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

Mentions: As multivalent polyamines significantly influence the nucleation duration, we studied whether spermidine affects the critical size of the nucleus. In Figure 7B, the slope of the curve without spermidine, n, is about −3, which indicates that 5 dimers form the critical nucleus, in agreement with previous reports (the slope is about −2.4 in [35], −3.7 in [33] and −4 in [36]). However this curve could not be used to analyze the evolution of the critical nucleus in the presence of spermidine because the experimental data are not properly fitted by a straight line. To proceed differently, we use the theory developed by Flyvbjerg et al. [34]. Assuming some scaling properties of the assembly curves, Flyvbjerg et al. obtained that the critical size of the nucleus is then 3p, where p is the slope of log [I(t)/I(∞)] versus log(t) ((I(t)) is the scattered light intensity at a time t). One of these scaling properties states that the tenth time scales like I(∞)−3 (equ. 3  in ref  [34]), which indicates that nucleation duration is related to the final mass of microtubules. Our experiments confirmed this scaling property without spermidine but, in the presence of 100 µM spermidine, the power law dependence does not fit properly the data (see Figure S2). Facilitated nucleation explains this finding since the tenth time is roughly constant for tubulin concentrations higher than CL. In spite of this, we show in Figure 8 the slope of [I(t)/I(∞)] versus log(t) extracted from light scattering curves because it is an interesting indicator of the critical size of the nucleus [34],[35]. The results show that spermidine does not change significantly the mean value of the assembly slope at early times (Figure 8), 2.36 for control and 2.32 with spermidine. This leads to a critical nucleus of about 7 dimers, slightly larger than 5 dimers obtained in Figure 7B. The critical size of the nucleus may then be not affected by the addition of spermidine. However, in the absence of a valid theory, this prediction remains 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 of                                spermidine.Initial slope of log                                [(I(t)/I(∞)] versus tubulin                                concentration. This slope is an interesting indicator of the number                                of tubulin dimers in the critical nuclei. The mean slope over                                tubulin concentrations is similar with or without spermidine, 2.36                                and 2.32 respectively. Spermidine may then not affect the critical                                size of the nucleus but this remains to be demonstrated with a valid                                theory. Inset: examples of log–log plot                                of [(I(t)/I(∞)]. The                                slopes were extracted at early times when the curves display a                                straight line.
© Copyright Policy
Related In: Results  -  Collection

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

pcbi-1000255-g008: Scaling properties and nucleus size in the presence of spermidine.Initial slope of log [(I(t)/I(∞)] versus tubulin concentration. This slope is an interesting indicator of the number of tubulin dimers in the critical nuclei. The mean slope over tubulin concentrations is similar with or without spermidine, 2.36 and 2.32 respectively. Spermidine may then not affect the critical size of the nucleus but this remains to be demonstrated with a valid theory. Inset: examples of log–log plot of [(I(t)/I(∞)]. The slopes were extracted at early times when the curves display a straight line.
Mentions: As multivalent polyamines significantly influence the nucleation duration, we studied whether spermidine affects the critical size of the nucleus. In Figure 7B, the slope of the curve without spermidine, n, is about −3, which indicates that 5 dimers form the critical nucleus, in agreement with previous reports (the slope is about −2.4 in [35], −3.7 in [33] and −4 in [36]). However this curve could not be used to analyze the evolution of the critical nucleus in the presence of spermidine because the experimental data are not properly fitted by a straight line. To proceed differently, we use the theory developed by Flyvbjerg et al. [34]. Assuming some scaling properties of the assembly curves, Flyvbjerg et al. obtained that the critical size of the nucleus is then 3p, where p is the slope of log [I(t)/I(∞)] versus log(t) ((I(t)) is the scattered light intensity at a time t). One of these scaling properties states that the tenth time scales like I(∞)−3 (equ. 3  in ref  [34]), which indicates that nucleation duration is related to the final mass of microtubules. Our experiments confirmed this scaling property without spermidine but, in the presence of 100 µM spermidine, the power law dependence does not fit properly the data (see Figure S2). Facilitated nucleation explains this finding since the tenth time is roughly constant for tubulin concentrations higher than CL. In spite of this, we show in Figure 8 the slope of [I(t)/I(∞)] versus log(t) extracted from light scattering curves because it is an interesting indicator of the critical size of the nucleus [34],[35]. The results show that spermidine does not change significantly the mean value of the assembly slope at early times (Figure 8), 2.36 for control and 2.32 with spermidine. This leads to a critical nucleus of about 7 dimers, slightly larger than 5 dimers obtained in Figure 7B. The critical size of the nucleus may then be not affected by the addition of spermidine. However, in the absence of a valid theory, this prediction remains 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