Mouse KL2 is a unique MTSE involved in chromosome-based spindle organization and regulated by multiple kinases during female meiosis

Abstract

Microtubule-severing enzymes (MTSEs) play important roles in mitosis and meiosis of the primitive organisms. However, their roles in mammalian female meiosis, which accounts for over 80% of gamete-originated human reproductive diseases, remain unexplored. In the current study, we reported that katanin-like 2 (KL2) was the only MTSE concentrating at chromosomes. Furthermore, the knockdown of KL2 significantly reduced the chromosome-based increase in the microtubule (MT) polymer, increased aberrant kinetochore-MT (K-MT) attachment, delayed meiosis, and severely affected normal fertility. We demonstrated that the inhibition of aurora B, a key kinase for correcting aberrant K-MT attachment, significantly eliminated KL2 expression from chromosomes. Additionally, KL2 interacted with phosphorylated eukaryotic elongation factor-2 kinase, and they competed for chromosome binding. Phosphorylated KL2 was also localized at spindle poles, with its phosphorylation regulated by extracellular signal-regulated kinase 1/2. In summary, the current study reveals a novel function of MTSEs in mammalian female meiosis and demonstrates that multiple kinases coordinate to regulate the levels of KL2 at chromosomes.

Keywords

mouse / KL2 / MTSE / kinase / female meiosis

Fundings

This work was financially supported by the Youth Program of National Natural Science Foundation of China (Grant No. 82001539 to Leilei Gao), the Zhejiang Province Health Innovation Talent Project (Grant No. 2021RC001 to Zhen Jin), the General Program of the National Natural Science Foundation of China (Grant No. 31671561 to Dong Zhang), and the Regional Program of National Natural Science Foundation of China (Grant No. 82260126 to Xiaocong Liu).

Acknowledgments

We thank Dr. Daniel W. Buster from Cancer Center of the University of Arizona, Professor David J. Sharp and Professor Hernando J. Sosa from the Department of Physiology & Biophysics of Albert Einstein College of Medicine, Professor Hongshan Chen from School of Pharmacy, Nanjing Medical University, Professor Xingyin Liu from State Key Lab of Reproductive Medicine and Offspring Health, Nanjing Medical University, and Professor Jianhuan Chen from Wuxi Medical School, Jiangnan University for their advice and proofreading of the manuscript.

References

[1] Nogales E, Wang H. Structural intermediates in microtubule assembly and disassembly: how and why?[J]. Curr Opin Cell Biol, 2006, 18(2): 179-184. doi: 10.1016/j.ceb.2006.02.009

[2] Brouhard GJ. Dynamic instability 30 years later: complexities in microtubule growth and catastrophe[J]. Mol Biol Cell, 2015, 26(7): 1207-1210. doi: 10.1091/mbc.E13-10-0594

[3] Gardner MK, Zanic M, Howard J. Microtubule catastrophe and rescue[J]. Curr Opin Cell Biol, 2013, 25(1): 14-22. doi: 10.1016/j.ceb.2012.09.006

[4] Walczak CE, Heald R. Mechanisms of mitotic spindle assembly and function[J]. Int Rev Cytol, 2008, 265: 111-158. https://www.sciencedirect.com/science/article/abs/pii/S0074769607650037

[5] Errico A, Ballabio A, Rugarli EI. Spastin, the protein mutated in autosomal dominant hereditary spastic paraplegia, is involved in microtubule dynamics[J]. Hum Mol Genet, 2002, 11(2): 153-163. doi: 10.1093/hmg/11.2.153

[6] Smith LB, Milne L, Nelson N, et al. KATNAL1 regulation of sertoli cell microtubule dynamics is essential for spermiogenesis and male fertility[J]. PLoS Genet, 2012, 8(5): e1002697. doi: 10.1371/journal.pgen.1002697

[7] Mukherjee S, Valencia JDD, Stewman S, et al. Human Fidgetin is a microtubule severing the enzyme and minus-end depolymerase that regulates mitosis[J]. Cell Cycle, 2012, 11(12): 2359-2366. doi: 10.4161/cc.20849

[8] Nakamura M, Ehrhardt DW, Hashimoto T. Microtubule and katanin-dependent dynamics of microtubule nucleation complexes in the acentrosomal Arabidopsis cortical array[J]. Nat Cell Biol, 2010, 12(11): 1064-1070. doi: 10.1038/ncb2110

[9] Sharma N, Bryant J, Wloga D, et al. Katanin regulates dynamics of microtubules and biogenesis of motile cilia[J]. J Cell Biol, 2007, 178(6): 1065-1079. doi: 10.1083/jcb.200704021

[10] Monroe N, Hill CP. Meiotic clade AAA ATPases: protein polymer disassembly machines[J]. J Mol Biol, 2016, 428(9): 1897-1911. doi: 10.1016/j.jmb.2015.11.004

[11] Vale RD. AAA proteins: lords of the ring[J]. J Cell Biol, 2000, 150(1): F13-F20. doi: 10.1083/jcb.150.1.F13

[12] Roll-Mecak A, Vale RD. Structural basis of microtubule severing by the hereditary spastic paraplegia protein spastin[J]. Nature, 2008, 451(7176): 363-367. doi: 10.1038/nature06482

[13] Johjima A, Noi K, Nishikori S, et al. Microtubule severing by katanin p60 AAA+ ATPase requires the C-terminal acidic tails of both α- and β-tubulins and basic amino acid residues in the AAA+ ring pore[J]. J Biol Chem, 2015, 290(18): 11762-11770. doi: 10.1074/jbc.M114.614768

[14] Sharp DJ, Ross JL. Microtubule-severing enzymes at the cutting edge[J]. J Cell Sci, 2012, 125(Pt 11): 2561-2569.

[15] Roll-Mecak A, McNally FJ. Microtubule-severing enzymes[J]. Curr Opin Cell Biol, 2010, 22(1): 96-103. doi: 10.1016/j.ceb.2009.11.001

[16] Friel CT, Howard J. The kinesin-13 MCAK has an unconventional ATPase cycle adapted for microtubule depolymerization[J]. EMBO J, 2011, 30(19): 3928-3939. doi: 10.1038/emboj.2011.290

[17] Tanenbaum ME, Macurek L, van der Vaart B, et al. A complex of Kif18b and MCAK promotes microtubule depolymerization and is negatively regulated by Aurora kinases[J]. Curr Biol, 2011, 21(16): 1356-1365. doi: 10.1016/j.cub.2011.07.017

[18] Howard J, Hyman AA. Microtubule polymerases and depolymerases[J]. Curr Opin Cell Biol, 2007, 19(1): 31-35. doi: 10.1016/j.ceb.2006.12.009

[19] Wordeman L. Microtubule-depolymerizing kinesins[J]. Curr Opin Cell Biol, 2005, 17(1): 82-88. doi: 10.1016/j.ceb.2004.12.003

[20] Hoffmann I. Centrosomes in mitotic spindle assembly and orientation[J]. Curr Opin Struct Biol, 2021, 66: 193-198. doi: 10.1016/j.sbi.2020.11.003

[21] Maiato H, DeLuca J, Salmon ED, et al. The dynamic kinetochore-microtubule interface[J]. J Cell Sci, 2004, 117(Pt 23): 5461-5477.

[22] Lee H. How chromosome mis-segregation leads to cancer: lessons from BubR1 mouse models[J]. Mol Cells, 2014, 37(10): 713-718. doi: 10.14348/molcells.2014.0233

[23] Meraldi P, Honda R, Nigg EA. Aurora kinases link chromosome segregation and cell division to cancer susceptibility[J]. Curr Opin Genet Dev, 2004, 14(1): 29-36. doi: 10.1016/j.gde.2003.11.006

[24] McNally FJ, Vale RD. Identification of katanin, an ATPase that severs and disassembles stable microtubules[J]. Cell, 1993, 75(3): 419-429. doi: 10.1016/0092-8674(93)90377-3

[25] Hartman JJ, Mahr J, McNally K, et al. Katanin, a microtubule-severing protein, is a novel AAA ATPase that targets to the centrosome using a WD40-containing subunit[J]. Cell, 1998, 93(2): 277-287. doi: 10.1016/S0092-8674(00)81578-0

[26] Buster D, McNally K, McNally FJ. Katanin inhibition prevents the redistribution of γ-tubulin at mitosis[J]. J Cell Sci, 2002, 115(Pt 5): 1083-1092.

[27] Yang HY, McNally K, McNally FJ. MEI-1/katanin is required for translocation of the meiosis I spindle to the oocyte cortex in C. elegans[J]. Dev Biol, 2003, 260(1): 245-259. doi: 10.1016/S0012-1606(03)00216-1

[28] Loughlin R, Wilbur JD, McNally FJ, et al. Katanin contributes to interspecies spindle length scaling in Xenopus[J]. Cell, 2011, 147(6): 1397-1407. doi: 10.1016/j.cell.2011.11.014

[29] Hazan J, Fonknechten N, Mavel D, et al. Spastin, a new AAA protein, is altered in the most frequent form of autosomal dominant spastic paraplegia[J]. Nat Genet, 1999, 23(3): 296-303. doi: 10.1038/15472

[30] Breslow DK, Holland AJ. Mechanism and regulation of centriole and cilium biogenesis[J]. Annu Rev Biochem, 2019, 88: 691-724. doi: 10.1146/annurev-biochem-013118-111153

[31] Errico A, Claudiani P, D'Addio M, et al. Spastin interacts with the centrosomal protein NA14, and is enriched in the spindle pole, the midbody and the distal axon[J]. Hum Mol Genet, 2004, 13(18): 2121-2132. doi: 10.1093/hmg/ddh223

[32] Zhang D, Rogers GC, Buster DW, et al. Three microtubule severing enzymes contribute to the "Pacman-flux" machinery that moves chromosomes[J]. J Cell Biol, 2007, 177(2): 231-242. doi: 10.1083/jcb.200612011

[33] Luke-Glaser S, Pintard L, Tyers M, et al. The AAA-ATPase FIGL-1 controls mitotic progression, and its levels are regulated by the CUL-3^MEL-26 E3 ligase in the C. elegans germ line[J]. J Cell Sci, 2007, 120(Pt 18): 3179-3187.

[34] Sonbuchner TM, Rath U, Sharp DJ. KL1 is a novel microtubule severing enzyme that regulates mitotic spindle architecture[J]. Cell Cycle, 2010, 9(12): 2403-2411. doi: 10.4161/cc.9.12.11916

[35] Ververis A, Christodoulou A, Christoforou M, et al. A novel family of katanin-like 2 protein isoforms (KATNAL2), interacting with nucleotide-binding proteins Nubp1 and Nubp2, are key regulators of different MT-based processes in mammalian cells[J]. Cell Mol Life Sci, 2016, 73(1): 163-184. doi: 10.1007/s00018-015-1980-5

[36] Filges I, Manokhina I, Peñaherrera MS, et al. Recurrent triploidy due to a failure to complete maternal meiosis II: whole-exome sequencing reveals candidate variants[J]. Mol Hum Reprod, 2015, 21(4): 339-346. doi: 10.1093/molehr/gau112

[37] Feng R, Sang Q, Kuang Y, et al. Mutations in TUBB8 and human oocyte meiotic arrest[J]. N Engl J Med, 2016, 374(3): 223-232. doi: 10.1056/NEJMoa1510791

[38] Kogo H, Kowa-Sugiyama H, Yamada K, et al. Screening of genes involved in chromosome segregation during meiosis I: toward the identification of genes responsible for infertility in humans[J]. J Hum Genet, 2010, 55(5): 293-299. doi: 10.1038/jhg.2010.26

[39] McNally F J, Roll-Mecak A. Microtubule-severing enzymes: From cellular functions to molecular mechanism[J]. J Cell Biol, 2018, 17(12): 4057-4069. https://rupress.org/jcb/article/217/12/4057/120287/Microtubule-severing-enzymes-From-cellular

[40] Balboula AZ, Schindler K. Selective disruption of aurora C kinase reveals distinct functions from aurora B kinase during meiosis in mouse oocytes[J]. PLoS Genet, 2014, 10(2): e1004194. doi: 10.1371/journal.pgen.1004194

[41] Zhang D, Grode KD, Stewman SF, et al. Drosophila katanin is a microtubule depolymerase that regulates cortical-microtubule plus-end interactions and cell migration[J]. Nat Cell Biol, 2011, 13(4): 361-369. doi: 10.1038/ncb2206

[42] Srayko M, O'Toole ET, Hyman AA, et al. Katanin disrupts the microtubule lattice and increases polymer number in C. elegans meiosis[J]. Curr Biol, 2006, 16(19): 1944-1949. doi: 10.1016/j.cub.2006.08.029

[43] Ems-McClung SC, Hainline SG, Devare J, et al. Aurora B inhibits MCAK activity through a phosphoconformational switch that reduces microtubule association[J]. Curr Biol, 2013, 23(24): 2491-2499. doi: 10.1016/j.cub.2013.10.054

[44] Ohkura H. Meiosis: an overview of key differences from mitosis[J]. Cold Spring Harb Perspect Biol, 2015, 7(5): a015859. doi: 10.1101/cshperspect.a015859

[45] Li X, Bloomfield M, Bridgeland A, et al. A fine balance among key biophysical factors is required for recovery of bipolar mitotic spindle from monopolar and multipolar abnormalities[J]. Mol Biol Cell, 2023, 34(9): ar90. doi: 10.1091/mbc.E22-10-0485

[46] Rogers GC, Rogers SL, Schwimmer TA, et al. Two mitotic kinesins cooperate to drive sister chromatid separation during anaphase[J]. Nature, 2004, 427(6972): 364-370. doi: 10.1038/nature02256

[47] McNally K, Berg E, Cortes DB, et al. Katanin maintains meiotic metaphase chromosome alignment and spindle structure in vivo and has multiple effects on microtubules in vitro[J]. Mol Biol Cell, 2014, 25(7): 1037-1049. doi: 10.1091/mbc.e13-12-0764

[48] McNally K, Audhya A, Oegema K, et al. Katanin controls mitotic and meiotic spindle length[J]. J Cell Biol, 2006, 175(6): 881-891. doi: 10.1083/jcb.200608117

[49] Srayko M, Buster DW, Bazirgan OA, et al. MEI-1/MEI-2 katanin-like microtubule severing activity is required for Caenorhabditis elegans meiosis[J]. Genes Dev, 2000, 14(9): 1072-1084. doi: 10.1101/gad.14.9.1072

[50] Moore CEJ, Wang X, Xie J, et al. Elongation factor 2 kinase promotes cell survival by inhibiting protein synthesis without inducing autophagy[J]. Cell Signal, 2016, 28(4): 284-293. doi: 10.1016/j.cellsig.2016.01.005

[51] Piserchio A, Isiorho EA, Long K, et al. Structural basis for the calmodulin-mediated activation of eukaryotic elongation factor 2 kinase[J]. Sci Adv, 2022, 8(27): eabo2039. doi: 10.1126/sciadv.abo2039

[52] Smith EM, Proud CG. cdc2-cyclin B regulates eEF2 kinase activity in a cell cycle- and amino acid-dependent manner[J]. EMBO J, 2008, 27(7): 1005-1016. doi: 10.1038/emboj.2008.39

[53] Gomes JE, Tavernier N, Richaudeau B, et al. Microtubule severing by the katanin complex is activated by PPFR-1-dependent MEI-1 dephosphorylation[J]. J Cell Biol, 2013, 202(3): 431-439. doi: 10.1083/jcb.201304174

[54] Whitehead E, Heald R, Wilbur JD. N-terminal phosphorylation of p60 katanin directly regulates microtubule severing[J]. J Mol Biol, 2013, 425(2): 214-221. doi: 10.1016/j.jmb.2012.11.022

[55] Wang R, He G, Nelman-Gonzalez M, et al. Regulation of Cdc25C by ERK-MAP kinases during the G₂/M transition[J]. Cell, 2007, 128(6): 1119-1132. doi: 10.1016/j.cell.2006.11.053