Mechanobiology of the Mitotic Spindle

The mitotic spindle is a microtubule-based assembly that separates the chromosomes during cell division. As basically mechanical micro machine, understanding of its functioning constantly motivating development experimental approaches based on perturbations, which are complementary to and work together with classical genetics biochemistry methods. Recent data emerging from these in combination theoretical modeling led novel ideas significant revisions basic concepts field. In this Perspective, we discuss advances mechanics, focusing microtubule forces control chromosome movements. Segregation genome mother into two daughter cells division one fundamental processes life. Physical separation opposite poles carried out by (Figure 1), fascinating complex machine made microtubules numerous other proteins (McIntosh et al., 2012McIntosh J.R. Molodtsov M.I. Ataullakhanov F.I. Biophysics mitosis.Q. Rev. Biophys. 2012; 45: 147-207Crossref PubMed Scopus (71) Google Scholar; Pavin Toli?, 2016Pavin N. Toli? I.M. Self-organization spindle.Annu. 2016; 279-298Crossref Prosser Pelletier, 2017Prosser S.L. Pelletier L. Mitotic animal cells: fine balancing act.Nat. Mol. Cell Biol. 2017; 18: 187-201Crossref (131) Scholar). principal job segregate without mistakes. However, sometimes errors segregation occur they can cause aneuploidy, state characterized wrong number cell. This type aberration found almost all human cancers common miscarriages genetic disorders, such as Down syndrome (Santaguida Amon, 2015Santaguida S. Amon A. Short- long-term effects mis-segregation aneuploidy.Nat. 2015; 16: 473-485Crossref Knouse 2017Knouse K.A. Davoli T. Elledge S.J. Aneuploidy cancer: Seq-ing answers old questions.Annu. Cancer 1: 335-354Crossref Webster Schuh, 2017Webster Schuh M. Mechanisms aneuploidy eggs.Trends 27: 55-68Abstract Full Text PDF (61) Thus, studying not only important research understand how functions but also better causes serious diseases. Spindles have been studied for more than 150 years Hays, 2016McIntosh Hays A brief history mechanisms.Biology (Basel). 5: 55Google it still clear orchestrates chromosomes. Why biologically crucial understood? Several thousands studies used various molecular biology provided list hundreds involved proteins, including tens motor walk along microtubules, well massive amount spindle. But always easy make coherent picture because main players interactions. Some perform different at locations. For example, Eg5 kinesin-5 family slide apart center bundle (Mann Wadsworth, 2018bMann B.J. Wadsworth P. Kinesin-5 regulation function mitosis.Trends 2018; 29: 66-79Abstract (14) An additional complication arises when same function, Kif15 kinesin-12 both (Tanenbaum 2009Tanenbaum M.E. Mac?rek Janssen Geers E.F. Alvarez-Fernández Medema R.H. cooperates eg5 promote bipolar assembly.Curr. 2009; 19: 1703-1711Abstract (171) These examples show why identify role each protein interact perspective article, mechanobiology mainly mammalian cells, putting context methods experimentally dissect advantages combining them approaches, focus movements positioning. complexity variety biochemistry. essentially requires perturbations. One most fruitful tools has laser ablation, allows cutting direct identification direction movement fragments 2A). rationale if moved toward other, was under compression before cut 2A, left), whereas signature tension middle). If rotate, implies rotational were present right). be complex, linear reasoning holds any material study acting kinetochore, linking microtubules. pioneering study, kinetochore ablated laser, resulted sister associated pole, demonstrating inter-kinetochore (McNeill Berns, 1981McNeill P.A. Berns M.W. Chromosome behavior after microirradiation single PtK2 cells.J. 1981; 88: 543-553Crossref Since then, ablation works that, explored mechanics kinetochores, centrosomes (Aist 1993Aist Liang H. Astral anaphase B: microbeam study.J. Sci. 1993; 104: 1207-1216Crossref Skibbens 1995Skibbens R.V. Rieder C.L. Salmon E.D. Kinetochore motility severing between centromeres using microsurgery: evidence directional instability position regulated tension.J. 1995; 108: 2537-2548PubMed Khodjakov Rieder, 1996Khodjakov Kinetochores moving away their pole do exert pushing force chromosome.J. 1996; 135: 315-327Crossref (98) 2000Khodjakov Cole R.W. Oakley B.R. Centrosome-independent formation vertebrates.Curr. 2000; 10: 59-67Abstract Maiato 2004Maiato Kinetochore-driven fibers contributes mitosis.J. 2004; 167: 831-840Crossref (211) Dick Gerlich, 2013Dick A.E. Gerlich D.W. Kinetic framework checkpoint signalling.Nat. 2013; 15: 1370-1377Crossref (102) Elting 2014Elting Hueschen Udy D.B. Dumont Force minus ends moves chromosomes.J. 2014; 206: 245-256Crossref (59) Sikirzhytski 2014Sikirzhytski V. Magidson Steinman J.B. He J. Le Berre Tikhonenko I. Ault J.G. McEwen B.F. Chen J.K. Sui al.Direct kinetochore-spindle connections required segregation.J. 231-243Crossref (58) Cojoc 2016Cojoc G. Roscioli E. Zhang García-Ulloa Shah J.V. Cimini D. Gregan Laser microsurgery reveals conserved viscoelastic kinetochore.J. 212: 767-776Crossref (15) Kajtez 2016Kajtez Solomatina Novak Polak B. Vukuši? K. Rüdiger Milas Šumanovac Šestak Risteski al.Overlap link k-fibres balance bi-oriented kinetochores.Nat. Commun. 7: 10298Crossref (48) 2017Vukuši? Bu?a R. Bosilj Microtubule sliding within bridging fiber pushes chromosomes.Dev. Cell. 43: 11-23.e6Abstract (27) Whereas perturbs existing spindle, techniques allow applying external groundbreaking studies, Bruce Nicklas microneedle stretch, move, or halt measure 2B) (Nicklas Staehly, 1967Nicklas R.B. Staehly C.A. micromanipulation. attachment spindle.Chromosoma. 1967; 21: 1-16Crossref (104) Nicklas, 1983Nicklas Measurements produced anaphase.J. 1983; 97: 542-548Crossref External applied (Takagi 2019Takagi Sakamoto Shiratsuchi Maeda Y.T. Shimamoto Y. Mechanically distinct arrays determine length response meiotic spindle.Dev. 2019; 49: 267-278.e5Abstract (7) Scholar) squeezing whole (Dumont Mitchison, 2009aDumont Mitchison T.J. Compression regulates mechanochemical switch poles.Curr. 1086-1095Abstract (70) Itabashi 2012Itabashi Terada Kuwana Kan Shimoyama Ishiwata S.a.i. Mechanical impulses metaphase progression cell.Proc. Natl. Acad. USA. 109: 7320-7325Crossref (26) change throughout properties 2C). Optical magnetic tweezers displace providing insight coming cortex 2D) (Toli?-Nørrelykke 2005Toli?-Nørrelykke Sacconi Stringari C. Raabe Pavone F.S. Nuclear division-plane positioning revealed optical micromanipulation.Curr. 2005; 1212-1216Abstract (0) Garzon-Coral 2016Garzon-Coral Fantana H.A. Howard force-generating machinery maintains mitosis.Science. 352: 1124-1127Crossref combined light microscopy see structure will perturbed outcome experiment. use fluorescently labeled helps architecture, localization over time perturbation. developments super-resolution open possibilities explore detailed (Sahl 2017Sahl Hell S.W. Jakobs Fluorescence nanoscopy biology.Nat. 685-701Crossref (325) gold standard where identified, electron microscopy, unfortunately cannot live cells. All perturbations reveal aspects working complete mechanobiology. Experiments typically provide movies contain ample information about dynamics. To key functioning, find patterns propose hypotheses. first step, hypotheses expressed cartoon models. models tested quantitative. situations, theory great tool becoming increasingly biology, general. Designing model us formulate hypotheses, solving verifying concepts. general, simplified view real system, thus, describe features, becomes powerful describes (Mogilner 2006Mogilner Wollman Marshall W.F. Quantitative biology: what good for?.Dev. 2006; 11: 279-287Abstract (86) successful should reproduce system pass tests exciting predictions would happen new experiments otherwise an obvious choice. example kinetochores captured part (Wollman 2005Wollman Cytrynbaum E.N. Jones J.T. Meyer Scholey J.M. Mogilner Efficient capture bias ‘search-and-capture’ process mitotic-spindle 828-832Abstract (187) Paul 2009Paul Silkworth W.T. Nardi I.K. Computer simulations predict rotations accelerate compromising accuracy.Proc. 106: 15708-15713Crossref (63) 2015Magidson Yang O'Connell C.B. Adaptive changes architecture facilitate proper assembly.Nat. 17: 1134-1144Crossref prevailing search-and-capture mechanism (Kirschner 1986Kirschner Beyond self-assembly: morphogenesis.Cell. 1986; 329-342Abstract (904) revised, significantly longer observed computational predicted enlarging speed up substantially, inspired (Magidson Similarly, interpolar bundles (Nédélec, 2002Nédélec F. generating stable antiparallel interactions.J. 2002; 158: 1005-1015Crossref (150) Scholar), another aspect assembly, initiated body (Dinarina 2009Dinarina Pugieux Corral M.M. Loose Spatz Karsenti Nédélec Chromatin shapes spindle.Cell. 138: 502-513Abstract Loughlin 2010Loughlin Heald predicts Xenopus organization.J. 2010; 191: 1239-1249Crossref (89) Things become even interesting limited scope scientific question motivated design theory, rather bigger picture. power comes 19th century physics James Clerk Maxwell developed electromagnetism, he existence electromagnetic waves velocity. Surprisingly, velocity light, him wave. We expect future surprising especially large relies self-regulating mechanisms length. slow down chemical reactions. affects polymerization, depends difference rates addition removal subunits. growing tip encounters obstacle decreases rate gap needed subunit, appears fluctuates occurs less frequently, resulting slowdown polymerization (Dogterom Yurke, 1997Dogterom Yurke Measurement force-velocity relation microtubules.Science. 1997; 278: 856-860Crossref (357) depolymerization generate move cargo attached plus end (Lombillo 1995Lombillo V.A. Stewart R.J. McIntosh Minus-end-directed motion kinesin-coated microspheres driven depolymerization.Nature. 373: 161-164Crossref Grishchuk 2005Grishchuk E.L. production disassembling microtubules.Nature. 438: 384-388Crossref (189) dynamics detachment cargo, (Akiyoshi 2010Akiyoshi Sarangapani K.K. Powers A.F. Nelson C.R. Reichow Arellano-Santoyo Gonen Ranish J.A. Asbury Biggins Tension directly stabilizes reconstituted kinetochore-microtubule attachments.Nature. 468: 576-579Crossref (251) contrast bonds, increased dissociation (Kramers, 1940Kramers Brownian field diffusion reactions.Physica. 1940; 284-304Crossref (6610) Howard, 2009Howard signaling networks cytoskeletal proteins.Annu. 38: 217-234Crossref (66) depolymerizing 3A) resembling catch-bond (Sarangapani Asbury, 2014Sarangapani Catch release: hook right mitosis?.Trends Genet. 30: 150-159Abstract Length needs regulated. length-dependent accumulation end, regulate 3B). Motors, kinesin-8 particular, bind entire microtubule, collects motors. motors processively reach detachment, accumulate known antenna (Varga 2006Varga Helenius Tanaka Hyman A.A. T.U. Yeast depolymerizes manner.Nat. 8: 957-962Crossref (298) Scholar, Varga 2009Varga Leduc Bormuth Diez Kinesin-8 act cooperatively mediate depolymerization.Cell. 1174-1183Abstract (192) Once induce catastrophe (Tischer 2009Tischer Brunner Dogterom Force- kinesin-8-dependent spatial fission yeast dynamics.Mol. Syst. 250Crossref (74) limiting promotes (Stumpff 2008Stumpff von Dassow Wagenbach Wordeman Kif18A suppresses alignment.Dev. 2008; 14: 252-262Abstract (202) Klemm 2018Klemm A.H. Glun?i? Metaphase dynamic instability.Mol. 1332-1345Crossref (6) Similar individual overlap midzone kinesin-4, depolymerase recruited crosslinker PRC1 manner, suggested vitro (Bieling 2010Bieling Telley I.A. Surrey minimal module controls overlaps.Cell. 142: 420-432Abstract Within overlap, Ase1/PRC1 prevents completely thereby presumably entropic generated molecules diffusing (Braun 2011Braun Lansky Z. Fink Ruhnow Janson braking Ase1 overlapping apart.Nat. 2011; 13: 1259-1264Crossref (69) 2015Lansky Braun Lüdecke Schlierf Ten Wolde P.R. Diffusible crosslinkers directed networks.Cell. 160: 1159-1168Abstract (75) crosslinkers, overlaps respect other. depend shown generates higher (Shimamoto 2015Shimamoto Forth Kapoor T.M. Measuring ensembles crosslinking microtubules.Dev. 34: 669-681Abstract (57) summary, determines self-organizes ultimately segregates 2009b; Oriola 2018Oriola Needleman D.J. Brugués 47: 655-673Crossref life starts interactions mediated microtubule-associated prometaphase anaphase, followed disassembly telophase. gain phases mitosis belongs cell, phase already big challenge. steady state, makes responses simpler interpret anaphase. defined eye-catching alignment near equatorial plane questions keep close equator. forming fiber, pulls demonstrated 4, point 1) At polymerize, discovered microinjecting biotinylated tubulin sites growth 2) (Mitchison 1986Mitchison Evans Schulze Kirschner Sites 515-527Abstract continuously translocate poleward flux, visible photoactivated segments 3) (Mitchison, 1989Mitchison Polewards flux spindle: photoactivation fluorescence.J. 1989; 637-652Crossref constant simultaneously polymerize depolymerize drawing). pulling opposed elastic stretched chromatin (Pickett-Heaps 1982Pickett-Heaps J.D. Tippit D.H. Porter K.R. Rethinking mitosis.Cell. 1982; 729-744Abstract finding supported measurements showing bioriented further nonattached ones 4) (Waters 1996Waters J.C. Oscillating newt lung are, average, rarely push.J. 2823-2831Crossref time, polar ejection forces, exerted arms transmitted 5) (Rieder 1986Rieder Davison E.A. Jensen L.C. Cassimeris Oscillatory monooriented relative result aster half-spindle.J. 103: 581-591Crossref pull non-kinetochore push Salmon, 1994Rieder Motile dictate vertebrate spindle.J. 1994; 124: 223-233Crossref consequences concepts, introduced. early explain congression (Khodjakov 1999Khodjakov Gabashvili I.S. Dumb" versus "smart" somatic cells.Cell Motil. Cytoskeleton. 1999; 179-185Crossref later elegant (Joglekar Hunt, 2002J