human preimplantation development.Cell Stem Cell 25 ,
697 – 712.e6 (2019). doi:10.1016/j.stem.2019.09.004;
pmid: 31588047- Q. Konget al., Lineage specification and pluripotency
revealed by transcriptome analysis from oocyte to blastocyst
in pig.FASEB J. 34 , 691–705 (2020). doi:10.1096/
fj.201901818RR; pmid: 31914626 - R. Yanet al., Decoding dynamic epigenetic landscapes in
human oocytes using single-cell multi-omics sequencing.
Cell Stem Cell 28 , 1641–1656.e7 (2021). doi:10.1016/
j.stem.2021.04.012; pmid: 33957080 - I. Bennabiet al., Shifting meiotic to mitotic spindle assembly
in oocytes disrupts chromosome alignment.EMBO Rep. 19 ,
368 – 381 (2018). doi:10.15252/embr.201745225;
pmid: 29330318 - V. Mountainet al., The kinesin-related protein, HSET,
opposes the activity of Eg5 and cross-links microtubules in
the mammalian mitotic spindle.J. Cell Biol. 147 , 351– 366
(1999). doi:10.1083/jcb.147.2.351; pmid: 10525540 - Y. Caoet al., Microtubule minus-end binding protein
CAMSAP2 and kinesin-14 motor KIFC3 control dendritic
microtubule organization.Curr. Biol. 30 , 899–908.e6 (2020).
doi:10.1016/j.cub.2019.12.056; pmid: 32084403 - Z. Y. She, W. X. Yang, Molecular mechanisms of kinesin-14
motors in spindle assembly and chromosome segregation.
J. Cell Sci. 130 , 2097–2110 (2017). doi:10.1242/jcs.200261;
pmid: 28668932 - S. Cai, L. N. Weaver, S. C. Ems-McClung, C. E. Walczak,
Kinesin-14 family proteins HSET/XCTK2 control spindle
length by cross-linking and sliding microtubules.Mol. Biol.
Cell 20 , 1348–1359 (2009). doi:10.1091/mbc.e08-09-0971;
pmid: 19116309 - C. E. Walczak, S. Verma, T. J. Mitchison, XCTK2: A kinesin-
related protein that promotes mitotic spindle assembly in
Xenopus laevisegg extracts.J. Cell Biol. 136 , 859– 870
(1997). doi:10.1083/jcb.136.4.859; pmid: 9049251 - T. J. Mullen, S. M. Wignall, Interplay between microtubule
bundling and sorting factors ensures acentriolar spindle
stability duringC. elegansoocyte meiosis.PLOS Genet. 13 ,
e1006986 (2017). doi:10.1371/journal.pgen.1006986;
pmid: 28910277 - C. H. Chuang, A. J. Schlientz, J. Yang, B. Bowerman,
Microtubule assembly and pole coalescence: Early steps in
Caenorhabditis elegansoocyte meiosis I spindle assembly.
Biol. Open 9 , bio.052308 (2020). doi:10.1242/bio.052308;
pmid: 32493729 - C. E. Walczak, I. Vernos, T. J. Mitchison, E. Karsenti, R. Heald,
A model for the proposed roles of different microtubule-
based motor proteins in establishing spindle bipolarity.
Curr. Biol. 8 , 903–913 (1998). doi:10.1016/S0960-9822(07)
00370-3; pmid: 9707401 - S. Morales-Mulia, J. M. Scholey, Spindle pole organization in
DrosophilaS2 cells by dynein, abnormal spindle protein
(Asp), and KLP10A.Mol. Biol. Cell 16 , 3176–3186 (2005).
doi:10.1091/mbc.e04-12-1110; pmid: 15888542 - G. Goshima, F. Nédélec, R. D. Vale, Mechanisms for focusing
mitotic spindle poles by minus end-directed motor proteins.
J. Cell Biol. 171 , 229–240 (2005). doi:10.1083/
jcb.200505107; pmid: 16247025 - J. Baumbach, Z. A. Novak, J. W. Raff, A. Wainman, Dissecting
the function and assembly of acentriolar microtubule
organizing centers inDrosophilacells in vivo.PLOS Genet. 11 ,
e1005261 (2015). doi:10.1371/journal.pgen.1005261;
pmid: 26020779 - M. Kwonet al., Mechanisms to suppress multipolar divisions
in cancer cells with extra centrosomes.Genes Dev. 22 ,
2189 – 2203 (2008). doi:10.1101/gad.1700908;
pmid: 18662975 - J. Kleylein-Sohnet al., Acentrosomal spindle organization
renders cancer cells dependent on the kinesin HSET.J. Cell
Sci. 125 , 5391–5402 (2012). doi:10.1242/jcs.107474;
pmid: 22946058 - N. Kim, K. Song, KIFC1 is essential for bipolar spindle
formation and genomic stability in the primary human
fibroblast IMR-90 cell.Cell Struct. Funct. 38 , 21–30 (2013).
doi:10.1247/csf.12014; pmid: 23318213
118. P. L. Chavaliet al., A CEP215-HSET complex links
centrosomes with spindle poles and drives centrosome
clustering in cancer.Nat. Commun. 7 , 11005 (2016).
doi:10.1038/ncomms11005; pmid: 26987684
119. B. Vitreet al., IFT proteins interact with HSET to promote
supernumerary centrosome clustering in mitosis.EMBO Rep.
21 , e49234 (2020). doi:10.15252/embr.201949234;
pmid: 32270908
120. E. A. Nigg, Centrosome aberrations: Cause or consequence of
cancer progression?Nat. Rev. Cancer 2 , 815–825 (2002).
doi:10.1038/nrc924; pmid: 12415252
121. J. Fu, I. M. Hagan, D. M. Glover, The centrosome and its
duplication cycle.Cold Spring Harb. Perspect. Biol. 7 ,
a015800 (2015). doi:10.1101/cshperspect.a015800;
pmid: 25646378
122. T. Yaoet al., Live-cell imaging of nuclear-chromosomal
dynamics in bovine in vitro fertilised embryos.Sci. Rep. 8 ,
7460 (2018). doi:10.1038/s41598-018-25698-w;
pmid: 29748644
123. T. Cavazzaet al., Parental genome unification is highly error-
prone in mammalian embryos.Cell 184 , 2860–2877.e22
(2021). doi:10.1016/j.cell.2021.04.013; pmid: 33964210
124. I. Schneider, M. de Ruijter-Villani, M. J. Hossain, T. A. E. Stout,
J. Ellenberg, Dual spindles assemble in bovine zygotes
despite the presence of paternal centrosomes.J. Cell Biol.
220 , e202010106 (2021). doi:10.1083/jcb.202010106;
pmid: 34550316
125. A. I. Mihajlović, J. Haverfield, G. FitzHarris, Distinct classes
of lagging chromosome underpin age-related oocyte
aneuploidy in mouse.Dev. Cell 56 , 2273–2283.e3 (2021).
doi:10.1016/j.devcel.2021.07.022; pmid: 34428397
126. J. Roeles, G. Tsiavaliaris, Actin-microtubule interplay
coordinates spindle assembly in human oocytes.Nat.
Commun. 10 , 4651 (2019). doi:10.1038/s41467-019-12674-9;
pmid: 31604948
127. B. T. Bajaret al., Improving brightness and photostability of
green and red fluorescent proteins for live cell imaging and
FRET reporting.Sci. Rep. 6 , 20889 (2016). doi:10.1038/
srep20889; pmid: 26879144
128. G. H. Patterson, J. Lippincott-Schwartz, A photoactivatable
GFP for selective photolabeling of proteins and cells.
Science 297 , 1873–1877 (2002). doi:10.1126/science.1074952;
pmid: 12228718
129. D. S. Bindelset al., mScarlet: A bright monomeric red
fluorescent protein for cellular imaging.Nat. Methods 14 ,
53 – 56 (2017). doi:10.1038/nmeth.4074; pmid: 27869816
130. E. R. Liman, J. Tytgat, P. Hess, Subunit stoichiometry of a
mammalian K+channel determined by construction of
multimeric cDNAs.Neuron 9 , 861–871 (1992). doi:10.1016/
0896-6273(92)90239-A; pmid: 1419000
131. Q. Zhanget al., Nudel promotes axonal lysosome clearance
and endo-lysosome formation via dynein-mediated transport.
Traffic 10 , 1337–1349 (2009). doi:10.1111/j.1600-
0854.2009.00945.x; pmid: 19522757
132. S. Pfender, V. Kuznetsov, S. Pleiser, E. Kerkhoff, M. Schuh,
Spire-type actin nucleators cooperate with Formin-2 to drive
asymmetric oocyte division.Curr. Biol. 21 , 955–960 (2011).
doi:10.1016/j.cub.2011.04.029; pmid: 21620703
133. D.Clift,C.So,W.A.McEwan,L.C.James,M.Schuh,Acute
and rapid degradation of endogenous proteins by Trim-
Away.Nat. Protoc. 13 , 2149–2175 (2018). doi:10.1038/
s41596-018-0028-3; pmid: 30250286
134. S. Hua, K. Jiang, Expression and purification of microtubule-
associated proteins from HEK293T cells for in vitro
reconstitution.Methods Mol. Biol. 2101 , 19–26 (2020).
doi:10.1007/978-1-0716-0219-5_2; pmid: 31879895
135. J. Bucevičius, G. Kostiuk, R. Gerasimaitė, T. Gilat,
G. Lukinavičius, Enhancing the biocompatibility of rhodamine
fluorescent probes by a neighbouring group effect.Chem.
Sci. 11 , 7313–7323 (2020). doi:10.1039/D0SC02154G;
pmid: 33777348
136. N. Tanaka, W. Meng, S. Nagae, M. Takeichi, Nezha/CAMSAP3
and CAMSAP2 cooperate in epithelial-specific organization
of noncentrosomal microtubules.Proc. Natl. Acad. Sci. U.S.A.
109 , 20029–20034 (2012). doi:10.1073/pnas.1218017109;
pmid: 23169647
137. A. Z. Politiet al., Quantitative mapping of fluorescently
tagged cellular proteins using FCS-calibrated four-
dimensional imaging.Nat. Protoc. 13 , 1445–1464 (2018).
doi:10.1038/nprot.2018.040; pmid: 29844523
138. N. L. Schieberet al., Minimal resin embedding of multicellular
specimens for targeted FIB-SEM imaging.Methods Cell Biol.
140 , 69–83 (2017). doi:10.1016/bs.mcb.2017.03.005;
pmid: 28528642
139. B. Li, C. N. Dewey, RSEM: Accurate transcript quantification
from RNA-Seq data with or without a reference genome.
BMC Bioinformatics 12 , 323 (2011). doi:10.1186/
1471-2105-12-323; pmid: 21816040
140. G. P. Wagner, K. Kin, V. J. Lynch, Measurement of mRNA
abundance using RNA-seq data: RPKM measure is
inconsistent among samples.Theory Biosci. 131 , 281– 285
(2012). doi:10.1007/s12064-012-0162-3; pmid: 22872506
ACKNOWLEDGMENTS
We are grateful to the patients who participated in this study. We
thank the staff from the Animal Facility and Live-Cell Imaging
Facility at the Max Planck Institute for Multidisciplinary Sciences for
technical assistance; the clinicians, nursing team, and embryology
team at the clinics for their support of this study; L. Abdelhalim,
E. Bellou, and L. Wartosch for help with human oocytes; C. Mauksch
for help with optimizing thawing of vitrified human oocytes;
E. Bellou, T. Cavazza, and M. Daniel for help with bovine and
porcine ovaries; T. Ruhwedel for help with sample preparation for
electron microscopy; E. Bellou, A. Politi, and F. Xie for helpful
discussions; A. Andersen, T. Cavazza, P. Lénárt, and L. Wartosch for
critical comments on the manuscript; T. Hiiragi, the M. J. Fox Foundation,
M. Mancini, and X. Zhu for cDNAs and constructs; and D. A. Compton,
E. Nigg, G. Goshima, P. Meraldi, A. McAinsh, M. Takeichi,
and R. Uehara for antibodies.Funding:The research leading to
these results was funded by the Max Planck Society and the DFG
under a Leibniz Prize to M.S. (SCHU 3047/1-1) and a grant to
W.M. [MO 1084/2-1 (FOR2848, P8)]. C.So is a recipient of the
Max Planck Croucher Postdoctoral Fellowship. W.M. and M.S. were
supported by the Deutsche Forschungsgemeinschaft (DFG,
German Research Foundation) under Germany’s Excellence
Strategy–EXC 2067/1-390729940.Author contributions:C.So
and M.S. conceived the study, designed experiments and methods
for data analysis; C.So performed all experiments and analyzed
the data with the following exceptions: K.M. performed Trim-Away
of NUMA in human oocytes; J.U. performed inhibition with
P150-CC1 in human oocytes; K.H. performed live imaging of
porcine oocytes and optimized live imaging of bovine oocytes;
A.M.S. prepared electron microscopy samples with C.So and
performed FIB-SEM; K.B.S. optimized Trim-Away of NUMA and
inhibition with P150-CC1 in human oocytes; and J.B. and G.L.
synthesized 5-SiR-CTX and 5-SiR-Hoechst. C.So and M.S. wrote the
manuscript and prepared the figures with input from all authors;
W.M. supervised the electron microscopy experiments; C.Si. and
A.T.-S. supervised the collection and vitrification of human oocytes
at Fertility Center Berlin; H.E. and R.M. supervised the collection
of human oocytes at Kinderwunschzentrum Göttingen; M.B. and
K.E. supervised the collection of human oocytes at Bourn Hall
Clinic; and M.S. supervised the entire study.Competing interests:
C.So and M.S. filed a patent application (EP21199120.3) based on
data presented here. The other authors declare no competing
financial interests.Data and materials availability:Plasmids are
available from M.S. under a material transfer agreement with the Max
Planck Society. All data needed to evaluate the conclusions in the
paper are present in the main text or the supplementary materials.SUPPLEMENTARY MATERIALS
science.org/doi/10.1126/science.abj3944
Figs. S1 to S14
Tables S1 and S2
MDAR Reproducibility Checklist
Movies S1 to S1810 May 2021; resubmitted 2 November 2021
Accepted 11 January 2022
10.1126/science.abj3944Soet al.,Science 375 , eabj3944 (2022) 11 February 2022 19 of 19
RESEARCH | RESEARCH ARTICLE