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Cited by 39Microtubule and mitosis dynamicsCancer Treatment and PharmacologySynthesis and biological activity

Microtubule detyrosination by VASH1/SVBP is regulated by the conformational state of tubulin in the lattice

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Yue, Yang, Hotta, Takashi, Higaki, Takumi, Verhey, Kristen J., Ohi, Ryoma·2023·Current Biology
DOI 10.1016/j.cub.2023.07.062PMC PMC10592207
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Abstract

Tubulin, a heterodimer of α- and β-tubulin, is a GTPase that assembles into microtubule (MT) polymers whose dynamic properties are intimately coupled to nucleotide hydrolysis. In cells, the organization and dynamics of MTs are further tuned by post-translational modifications (PTMs), which control the ability of MT-associated proteins (MAPs) and molecular motors to engage MTs. Detyrosination is a PTM of α-tubulin, wherein its C-terminal tyrosine residue is enzymatically removed by either the vasohibin (VASH) or MT-associated tyrosine carboxypeptidase (MATCAP) peptidases. How these enzymes generate specific patterns of MT detyrosination in cells is not known. Here, we use a novel antibody-based probe to visualize the formation of detyrosinated MTs in real time and employ single-molecule imaging of VASH1 bound to its regulatory partner small-vasohibin binding protein (SVBP) to understand the process of MT detyrosination in vitro and in cells. We demonstrate that the activity, but not binding, of VASH1/SVBP is much greater on mimics of guanosine triphosphate (GTP)-MTs than on guanosine diphosphate (GDP)-MTs. Given emerging data showing that tubulin subunits in GTP-MTs are in expanded conformation relative to tubulin subunits in GDP-MTs, we reasoned that the lattice conformation of MTs is a key factor that gates the activity of VASH1/SVBP. We show that Taxol, a drug known to expand the MT lattice, promotes MT detyrosination and that CAMSAP2 and CAMSAP3 are two MAPs that spatially regulate detyrosination in cells. Collectively, our work shows that VASH1/SVBP detyrosination is regulated by the conformational state of tubulin in the MT lattice and that this is spatially determined in cells by the activity of MAPs.

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Research works printed but not linked: 57

  1. Brouhard, G.J., and Rice, L.M. (2018). Microtubule dynamics: an interplay of biochemistry and mechanics. Nat. Rev. Mol. Cell Biol. 19, 451-463. https://doi.org/10.1038/s41580-018-0009-y. · DOI
  2. Alushin, G.M., Lander, G.C., Kellogg, E.H., Zhang, R., Baker, D., and Nogales, E. (2014). High-resolution microtubule structures reveal the structural transitions in ab-tubulin upon GTP hydrolysis. Cell 157, 1117- 1129. https://doi.org/10.1016/j.cell.2014.03.053. · DOI · Paper page
  3. Howes, S.C., Geyer, E.A., LaFrance, B., Zhang, R., Kellogg, E.H., Westermann, S., Rice, L.M., and Nogales, E. (2017). Structural differences between yeast and mammalian microtubules revealed by cryo-EM. J. Cell Biol. 216, 2669-2677. https://doi.org/10.1083/jcb.201612195. · DOI · Paper page
  4. Manka, S.W., and Moores, C.A. (2018). The role of tubulin-tubulin lattice contacts in the mechanism of microtubule dynamic instability. Nat. Struct. Mol. Biol. 25, 607-615. https://doi.org/10.1038/s41594-018-0087-8. · DOI · Paper page
  5. Zhang, R., Alushin, G.M., Brown, A., and Nogales, E. (2015). Mechanistic origin of microtubule dynamic instability and its modulation by EB pro- teins. Cell 162, 849-859. https://doi.org/10.1016/j.cell.2015.07.012. · DOI · Paper page
  6. Zhang, R., LaFrance, B., and Nogales, E. (2018). Separating the effects of nucleotide and EB binding on microtubule structure. Proc. Natl. Acad. Sci. USA 115, E6191-E6200. https://doi.org/10.1073/pnas.1802637115. · DOI · Paper page
  7. Janke, C., and Magiera, M.M. (2020). The tubulin code and its role in con- trolling microtubule properties and functions. Nat. Rev. Mol. Cell Biol. 21, 307-326. https://doi.org/10.1038/s41580-020-0214-3. · DOI · Paper page
  8. Roll-Mecak, A. (2020). The tubulin code in microtubule dynamics and in- formation encoding. Dev. Cell 54, 7-20. https://doi.org/10.1016/j.devcel. 2020.06.008. · DOI
  9. Rodrĭguez, J.A., Arce, C.A., Barra, H.S., and Caputto, R. (1973). Release of tyrosine incorporated as a single unit into rat brain protein. Biochem. Biophys. Res. Commun. 54, 335-340. https://doi.org/10.1016/0006- 291x(73)90927-3. · DOI
  10. Peris, L., Wagenbach, M., Lafaneche `re, L., Brocard, J., Moore, A.T., Kozielski, F., Job, D., Wordeman, L., and Andrieux, A. (2009). Motor- dependent microtubule disassembly driven by tubulin tyrosination. J. Cell Biol. 185, 1159-1166. https://doi.org/10.1083/jcb.200902142. · DOI
  11. Hotta, T., McAlear, T.S., Yue, Y., Higaki, T., Haynes, S.E., Nesvizhskii, A.I., Sept, D., Verhey, K.J., Bechstedt, S., and Ohi, R. (2022). EML2-S consti- tutes a new class of proteins that recognizes and regulates the dynamics of tyrosinated microtubules. Curr. Biol. 32, 3898-3910.e14. https://doi. org/10.1016/j.cub.2022.07.027. · DOI
  12. Steinmetz, M.O., and Akhmanova, A. (2008). Capturing protein tails by CAP-Gly domains. Trends Biochem. Sci. 33, 535-545. https://doi.org/ 10.1016/j.tibs.2008.08.006. · DOI
  13. McKenney, R.J., Huynh, W., Vale, R.D., and Sirajuddin, M. (2016). Tyrosination of alpha-tubulin controls the initiation of processive dynein- dynactin motility. EMBO J. 35, 1175-1185. https://doi.org/10.15252/ embj.201593071. · DOI · Paper page
  14. Nieuwenhuis, J., Adamopoulos, A., Bleijerveld, O.B., Mazouzi, A., Stickel, E., Celie, P., Altelaar, M., Knipscheer, P., Perrakis, A., Blomen, V.A., et al. (2017). Vasohibins encode tubulin detyrosinating activity. Science 358, 1453-1456. https://doi.org/10.1126/science.aao5676. · DOI · Paper page
  15. Souphron, J., Bodakuntla, S., Jijumon, A.S., Lakisic, G., Gautreau, A.M., Janke, C., and Magiera, M.M. (2019). Purification of tubulin with controlled post-translational modifications by polymerization-depolymerization cycles. Nat. Protoc. 14, 1634-1660. https://doi.org/10.1038/s41596-019-0153-7. · DOI
  16. Austin, B.P., To ¨zs er, J., Bagossi, P., Tropea, J.E., and Waugh, D.S. (2011). The substrate specificity of Metarhizium anisopliae and Bos taurus car- boxypeptidases A: insights into their use as tools for the removal of affinity tags. Protein Expr. Purif. 77, 53-61. https://doi.org/10.1016/j.pep.2010. 11.005. · DOI
  17. Cialek, C.A., Galindo, G., Koch, A.L., Saxton, M.N., and Stasevich, T.J. (2021). Bead loading proteins and nucleic acids into adherent human cells. J. Vis. Exp. https://doi.org/10.3791/62559. · DOI
  18. Ramirez-Rios, S., Choi, S.R., Sanyal, C., Blum, T.B., Bosc, C., Krichen, F., Denarier, E., Soleilhac, J.M., Blot, B., Janke, C., et al. (2023). VASH1-SVBP and VASH2-SVBP generate different detyrosination profiles on microtu- bules. J. Cell Biol. 222, https://doi.org/10.1083/jcb.202205096. · DOI
  19. Liu, H., and Shima, T. (2022). Preference of CAMSAP3 for microtubules with expanded lattice contributes to stabilization of the minus end. Preprint at bioRxiv. https://doi.org/10.1101/2022.08.21.504720. · DOI
  20. de Jager, L., Jansen, K.I., Kapitein, L.C., Fo ¨rster, F., and Howes, S.C. (2022). Increased microtubule lattice spacing correlates with selective binding of kinesin-1 in cells. Preprint at bioRxiv. https://doi.org/10.1101/ 2022.05.25.493428. · DOI · Paper page
  21. Tanaka, N., Meng, W., Nagae, S., and Takeichi, M. (2012). Nezha/ CAMSAP3 and CAMSAP2 cooperate in epithelial-specific organization of noncentrosomal microtubules. Proc. Natl. Acad. Sci. USA 109, 20029-20034. https://doi.org/10.1073/pnas.1218017109. · DOI
  22. Jiang, K., Hua, S., Mohan, R., Grigoriev, I., Yau, K.W., Liu, Q., Katrukha, E.A., Altelaar, A.F., Heck, A.J., Hoogenraad, C.C., et al. (2014). Microtubule minus-end stabilization by polymerization-driven CAMSAP deposition. Dev. Cell 28, 295-309. https://doi.org/10.1016/j.devcel.2014. 01.001. · DOI
  23. Liu, H., and Shima, T. (2023). Preference of CAMSAP3 for expanded microtubule lattice contributes to stabilization of the minus end. Life Sci. Alliance 6, 6. https://doi.org/10.26508/lsa.202201714. · DOI
  24. Atherton, J., Jiang, K., Stangier, M.M., Luo, Y., Hua, S., Houben, K., van Hooff, J.J.E., Joseph, A.P., Scarabelli, G., Grant, B.J., et al. (2017). A struc- tural model for microtubule minus-end recognition and protection by CAMSAP proteins. Nat. Struct. Mol. Biol. 24, 931-943. https://doi.org/ 10.1038/nsmb.3483. · DOI
  25. Atherton, J., Luo, Y., Xiang, S., Yang, C., Rai, A., Jiang, K., Stangier, M., Vemu, A., Cook, A.D., Wang, S., et al. (2019). Structural determinants of microtubule minus end preference in CAMSAP CKK domains. Nat. Commun. 10, 5236. https://doi.org/10.1038/s41467-019-13247-6. · DOI · Paper page
  26. Baines, A.J., Bignone, P.A., King, M.D., Maggs, A.M., Bennett, P.M., Pinder, J.C., and Phillips, G.W. (2009). The CKK domain (DUF1781) binds microtubules and defines the CAMSAP/ssp4 family of animal proteins. Mol. Biol. Evol. 26, 2005-2014. https://doi.org/10.1093/molbev/msp115. · DOI
  27. Moutin, M.J., Bosc, C., Peris, L., and Andrieux, A. (2021). Tubulin post- translational modifications control neuronal development and functions. Dev. Neurobiol. 81, 253-272. https://doi.org/10.1002/dneu.22774. · DOI
  28. Schulze, E., Asai, D.J., Bulinski, J.C., and Kirschner, M. (1987). Posttranslational modification and microtubule stability. J. Cell Biol. 105, 2167-2177. https://doi.org/10.1083/jcb.105.5.2167. · DOI
  29. Geeraert, C., Ratier, A., Pfisterer, S.G., Perdiz, D., Cantaloube, I., Rouault, A., Pattingre, S., Proikas-Cezanne, T., Codogno, P., and Pou ¨s, C. (2010). Starvation-induced hyperacetylation of tubulin is required for the stimula- tion of autophagy by nutrient deprivation. J. Biol. Chem. 285, 24184- 24194. https://doi.org/10.1074/jbc.M109.091553. · DOI
  30. Mackeh, R., Lorin, S., Ratier, A., Mejdoubi-Charef, N., Baillet, A., Bruneel, A., Hamaı ¨, A., Codogno, P., Pou ¨s, C., and Perdiz, D. (2014). Reactive ox- ygen species, AMP-activated protein kinase, and the transcription cofactor p300 regulate alpha-tubulin acetyltransferase-1 (aTAT-1/MEC- 17)-dependent microtubule hyperacetylation during cell stress. J. Biol. Chem. 289, 11816-11828. https://doi.org/10.1074/jbc.M113.507400. · DOI
  31. Ryu, N.M., and Kim, J.M. (2020). The role of the alpha-tubulin acetyltrans- ferase aTAT1 in the DNA damage response. J. Cell Sci. 133, https://doi. org/10.1242/jcs.246702. · DOI
  32. Yu, X., Chen, X., Amrute-Nayak, M., Allgeyer, E., Zhao, A., Chenoweth, H., Clement, M., Harrison, J., Doreth, C., Sirinakis, G., et al. (2021). MARK4 controls ischaemic heart failure through microtubule detyrosination. Nature 594, 560-565. https://doi.org/10.1038/s41586-021-03573-5. · DOI
  33. Cook, T.A., Nagasaki, T., and Gundersen, G.G. (1998). Rho guanosine tri- phosphatase mediates the selective stabilization of microtubules induced by lysophosphatidic acid. J. Cell Biol. 141, 175-185. https://doi.org/10. 1083/jcb.141.1.175. · DOI
  34. Gundersen, G.G., and Bulinski, J.C. (1988). Selective stabilization of mi- crotubules oriented toward the direction of cell migration. Proc. Natl. Acad. Sci. USA 85, 5946-5950. https://doi.org/10.1073/pnas.85. 16.5946. · DOI
  35. Wen, Y., Eng, C.H., Schmoranzer, J., Cabrera-Poch, N., Morris, E.J., Chen, M., Wallar, B.J., Alberts, A.S., and Gundersen, G.G. (2004). EB1 and APC bind to mDia to stabilize microtubules downstream of Rho and promote cell migration. Nat. Cell Biol. 6, 820-830. https://doi.org/10. 1038/ncb1160. · DOI
  36. Cassimeris, L., Guglielmi, L., Denis, V., Larroque, C., and Martineau, P. (2013). Specific in vivo labeling of tyrosinated alpha-tubulin and measure- ment of microtubule dynamics using a GFP tagged, cytoplasmically ex- pressed recombinant antibody. PLoS One 8, e59812. https://doi.org/10. 1371/journal.pone.0059812. · DOI
  37. Mikhaylova, M., Cloin, B.M., Finan, K., van den Berg, R., Teeuw, J., Kijanka, M.M., Sokolowski, M., Katrukha, E.A., Maidorn, M., Opazo, F., et al. (2015). Resolving bundled microtubules using anti-tubulin nanobod- ies. Nat. Commun. 6, 7933. https://doi.org/10.1038/ncomms8933. · DOI
  38. Kesarwani, S., Lama, P., Chandra, A., Reddy, P.P., Jijumon, A.S., Bodakuntla, S., Rao, B.M., Janke, C., Das, R., and Sirajuddin, M. (2020). Genetically encoded live-cell sensor for tyrosinated microtubules. J. Cell Biol. 219, https://doi.org/10.1083/jcb.201912107. · DOI · Paper page
  39. Li, F., Li, Y., Ye, X., Gao, H., Shi, Z., Luo, X., Rice, L.M., and Yu, H. (2020). Cryo-EM structure of VASH1-SVBP bound to microtubules. eLife 9, https://doi.org/10.7554/eLife.58157. · DOI
  40. Kellogg, E.H., Hejab, N.M.A., Howes, S., Northcote, P., Miller, J.H., Dı ´az, J.F., Downing, K.H., and Nogales, E. (2017). Insights into the distinct mechanisms of action of taxane and non-taxane microtubule stabilizers from cryo-EM structures. J. Mol. Biol. 429, 633-646. https://doi.org/10. 1016/j.jmb.2017.01.001. · DOI · Paper page
  41. Gundersen, G.G., Khawaja, S., and Bulinski, J.C. (1987). Postpolymerization detyrosination of alpha-tubulin: a mechanism for sub- cellular differentiation of microtubules. J. Cell Biol. 105, 251-264. https:// doi.org/10.1083/jcb.105.1.251. · DOI
  42. Dı ´az, J.F., Barasoain, I., and Andreu, J.M. (2003). Fast kinetics of Taxol binding to microtubules. Effects of solution variables and microtubule- associated proteins. J. Biol. Chem. 278, 8407-8419. https://doi.org/10. 1074/jbc.M211163200. · DOI
  43. Shannon, K.B., Canman, J.C., Ben Moree, C., Tirnauer, J.S., and Salmon, E.D. (2005). Taxol-stabilized microtubules can position the cytokinetic furrow in mammalian cells. Mol. Biol. Cell 16, 4423-4436. https://doi. org/10.1091/mbc.E04-11-0974. · DOI
  44. Prota, A.E., Lucena-Agell, D., Ma, Y., Estevez-Gallego, J., Li, S., Bargsten, K., Josa-Prado, F., Altmann, K.H., Gaillard, N., Kamimura, S., et al. (2023). Structural insight into the stabilization of microtubules by taxanes. eLife 12, https://doi.org/10.7554/eLife.84791. · DOI · Paper page
  45. Thawani, A., and Petry, S. (2021). Molecular insight into how gamma- TuRC makes microtubules. J. Cell Sci. 134, https://doi.org/10.1242/jcs. 245464. · DOI
  46. Rai, D., Hua, S., Monster, J.L., Stucchi, R., Stecker, K., Zhang, Y., Katrukha, E.A., Altelaar, M., Wieczorek, M., Jiang, K., et al. (2022). CAMSAP-driven microtubule release from g-TuRC and its regulation by nucleation-promoting factors. Preprint at bioRxiv. https://doi.org/10. 1101/2022.08.03.502613. · DOI
  47. Sanyal, C., Pietsch, N., Ramirez Rios, S., Peris, L., Carrier, L., and Moutin, M.J. (2023). The detyrosination/re-tyrosination cycle of tubulin and its role and dysfunction in neurons and cardiomyocytes. Semin. Cell Dev. Biol. 137, 46-62. https://doi.org/10.1016/j.semcdb.2021.12.006. · DOI
  48. Shima, T., Morikawa, M., Kaneshiro, J., Kambara, T., Kamimura, S., Yagi, T., Iwamoto, H., Uemura, S., Shigematsu, H., Shirouzu, M., et al. (2018). Kinesin-binding-triggered conformation switching of microtubules con- tributes to polarized transport. J. Cell Biol. 217, 4164-4183. https://doi. org/10.1083/jcb.201711178. · DOI · Paper page
  49. Duan, A.R., Jonasson, E.M., Alberico, E.O., Li, C., Scripture, J.P., Miller, R.A., Alber, M.S., and Goodson, H.V. (2017). Interactions between tau and different conformations of tubulin: implications for tau function and mechanism. J. Mol. Biol. 429, 1424-1438. https://doi.org/10.1016/j.jmb. 2017.03.018. · DOI
  50. Peet, D.R., Burroughs, N.J., and Cross, R.A. (2018). Kinesin expands and stabilizes the GDP-microtubule lattice. Nat. Nanotechnol. 13, 386-391. https://doi.org/10.1038/s41565-018-0084-4. · DOI · Paper page
  51. Schindelin, J., Arganda-Carreras, I., Frise, E., Kaynig, V., Longair, M., Pietzsch, T., Preibisch, S., Rueden, C., Saalfeld, S., Schmid, B., et al. (2012). Fiji: an open-source platform for biological-image analysis. Nat. Methods 9, 676-682. https://doi.org/10.1038/nmeth.2019. · DOI · Paper page
  52. Hotta, T., Haynes, S.E., Blasius, T.L., Gebbie, M., Eberhardt, E.L., Sept, D., Cianfrocco, M., Verhey, K.J., Nesvizhskii, A.I., and Ohi, R. (2021). Parthenolide destabilizes microtubules by covalently modifying tubulin. Curr. Biol. 31, 900-907.e6. https://doi.org/10.1016/j.cub.2020. 11.055. · DOI
  53. Khandelia, P., Yap, K., and Makeyev, E.V. (2011). Streamlined platform for short hairpin RNA interference and transgenesis in cultured mammalian cells. Proc. Natl. Acad. Sci. USA 108, 12799-12804. https://doi.org/10. 1073/pnas.1103532108. · DOI
  54. Mo, G.C.H., Posner, C., Rodriguez, E.A., Sun, T., and Zhang, J. (2020). A rationally enhanced red fluorescent protein expands the utility of FRET biosensors. Nat. Commun. 11, 1848. https://doi.org/10.1038/s41467- 020-15687-x. · DOI
  55. Hotta, T., Plemmons, A., Gebbie, M., Ziehm, T.A., Blasius, T.L., Johnson, C., Verhey, K.J., Pearring, J.N., and Ohi, R. (2023). Mechanistic analysis of CCP1 in generating DC2 a-tubulin in mammalian cells and photoreceptor neurons. Biomolecules 13, https://doi.org/10.3390/biom13020357. · DOI
  56. Kikukawa, K., Yoshimura, K., Watanabe, A., and Higaki, T. (2021). Metal- nano-ink coating for monitoring and quantification of cotyledon epidermal cell morphogenesis. Front. Plant Sci. 12, 745980. https://doi.org/10.3389/ fpls.2021.745980. · DOI
  57. Castoldi, M., and Popov, A.V. (2003). Purification of brain tubulin through two cycles of polymerization-depolymerization in a high-molarity buffer. Protein Expr. Purif. 32, 83-88. https://doi.org/10.1016/S1046-5928(03) 00218-3. · DOI

Cited by

Sorted by citations

Authors
Yue, Y., Hotta, T., Higaki, T., Verhey, K. J., & Ohi, R.
Affiliations
  • University of Michigan (US)
  • International Research Organization for Advanced Science and Technology (JP)
Corresponding author
  • Verhey, Kristen J.
  • Ohi, Ryoma
ORCID
Venue
Current Biology
Publisher
Elsevier BV
Published
2023-10
Volume
33
Issue
19
Pages
4111-4123.e7
ISSN
0960-9822 (ISSN-L)
DOI
10.1016/j.cub.2023.07.062
PMC
PMC10592207
OpenAlex
W4386769655
Cited by
39
References
4
Citations by year
2026: 21 · 2025: 14 · 2024: 4
FWCI
5.45
Type
journal article
Language
en
Open access
Licence
Locations
  • Current Biology (publishedVersion)
  • PubMed (publishedVersion)
  • PubMed Central (acceptedVersion)
Funders
  • National Institutes of Health (R01 GM086610, R35 GM131744)
  • University of Michigan
Topics
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Cell Biology › Microtubule and mitosis dynamics
  • Health Sciences › Medicine › Oncology › Cancer Treatment and Pharmacology
  • Physical Sciences › Chemistry › Organic Chemistry › Synthesis and biological activity
Keywords
microtubule detyrosination, vasohibin-1, alpha-tubulin, tubulin conformation, microtubule lattice, post-translational modification, paclitaxel, CAMSAP3, microtubule-associated proteins, single-molecule imaging
MeSH terms
Guanosine Triphosphate, Microtubules, Tubulin, Tyrosine, Paclitaxel

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