Abstract
Microtubules polymerize from GTP-liganded tubulin dimers, but are essentially made of GDP-liganded tubulin. We investigate the tug-of-war resulting from the fact that GDP-liganded tubulin favors a curved configuration, but is forced to remain in a straight one when part of a microtubule. We point out that near the end of a microtubule, the proximity of the end shifts the balance in this tug-of-war, with some protofilament bending as result. This somewhat relaxes the microtubule lattice near its end, resulting in a structural cap. This structural cap thus is a simple mechanical consequence of two well-established facts: protofilaments made of GDP-liganded tubulin have intrinsic curvature, and microtubules are elastic, made from material that can yield to forces, in casu its own intrinsic forces. We explore possible properties of this structural cap, and demonstrate 1) how it allows both polymerization from GTP-liganded tubulin and rapid depolymerization in its absence; 2) how rescue can occur; 3) how a third, meta-stable intermediate state is possible and can explain some experimental results; and 4) how the tapered tips observed at polymerizing microtubule ends are stabilized during growth, though unable to accommodate a lateral cap. This scenario thus supports the widely accepted GTP-cap model by suggesting a stabilizing mechanism that explains the many aspects of dynamic instability.
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- Arnal I., Karsenti E., Hyman A. A. Structural transitions at microtubule ends correlate with their dynamic properties in Xenopus egg extracts. J Cell Biol. 2000 May 15;149(4):767–774. doi: 10.1083/jcb.149.4.767. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Arnal I., Wade R. H. How does taxol stabilize microtubules? Curr Biol. 1995 Aug 1;5(8):900–908. doi: 10.1016/s0960-9822(95)00180-1. [DOI] [PubMed] [Google Scholar]
- Billger M. A., Bhattacharjee G., Williams R. C., Jr, Bhatacharjee G. Dynamic instability of microtubules assembled from microtubule-associated protein-free tubulin: neither variability of growth and shortening rates nor "rescue" requires microtubule-associated proteins. Biochemistry. 1996 Oct 22;35(42):13656–13663. doi: 10.1021/bi9616965. [DOI] [PubMed] [Google Scholar]
- Caplow M. Microtubule dynamics. Curr Opin Cell Biol. 1992 Feb;4(1):58–65. doi: 10.1016/0955-0674(92)90059-l. [DOI] [PubMed] [Google Scholar]
- Caplow M., Ruhlen R. L., Shanks J. The free energy for hydrolysis of a microtubule-bound nucleotide triphosphate is near zero: all of the free energy for hydrolysis is stored in the microtubule lattice. J Cell Biol. 1994 Nov;127(3):779–788. doi: 10.1083/jcb.127.3.779. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Caplow M., Shanks J. Evidence that a single monolayer tubulin-GTP cap is both necessary and sufficient to stabilize microtubules. Mol Biol Cell. 1996 Apr;7(4):663–675. doi: 10.1091/mbc.7.4.663. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Caudron N., Valiron O., Usson Y., Valiron P., Job D. A reassessment of the factors affecting microtubule assembly and disassembly in vitro. J Mol Biol. 2000 Mar 17;297(1):211–220. doi: 10.1006/jmbi.2000.3554. [DOI] [PubMed] [Google Scholar]
- Chrétien D., Fuller S. D., Karsenti E. Structure of growing microtubule ends: two-dimensional sheets close into tubes at variable rates. J Cell Biol. 1995 Jun;129(5):1311–1328. doi: 10.1083/jcb.129.5.1311. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Chrétien D., Fuller S. D. Microtubules switch occasionally into unfavorable configurations during elongation. J Mol Biol. 2000 May 12;298(4):663–676. doi: 10.1006/jmbi.2000.3696. [DOI] [PubMed] [Google Scholar]
- Chrétien D., Jáinosi I., Taveau J. C., Flyvbjerg H. Microtubule's conformational cap. Cell Struct Funct. 1999 Oct;24(5):299–303. doi: 10.1247/csf.24.299. [DOI] [PubMed] [Google Scholar]
- Desai A., Mitchison T. J. Microtubule polymerization dynamics. Annu Rev Cell Dev Biol. 1997;13:83–117. doi: 10.1146/annurev.cellbio.13.1.83. [DOI] [PubMed] [Google Scholar]
- Dougherty C. A., Himes R. H., Wilson L., Farrell K. W. Detection of GTP and Pi in wild-type and mutated yeast microtubules: implications for the role of the GTP/GDP-Pi cap in microtubule dynamics. Biochemistry. 1998 Aug 4;37(31):10861–10865. doi: 10.1021/bi980677n. [DOI] [PubMed] [Google Scholar]
- Downing K. H., Nogales E. Tubulin and microtubule structure. Curr Opin Cell Biol. 1998 Feb;10(1):16–22. doi: 10.1016/s0955-0674(98)80082-3. [DOI] [PubMed] [Google Scholar]
- Drechsel D. N., Hyman A. A., Cobb M. H., Kirschner M. W. Modulation of the dynamic instability of tubulin assembly by the microtubule-associated protein tau. Mol Biol Cell. 1992 Oct;3(10):1141–1154. doi: 10.1091/mbc.3.10.1141. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Drechsel D. N., Kirschner M. W. The minimum GTP cap required to stabilize microtubules. Curr Biol. 1994 Dec 1;4(12):1053–1061. doi: 10.1016/s0960-9822(00)00243-8. [DOI] [PubMed] [Google Scholar]
- Erickson H. P., O'Brien E. T. Microtubule dynamic instability and GTP hydrolysis. Annu Rev Biophys Biomol Struct. 1992;21:145–166. doi: 10.1146/annurev.bb.21.060192.001045. [DOI] [PubMed] [Google Scholar]
- Erickson H. P. The structure and assembly of microtubules. Ann N Y Acad Sci. 1975 Jun 30;253:60–77. doi: 10.1111/j.1749-6632.1975.tb19193.x. [DOI] [PubMed] [Google Scholar]
- Felgner H., Frank R., Biernat J., Mandelkow E. M., Mandelkow E., Ludin B., Matus A., Schliwa M. Domains of neuronal microtubule-associated proteins and flexural rigidity of microtubules. J Cell Biol. 1997 Sep 8;138(5):1067–1075. doi: 10.1083/jcb.138.5.1067. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Flyvbjerg H, Holy TE, Leibler S. Microtubule dynamics: Caps, catastrophes, and coupled hydrolysis. Phys Rev E Stat Phys Plasmas Fluids Relat Interdiscip Topics. 1996 Nov;54(5):5538–5560. doi: 10.1103/physreve.54.5538. [DOI] [PubMed] [Google Scholar]
- Flyvbjerg H, Holy TE, Leibler S. Stochastic dynamics of microtubules: A model for caps and catastrophes. Phys Rev Lett. 1994 Oct 24;73(17):2372–2375. doi: 10.1103/PhysRevLett.73.2372. [DOI] [PubMed] [Google Scholar]
- Fygenson DK, Braun E, Libchaber A. Phase diagram of microtubules. Phys Rev E Stat Phys Plasmas Fluids Relat Interdiscip Topics. 1994 Aug;50(2):1579–1588. doi: 10.1103/physreve.50.1579. [DOI] [PubMed] [Google Scholar]
- Gildersleeve R. F., Cross A. R., Cullen K. E., Fagen A. P., Williams R. C., Jr Microtubules grow and shorten at intrinsically variable rates. J Biol Chem. 1992 Apr 25;267(12):7995–8006. [PubMed] [Google Scholar]
- Horio T., Hotani H. Visualization of the dynamic instability of individual microtubules by dark-field microscopy. Nature. 1986 Jun 5;321(6070):605–607. doi: 10.1038/321605a0. [DOI] [PubMed] [Google Scholar]
- Hyman A. A., Chrétien D., Arnal I., Wade R. H. Structural changes accompanying GTP hydrolysis in microtubules: information from a slowly hydrolyzable analogue guanylyl-(alpha,beta)-methylene-diphosphonate. J Cell Biol. 1995 Jan;128(1-2):117–125. doi: 10.1083/jcb.128.1.117. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Hyman A. A., Karsenti E. Morphogenetic properties of microtubules and mitotic spindle assembly. Cell. 1996 Feb 9;84(3):401–410. doi: 10.1016/s0092-8674(00)81285-4. [DOI] [PubMed] [Google Scholar]
- Hyman A. A., Salser S., Drechsel D. N., Unwin N., Mitchison T. J. Role of GTP hydrolysis in microtubule dynamics: information from a slowly hydrolyzable analogue, GMPCPP. Mol Biol Cell. 1992 Oct;3(10):1155–1167. doi: 10.1091/mbc.3.10.1155. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Jánosi I. M., Chrétien D., Flyvbjerg H. Modeling elastic properties of microtubule tips and walls. Eur Biophys J. 1998;27(5):501–513. doi: 10.1007/s002490050160. [DOI] [PubMed] [Google Scholar]
- Keates R. A., Hallett F. R. Dynamic instability of sheared microtubules observed by quasi-elastic light scattering. Science. 1988 Sep 23;241(4873):1642–1645. doi: 10.1126/science.241.4873.1642. [DOI] [PubMed] [Google Scholar]
- Kirschner M. W. Microtubule assembly and nucleation. Int Rev Cytol. 1978;54:1–71. doi: 10.1016/s0074-7696(08)60164-3. [DOI] [PubMed] [Google Scholar]
- Mandelkow E. M., Mandelkow E., Milligan R. A. Microtubule dynamics and microtubule caps: a time-resolved cryo-electron microscopy study. J Cell Biol. 1991 Sep;114(5):977–991. doi: 10.1083/jcb.114.5.977. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Mandelkow E. M., Mandelkow E. Unstained microtubules studied by cryo-electron microscopy. Substructure, supertwist and disassembly. J Mol Biol. 1985 Jan 5;181(1):123–135. doi: 10.1016/0022-2836(85)90330-4. [DOI] [PubMed] [Google Scholar]
- Mejillano M. R., Barton J. S., Himes R. H. Stabilization of microtubules by GTP analogues. Biochem Biophys Res Commun. 1990 Jan 30;166(2):653–660. doi: 10.1016/0006-291x(90)90859-l. [DOI] [PubMed] [Google Scholar]
- Melki R., Carlier M. F. Thermodynamics of tubulin polymerization into zinc sheets: assembly is not regulated by GTP hydrolysis. Biochemistry. 1993 Apr 6;32(13):3405–3413. doi: 10.1021/bi00064a026. [DOI] [PubMed] [Google Scholar]
- Melki R., Fievez S., Carlier M. F. Continuous monitoring of Pi release following nucleotide hydrolysis in actin or tubulin assembly using 2-amino-6-mercapto-7-methylpurine ribonucleoside and purine-nucleoside phosphorylase as an enzyme-linked assay. Biochemistry. 1996 Sep 17;35(37):12038–12045. doi: 10.1021/bi961325o. [DOI] [PubMed] [Google Scholar]
- Mickey B., Howard J. Rigidity of microtubules is increased by stabilizing agents. J Cell Biol. 1995 Aug;130(4):909–917. doi: 10.1083/jcb.130.4.909. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Mitchison T., Kirschner M. Dynamic instability of microtubule growth. Nature. 1984 Nov 15;312(5991):237–242. doi: 10.1038/312237a0. [DOI] [PubMed] [Google Scholar]
- Müller-Reichert T., Chrétien D., Severin F., Hyman A. A. Structural changes at microtubule ends accompanying GTP hydrolysis: information from a slowly hydrolyzable analogue of GTP, guanylyl (alpha,beta)methylenediphosphonate. Proc Natl Acad Sci U S A. 1998 Mar 31;95(7):3661–3666. doi: 10.1073/pnas.95.7.3661. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Nogales E. A structural view of microtubule dynamics. Cell Mol Life Sci. 1999 Oct 1;56(1-2):133–142. doi: 10.1007/s000180050012. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Nogales E., Whittaker M., Milligan R. A., Downing K. H. High-resolution model of the microtubule. Cell. 1999 Jan 8;96(1):79–88. doi: 10.1016/s0092-8674(00)80961-7. [DOI] [PubMed] [Google Scholar]
- O'Toole E. T., Winey M., McIntosh J. R. High-voltage electron tomography of spindle pole bodies and early mitotic spindles in the yeast Saccharomyces cerevisiae. Mol Biol Cell. 1999 Jun;10(6):2017–2031. doi: 10.1091/mbc.10.6.2017. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Odde D. J., Cassimeris L., Buettner H. M. Kinetics of microtubule catastrophe assessed by probabilistic analysis. Biophys J. 1995 Sep;69(3):796–802. doi: 10.1016/S0006-3495(95)79953-2. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Odde D. J., Ma L., Briggs A. H., DeMarco A., Kirschner M. W. Microtubule bending and breaking in living fibroblast cells. J Cell Sci. 1999 Oct;112(Pt 19):3283–3288. doi: 10.1242/jcs.112.19.3283. [DOI] [PubMed] [Google Scholar]
- Panda D., Chakrabarti G., Hudson J., Pigg K., Miller H. P., Wilson L., Himes R. H. Suppression of microtubule dynamic instability and treadmilling by deuterium oxide. Biochemistry. 2000 May 2;39(17):5075–5081. doi: 10.1021/bi992217f. [DOI] [PubMed] [Google Scholar]
- Panda Dulal, Miller Herbert P., Wilson Leslie. Determination of the size and chemical nature of the stabilizing "cap" at microtubule ends using modulators of polymerization dynamics. Biochemistry. 2002 Feb 5;41(5):1609–1617. doi: 10.1021/bi011767m. [DOI] [PubMed] [Google Scholar]
- Quarmby L. Cellular Samurai: katanin and the severing of microtubules. J Cell Sci. 2000 Aug;113(Pt 16):2821–2827. doi: 10.1242/jcs.113.16.2821. [DOI] [PubMed] [Google Scholar]
- Simon J. R., Salmon E. D. The structure of microtubule ends during the elongation and shortening phases of dynamic instability examined by negative-stain electron microscopy. J Cell Sci. 1990 Aug;96(Pt 4):571–582. doi: 10.1242/jcs.96.4.571. [DOI] [PubMed] [Google Scholar]
- Spittle C., Charrasse S., Larroque C., Cassimeris L. The interaction of TOGp with microtubules and tubulin. J Biol Chem. 2000 Jul 7;275(27):20748–20753. doi: 10.1074/jbc.M002597200. [DOI] [PubMed] [Google Scholar]
- Steinmetz M. O., Kammerer R. A., Jahnke W., Goldie K. N., Lustig A., van Oostrum J. Op18/stathmin caps a kinked protofilament-like tubulin tetramer. EMBO J. 2000 Feb 15;19(4):572–580. doi: 10.1093/emboj/19.4.572. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Stewart R. J., Farrell K. W., Wilson L. Role of GTP hydrolysis in microtubule polymerization: evidence for a coupled hydrolysis mechanism. Biochemistry. 1990 Jul 10;29(27):6489–6498. doi: 10.1021/bi00479a022. [DOI] [PubMed] [Google Scholar]
- Tran P. T., Joshi P., Salmon E. D. How tubulin subunits are lost from the shortening ends of microtubules. J Struct Biol. 1997 Mar;118(2):107–118. doi: 10.1006/jsbi.1997.3844. [DOI] [PubMed] [Google Scholar]
- Tran P. T., Walker R. A., Salmon E. D. A metastable intermediate state of microtubule dynamic instability that differs significantly between plus and minus ends. J Cell Biol. 1997 Jul 14;138(1):105–117. doi: 10.1083/jcb.138.1.105. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Vandecandelaere A., Brune M., Webb M. R., Martin S. R., Bayley P. M. Phosphate release during microtubule assembly: what stabilizes growing microtubules? Biochemistry. 1999 Jun 22;38(25):8179–8188. doi: 10.1021/bi9830765. [DOI] [PubMed] [Google Scholar]
- Vorobjev I. A., Svitkina T. M., Borisy G. G. Cytoplasmic assembly of microtubules in cultured cells. J Cell Sci. 1997 Nov;110(Pt 21):2635–2645. doi: 10.1242/jcs.110.21.2635. [DOI] [PubMed] [Google Scholar]
- Voter W. A., O'Brien E. T., Erickson H. P. Dilution-induced disassembly of microtubules: relation to dynamic instability and the GTP cap. Cell Motil Cytoskeleton. 1991;18(1):55–62. doi: 10.1002/cm.970180106. [DOI] [PubMed] [Google Scholar]
- Walker R. A., Inoué S., Salmon E. D. Asymmetric behavior of severed microtubule ends after ultraviolet-microbeam irradiation of individual microtubules in vitro. J Cell Biol. 1989 Mar;108(3):931–937. doi: 10.1083/jcb.108.3.931. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Walker R. A., O'Brien E. T., Pryer N. K., Soboeiro M. F., Voter W. A., Erickson H. P., Salmon E. D. Dynamic instability of individual microtubules analyzed by video light microscopy: rate constants and transition frequencies. J Cell Biol. 1988 Oct;107(4):1437–1448. doi: 10.1083/jcb.107.4.1437. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Walker R. A., Pryer N. K., Salmon E. D. Dilution of individual microtubules observed in real time in vitro: evidence that cap size is small and independent of elongation rate. J Cell Biol. 1991 Jul;114(1):73–81. doi: 10.1083/jcb.114.1.73. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Waterman-Storer C. M., Salmon E. D. Actomyosin-based retrograde flow of microtubules in the lamella of migrating epithelial cells influences microtubule dynamic instability and turnover and is associated with microtubule breakage and treadmilling. J Cell Biol. 1997 Oct 20;139(2):417–434. doi: 10.1083/jcb.139.2.417. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Waterman-Storer C. M., Salmon E. D. Microtubule dynamics: treadmilling comes around again. Curr Biol. 1997 Jun 1;7(6):R369–R372. doi: 10.1016/s0960-9822(06)00177-1. [DOI] [PubMed] [Google Scholar]
- Wiese C., Zheng Y. A new function for the gamma-tubulin ring complex as a microtubule minus-end cap. Nat Cell Biol. 2000 Jun;2(6):358–364. doi: 10.1038/35014051. [DOI] [PubMed] [Google Scholar]