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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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AFM images of microtubules assembled in the presence of variousconcentrations of spermidine in buffer M with 30 µMtubulin.Scan area: 8×8 µm2. (A) Control. (B)200 µM spermidine. (C) 800 µM spermidine.Microtubules formed in the presence of moderate concentrations ofspermidine (<400 µm) had a normalappearance without any tendency for bundling. Higher spermidineconcentrations lead to the apparition of shorter MTs and someaberrant structures (oligomers, aggregates, see also Figure 3).
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pcbi-1000255-g002: AFM images of microtubules assembled in the presence of variousconcentrations of spermidine in buffer M with 30 µMtubulin.Scan area: 8×8 µm2. (A) Control. (B)200 µM spermidine. (C) 800 µM spermidine.Microtubules formed in the presence of moderate concentrations ofspermidine (<400 µm) had a normalappearance without any tendency for bundling. Higher spermidineconcentrations lead to the apparition of shorter MTs and someaberrant structures (oligomers, aggregates, see also Figure 3).

Mentions: We first checked by atomic force microscopy if the presence of polyamines, inthe physiological range, does not lead to aberrant microtubule structures.At low and moderate spermidine concentrations (<400µM in 25 mM MES-KOH pH 6.8, 20% glycerol),we observed the formation of long microtubules like in the absence ofpolyamines (Figures 2A,2B, and 3A). Athigher concentrations of spermidine (>400µM), very short microtubules, oligomers and (or)aggregates, and to a less extend MT bundles coexist (Figures 2C and 3B). It has already been observed thatmultivalent cations at high concentrations can promote the formation oflarge bundles [27]. The formation of small oligomers and(or) aggregates was also expected for a strong attraction regime(Regime III, see “elongation”section). The critical polyamine concentrations above whichaberrant structures appear increase with the ionic strength. For example, inthe presence of 800 µM spermidine and without KCl, smallaggregates or oligomers are observed whereas, with 100 mM KCl, only longmicrotubules are observed at 800 µM spermidine.


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)

AFM images of microtubules assembled in the presence of variousconcentrations of spermidine in buffer M with 30 µMtubulin.Scan area: 8×8 µm2. (A) Control. (B)200 µM spermidine. (C) 800 µM spermidine.Microtubules formed in the presence of moderate concentrations ofspermidine (<400 µm) had a normalappearance without any tendency for bundling. Higher spermidineconcentrations lead to the apparition of shorter MTs and someaberrant structures (oligomers, aggregates, see also Figure 3).
© Copyright Policy
Related In: Results  -  Collection

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

pcbi-1000255-g002: AFM images of microtubules assembled in the presence of variousconcentrations of spermidine in buffer M with 30 µMtubulin.Scan area: 8×8 µm2. (A) Control. (B)200 µM spermidine. (C) 800 µM spermidine.Microtubules formed in the presence of moderate concentrations ofspermidine (<400 µm) had a normalappearance without any tendency for bundling. Higher spermidineconcentrations lead to the apparition of shorter MTs and someaberrant structures (oligomers, aggregates, see also Figure 3).
Mentions: We first checked by atomic force microscopy if the presence of polyamines, inthe physiological range, does not lead to aberrant microtubule structures.At low and moderate spermidine concentrations (<400µM in 25 mM MES-KOH pH 6.8, 20% glycerol),we observed the formation of long microtubules like in the absence ofpolyamines (Figures 2A,2B, and 3A). Athigher concentrations of spermidine (>400µM), very short microtubules, oligomers and (or)aggregates, and to a less extend MT bundles coexist (Figures 2C and 3B). It has already been observed thatmultivalent cations at high concentrations can promote the formation oflarge bundles [27]. The formation of small oligomers and(or) aggregates was also expected for a strong attraction regime(Regime III, see “elongation”section). The critical polyamine concentrations above whichaberrant structures appear increase with the ionic strength. For example, inthe presence of 800 µM spermidine and without KCl, smallaggregates or oligomers are observed whereas, with 100 mM KCl, only longmicrotubules are observed at 800 µM spermidine.

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