion.

Mechulam A, Chernov KG, Mucher E, Hamon L, Curmi PA, Pastré D - PLoS Comput. Biol. (2009)

Bottom Line: We suggest for the first time that the action of multivalent cations on microtubule dynamics can result from facilitated diffusion of GTP-tubulin to the microtubule ends.The mechanism of facilitated diffusion requires an attraction force between two tubulins, which can result from the sharing of multivalent counterions.The results presented here show that polyamines can be of particular importance for the regulation of the microtubule network in vivo and provide the basis for further investigations into the effects of facilitated diffusion on cytoskeleton dynamics.

View Article: PubMed Central - PubMed

Affiliation: Laboratoire Structure-Activité des Biomolécules Normales et Pathologiques, Université Evry-Val d'Essonne, Evry, France.

ABSTRACT
We suggest for the first time that the action of multivalent cations on microtubule dynamics can result from facilitated diffusion of GTP-tubulin to the microtubule ends. Facilitated diffusion can promote microtubule assembly, because, upon encountering a growing nucleus or the microtubule wall, random GTP-tubulin sliding on their surfaces will increase the probability of association to the target sites (nucleation sites or MT ends). This is an original explanation for understanding the apparent discrepancy between the high rate of microtubule elongation and the low rate of tubulin association at the microtubule ends in the viscous cytoplasm. The mechanism of facilitated diffusion requires an attraction force between two tubulins, which can result from the sharing of multivalent counterions. Natural polyamines (putrescine, spermidine, and spermine) are present in all living cells and are potent agents to trigger tubulin self-attraction. By using an analytical model, we analyze the implication of facilitated diffusion mediated by polyamines on nucleation and elongation of microtubules. In vitro experiments using pure tubulin indicate that the promotion of microtubule assembly by polyamines is typical of facilitated diffusion. The results presented here show that polyamines can be of particular importance for the regulation of the microtubule network in vivo and provide the basis for further investigations into the effects of facilitated diffusion on cytoskeleton dynamics.

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Scaling properties and nucleus size in the presence ofspermidine.Initial slope of log[(I(t)/I(∞)] versus tubulinconcentration. This slope is an interesting indicator of the numberof tubulin dimers in the critical nuclei. The mean slope overtubulin concentrations is similar with or without spermidine, 2.36and 2.32 respectively. Spermidine may then not affect the criticalsize of the nucleus but this remains to be demonstrated with a validtheory. Inset: examples of log–log plotof [(I(t)/I(∞)]. Theslopes were extracted at early times when the curves display astraight line.
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pcbi-1000255-g008: Scaling properties and nucleus size in the presence ofspermidine.Initial slope of log[(I(t)/I(∞)] versus tubulinconcentration. This slope is an interesting indicator of the numberof tubulin dimers in the critical nuclei. The mean slope overtubulin concentrations is similar with or without spermidine, 2.36and 2.32 respectively. Spermidine may then not affect the criticalsize of the nucleus but this remains to be demonstrated with a validtheory. Inset: examples of log–log plotof [(I(t)/I(∞)]. Theslopes were extracted at early times when the curves display astraight line.

Mentions: As multivalent polyamines significantly influence the nucleation duration, westudied whether spermidine affects the critical size of the nucleus. InFigure 7B, the slopeof the curve without spermidine, n, is about −3,which indicates that 5 dimers form the critical nucleus, in agreement withprevious reports (the slope is about −2.4 in [35],−3.7 in [33] and −4 in [36]). However thiscurve could not be used to analyze the evolution of the critical nucleus inthe presence of spermidine because the experimental data are not properlyfitted by a straight line. To proceed differently, we use the theorydeveloped by Flyvbjerg et al. [34]. Assumingsome scaling properties of the assembly curves, Flyvbjerg et al. obtainedthat the critical size of the nucleus is then 3p, wherep is the slope of log[I(t)/I(∞)] versuslog(t) ((I(t)) is the scattered light intensity ata time t). One of these scaling properties states that thetenth time scales likeI(∞)−3 (equ. 3 in ref [34]), which indicates that nucleation duration isrelated to the final mass of microtubules. Our experiments confirmed thisscaling property without spermidine but, in the presence of 100 µMspermidine, the power law dependence does not fit properly the data (seeFigureS2). Facilitated nucleation explains this finding since the tenthtime is roughly constant for tubulin concentrations higher thanCL. In spite of this, we show in Figure 8 the slope of[I(t)/I(∞)] versuslog(t) extracted from light scattering curves becauseit is an interesting indicator of the critical size of the nucleus [34],[35]. The resultsshow that spermidine does not change significantly the mean value of theassembly slope at early times (Figure 8), 2.36 for control and 2.32 with spermidine. This leadsto a critical nucleus of about 7 dimers, slightly larger than 5 dimersobtained in Figure 7B.The critical size of the nucleus may then be not affected by the addition ofspermidine. However, in the absence of a valid theory, this predictionremains to be tested.


Polyamine sharing between tubulin dimers favours microtubule nucleation and elongation via facilitated diffusion.

Mechulam A, Chernov KG, Mucher E, Hamon L, Curmi PA, Pastré D - PLoS Comput. Biol. (2009)

Scaling properties and nucleus size in the presence ofspermidine.Initial slope of log[(I(t)/I(∞)] versus tubulinconcentration. This slope is an interesting indicator of the numberof tubulin dimers in the critical nuclei. The mean slope overtubulin concentrations is similar with or without spermidine, 2.36and 2.32 respectively. Spermidine may then not affect the criticalsize of the nucleus but this remains to be demonstrated with a validtheory. Inset: examples of log–log plotof [(I(t)/I(∞)]. Theslopes were extracted at early times when the curves display astraight line.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2599886&req=5

pcbi-1000255-g008: Scaling properties and nucleus size in the presence ofspermidine.Initial slope of log[(I(t)/I(∞)] versus tubulinconcentration. This slope is an interesting indicator of the numberof tubulin dimers in the critical nuclei. The mean slope overtubulin concentrations is similar with or without spermidine, 2.36and 2.32 respectively. Spermidine may then not affect the criticalsize of the nucleus but this remains to be demonstrated with a validtheory. Inset: examples of log–log plotof [(I(t)/I(∞)]. Theslopes were extracted at early times when the curves display astraight line.
Mentions: As multivalent polyamines significantly influence the nucleation duration, westudied whether spermidine affects the critical size of the nucleus. InFigure 7B, the slopeof the curve without spermidine, n, is about −3,which indicates that 5 dimers form the critical nucleus, in agreement withprevious reports (the slope is about −2.4 in [35],−3.7 in [33] and −4 in [36]). However thiscurve could not be used to analyze the evolution of the critical nucleus inthe presence of spermidine because the experimental data are not properlyfitted by a straight line. To proceed differently, we use the theorydeveloped by Flyvbjerg et al. [34]. Assumingsome scaling properties of the assembly curves, Flyvbjerg et al. obtainedthat the critical size of the nucleus is then 3p, wherep is the slope of log[I(t)/I(∞)] versuslog(t) ((I(t)) is the scattered light intensity ata time t). One of these scaling properties states that thetenth time scales likeI(∞)−3 (equ. 3 in ref [34]), which indicates that nucleation duration isrelated to the final mass of microtubules. Our experiments confirmed thisscaling property without spermidine but, in the presence of 100 µMspermidine, the power law dependence does not fit properly the data (seeFigureS2). Facilitated nucleation explains this finding since the tenthtime is roughly constant for tubulin concentrations higher thanCL. In spite of this, we show in Figure 8 the slope of[I(t)/I(∞)] versuslog(t) extracted from light scattering curves becauseit is an interesting indicator of the critical size of the nucleus [34],[35]. The resultsshow that spermidine does not change significantly the mean value of theassembly slope at early times (Figure 8), 2.36 for control and 2.32 with spermidine. This leadsto a critical nucleus of about 7 dimers, slightly larger than 5 dimersobtained in Figure 7B.The critical size of the nucleus may then be not affected by the addition ofspermidine. However, in the absence of a valid theory, this predictionremains to be tested.

Bottom Line: We suggest for the first time that the action of multivalent cations on microtubule dynamics can result from facilitated diffusion of GTP-tubulin to the microtubule ends.The mechanism of facilitated diffusion requires an attraction force between two tubulins, which can result from the sharing of multivalent counterions.The results presented here show that polyamines can be of particular importance for the regulation of the microtubule network in vivo and provide the basis for further investigations into the effects of facilitated diffusion on cytoskeleton dynamics.

View Article: PubMed Central - PubMed

Affiliation: Laboratoire Structure-Activité des Biomolécules Normales et Pathologiques, Université Evry-Val d'Essonne, Evry, France.

ABSTRACT
We suggest for the first time that the action of multivalent cations on microtubule dynamics can result from facilitated diffusion of GTP-tubulin to the microtubule ends. Facilitated diffusion can promote microtubule assembly, because, upon encountering a growing nucleus or the microtubule wall, random GTP-tubulin sliding on their surfaces will increase the probability of association to the target sites (nucleation sites or MT ends). This is an original explanation for understanding the apparent discrepancy between the high rate of microtubule elongation and the low rate of tubulin association at the microtubule ends in the viscous cytoplasm. The mechanism of facilitated diffusion requires an attraction force between two tubulins, which can result from the sharing of multivalent counterions. Natural polyamines (putrescine, spermidine, and spermine) are present in all living cells and are potent agents to trigger tubulin self-attraction. By using an analytical model, we analyze the implication of facilitated diffusion mediated by polyamines on nucleation and elongation of microtubules. In vitro experiments using pure tubulin indicate that the promotion of microtubule assembly by polyamines is typical of facilitated diffusion. The results presented here show that polyamines can be of particular importance for the regulation of the microtubule network in vivo and provide the basis for further investigations into the effects of facilitated diffusion on cytoskeleton dynamics.

Show MeSH