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EB1 Is Essential for Spindle Formation and Chromosome Alignment During Oocyte Meiotic Maturation in Mice

Published online by Cambridge University Press:  08 January 2021

Dongjie Zhou Open the ORCID record for Dongjie Zhou [Opens in a new window] ,
Zheng-Wen Nie  and
Xiang-Shun Cui
Dongjie Zhou
Affiliation:
Department of Animal Science, Chungbuk National University, 356 Room, S21-5 Dong, Chungdae-ro 1, Seowon-Gu, Cheongju, Chungbuk28644, South Korea
Zheng-Wen Nie
Affiliation:
Department of Animal Science, Chungbuk National University, 356 Room, S21-5 Dong, Chungdae-ro 1, Seowon-Gu, Cheongju, Chungbuk28644, South Korea
Xiang-Shun Cui*
Affiliation:
Department of Animal Science, Chungbuk National University, 356 Room, S21-5 Dong, Chungdae-ro 1, Seowon-Gu, Cheongju, Chungbuk28644, South Korea
*
*Author for correspondence: Xiang-Shun Cui, E-mail: [email protected]
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Abstract

The cytoskeleton plays an orchestrating role in polarized cell growth. Microtubules (MTs) not only play critical roles in chromosome alignment and segregation but also control cell shape, division, and motility. A member of the plus-end tracking proteins, end-binding protein 1 (EB1), regulates MT dynamics and plays vital roles in maintaining spindle symmetry and chromosome alignment during mitosis. However, the role of EB1 in mouse oocyte meiosis remains unknown. Here, we examined the localization patterns and expression levels of EB1 at different stages. EB1 protein level was found to be stable during meiosis. EB1 mainly localized along the spindle and had a similar localization pattern as that of α-tubulin. The EB1 protein was degraded with a Trim-Away method, and the results were further confirmed with western blotting and immunofluorescence. At 12 h of culture after EB1 knockdown (KD), a reduced number of mature MII oocytes were observed. EB1 KD led to spindle disorganization, chromosome misalignment, and missegregation; β-catenin protein binds to actin via the adherens junctional complex, which was significantly reduced in the EB1 KD oocytes. Collectively, we propose that the impairment of EB1 function manipulates spindle formation, thereby promoting chromosomal loss, which is expected to fuel aneuploidy and possibly fertilization failure.

Keywords

chromosome misalignmentEB1meiosisoocytespindle formation
Type
Biological Applications
Information
Microscopy and Microanalysis , Volume 27 , Issue 2 , April 2021 , pp. 385 - 391
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Copyright
Copyright © The Author(s), 2021. Published by Cambridge University Press

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References

Akhmanova, A & Steinmetz, MO (2010). Microtubule + TIPs at a glance. J Cell Sci 123, 3415–3419.CrossRefGoogle ScholarPubMed
Alberico, EO, Zhu, ZC, Wu, YO, Gardner, MK, Kovar, DR & Goodson, HV (2016). Interactions between the microtubule binding protein EB1 and F-actin. J Mol Biol 428(6), 1304–1314.CrossRefGoogle ScholarPubMed
Applewhite, DA, Grode, KD, Duncan, MC & Rogers, SL (2013). The actin microtubule cross-linking activity of Drosophila short stop is regulated by intramolecular inhibition. Mol Biol Cell 24, 2885–2893.CrossRefGoogle ScholarPubMed
Banerjee, B, Kestner, CA & Stukenberg, PT (2014). EB1 enables spindle microtubules to regulate centromeric recruitment of Aurora B. J Cell Biol 204, 947–963.CrossRefGoogle ScholarPubMed
Beinhauer, JD, Hagan, IM, Hegemann, JH & Fleig, U (1997). Mal3, the fission yeast homologue of the human APC-interacting protein EB-1 is required for microtubule integrity and the maintenance of cell form. J Cell Biol 139, 717–728.CrossRefGoogle ScholarPubMed
Bennabi, I, Terret, ME & Verlhac, MH (2016). Meiotic spindle assembly and chromosome segregation in oocytes. J Cell Biol 215, 611–619.CrossRefGoogle ScholarPubMed
Boettcher, M & McManus, MT (2015). Choosing the right tool for the job: RNAi, TALEN or CRISPR. Mol Cell 58, 575–585.CrossRefGoogle ScholarPubMed
Breuer, M & Verlhac, MH (2010). HURP permits MTOC sorting for robust meiotic spindle bipolarity, similar to extra centrosome clustering in cancer cells. J Cell Biol 191, 1251–1260.CrossRefGoogle ScholarPubMed
Chan, YW, Jeyaprakash, AA, Nigg, EA & Santamaria, A (2012). Aurora B controls kinetochore-microtubule attachments by inhibiting Ska complex KMN network interaction. J Cell Biol 196, 563–571.CrossRefGoogle ScholarPubMed
Chausovsky, A, Bershadsky, AD & Borisy, GG (2000). Cadherin-mediated regulation of microtubule dynamics. Nat Cell Biol 2, 797–804.CrossRefGoogle ScholarPubMed
Clift, D, McEwan, WA, Labzin, LI, Konieczny, V, Mogessie, B, James, LC & Schuh, M (2017). A method for the acute and rapid degradation of endogenous proteins. Cell 171, 1692–1706.CrossRefGoogle ScholarPubMed
Clift, D, So, C, McEwan, WA, James, LC & Schuh, M (2018). Acute and rapid degradation of endogenous proteins by Trim-Away. Nat Protoc 13, 2149–2175.CrossRefGoogle ScholarPubMed
Dikovskaya, D, Newton, IP & Näthke, IS (2004). The adenomatous polyposis coli protein is required for the formation of robust spindles formed in CSF Xenopus extracts. Mol Biol Cell 15, 2978–2991.CrossRefGoogle ScholarPubMed
Dikovskaya, D, Schiffmann, D, Newton, IP, Oakley, A, Kroboth, K, Sansom, O, Jamieson, TJ, Meniel, V, Clarke, A, Näthke, IS. (2007). Loss of APC induces polyploidy as a result of a combination of defects in mitosis and apoptosis. J Cell Biol 176, 183–195.CrossRefGoogle ScholarPubMed
Draviam, VM, Shapiro, I, Aldridge, B & Sorger, PK (2006). Misorientation and reduced stretching of aligned sister kinetochores promote chromosome missegregation in EB1- or APC-depleted cells. EMBO J 25, 2814–2827.CrossRefGoogle ScholarPubMed
Eisen, JS & Smith, JC (2008). Controlling morpholino experiments: Don't stop making antisense. Development 135, 1735–1743.CrossRefGoogle ScholarPubMed
Green, RA, Wollman, R & Kaplan, KB (2005). APC and EB1 function together in mitosis to regulate spindle dynamics and chromosome alignment. Mol Biol Cell 16, 4609–4622.CrossRefGoogle ScholarPubMed
Jeyaprakash, AA, et al. (2012). Structural and functional organization of the Ska complex, a key component of the kinetochore-microtubule interface. Mol Cell 46, 274–286.CrossRefGoogle ScholarPubMed
Jiang, K & Akhmanova, A (2011). Microtubule tip-interacting proteins: A view from both ends. Curr Opin Cell Biol 23, 94–101.CrossRefGoogle ScholarPubMed
Kim, SJ (2019). The chronic and unpredictable stress suppressed Kisspeptin expression during ovarian cycle in mice. JARB 34(1), 40–49.Google Scholar
King, JS, Veltman, DM, Georgiou, M, Baum, B & Insall, RH (2010). SCAR/WAVE is activated at mitosis and drives myosin-independent cytokinesis. J Cell Sci 123(Pt 13), 2246–2255.CrossRefGoogle ScholarPubMed
Komarova, Y, et al. (2009). Mammalian end binding proteins control persistent microtubule growth. J Cell Biol 184, 691–706.CrossRefGoogle ScholarPubMed
Kumar, P & Wittmann, T (2012). TIPs: SxIPping along microtubule ends. Trends Cell Biol 22, 418–428.CrossRefGoogle ScholarPubMed
Liang, S, Guo, J, Choi, JW, Shin, KT, Wang, HY, Jo, YJ, Kim, NH & Cui, XS (2017). Protein phosphatase 2A regulatory subunit B55α functions in mouse oocyte maturation and early embryonic development. Oncotarget 8(16), 26979–26991.CrossRefGoogle ScholarPubMed
Nagafuchi, A, Ishihara, S & Tsukita, S (1994). The roles of catenins in the cadherin-mediated cell adhesion: Functional analysis of E-cadherin-alpha catenin fusion molecules. J Cell Biol 127, 235–245.CrossRefGoogle ScholarPubMed
Paynton, BV, Rempel, R & Bachvarova, R (1988). Changes in state of adenylation and time course of degradation of maternal mRNAs during oocyte maturation and early embryonic development in the mouse. Dev Biol 129, 304–314.CrossRefGoogle ScholarPubMed
Pruyne, D & Bretscher, A (2000). Polarization of cell growth in yeast. J Cell Sci 113(Pt 4), 571–585.CrossRefGoogle ScholarPubMed
Rehberg, M & Gräf, R (2002). Dictyostelium EB1 is a genuine centrosomal component required for proper spindle formation. Mol Biol Cell 13(7), 2301–2310.CrossRefGoogle ScholarPubMed
Roeles, J & Tsiavaliaris, G (2019). Actin-microtubule interplay coordinates spindle assembly in human oocytes. Nat Commun 10, 4651.CrossRefGoogle ScholarPubMed
Schober, JM, Cain, JM, Komarova, YA & Borisy, GG (2009). Migration and actin protrusion in melanoma cells are regulated by EB1 protein. Cancer Lett 284(1), 30–36.CrossRefGoogle ScholarPubMed
Schuh, M & Ellenberg, J (2007). Self-organization of MTOCs replaces centrosome function during acentrosomal spindle assembly in live mouse oocytes. Cell 130, 484–498.CrossRefGoogle ScholarPubMed
Siegrist, SE & Doe, CQ (2007). Microtubule-induced cortical cell polarity. Genes Dev 21, 483–496.CrossRefGoogle ScholarPubMed
Sirajuddin, M, Rice, LM & Vale, RD (2014). Regulation of microtubule motors by tubulin isotypes and post-translational modifications. Nat Cell Biol 16, 335–344.CrossRefGoogle ScholarPubMed
Sjöblom, B, Ylänne, J & Djinović-Carugo, K (2008). Novel structural insights into F-actin-binding and novel functions of calponin homology domains. Curr Opin Struct Biol 18(6), 702–708.CrossRefGoogle ScholarPubMed
Su, X, Ohi, R & Pellman, D (2012). Move in for the kill: Motile microtubule regulators. Trends Cell Biol 22, 567–575.CrossRefGoogle ScholarPubMed
Su, YQ, Sugiura, K, Woo, Y, Wigglesworth, K, Kamdar, S, Affourtit, J & Eppig, JJ (2007). Selective degradation of transcripts during meiotic maturation of mouse oocytes. Dev Biol 302(1), 104–117.CrossRefGoogle ScholarPubMed
Tamura, N & Draviam, VM (2012). Microtubule plus-ends within a mitotic cell are ‘moving platforms’ with anchoring, signalling and force-coupling roles. Open Biol 2, 120–132.CrossRefGoogle ScholarPubMed
Tang, EI, Mok, KW, Lee, WM & Cheng, CY (2015). EB1 regulates tubulin and actin cytoskeletal networks at the sertoli cell blood-testis barrier in male rats: An in vitro study. Endocrinology 156(2), 680–693.CrossRefGoogle Scholar
Terenna, CR, Makushok, T, Velve-Casquillas, G, Baigl, D, Chen, Y, Bornens, M, Paoletti, A, Piel, M & Tran, PT (2008). Physical mechanisms redirecting cell polarity and cell shape in fission yeast. Curr Biol 18, 1748–1753.CrossRefGoogle ScholarPubMed
Thomas, GE, Bandopadhyay, K, Sutradhar, S, Renjith, MR, Singh, P, et al. (2016). EB1 regulates attachment of Ska1 with microtubules by forming extended structures on the microtubule lattice. Nat Commun 7, 11665.CrossRefGoogle ScholarPubMed
Tian, Y, Tian, X, Gawlak, G, O'Donnell, JJ 3rd, Sacks, DB & Birukova, AA (2014). IQGAP1 regulates endothelial barrier function via EB1-cortactin cross talk. Mol Cell Biol 34, 3546–3558.CrossRefGoogle ScholarPubMed
Tirnauer, JS & Bierer, BE. (2000). Eb1 Proteins Regulate Microtubule Dynamics, Cell Polarity, and Chromosome Stability. Journal of Cell Biology 149(4), 761–766. doi: http://dx.doi.org/10.1083/jcb.149.4.761.CrossRefGoogle ScholarPubMed
Tirnauer, JS, Grego, S, Salmon, ED & Mitchison, TJ (2002). EB1-microtubule interactions in Xenopus egg extracts: Role of EB1 in microtubule stabilization and mechanisms of targeting to microtubules. Mol Biol Cell 13, 3614–3626.Google ScholarPubMed
Valenta, T, Hausmann, G & Basler, K (2012). The many faces and functions of β-catenin. EMBO J 31(12), 2714–2736.CrossRefGoogle ScholarPubMed
Wang, Y, Zhou, X, Zhu, H, et al. (2005). Overexpression of EB1 in human esophageal squamous cell carcinoma (ESCC) may promote cellular growth by activating beta-catenin/TCF pathway. Oncogene 24, 6637–6645.CrossRefGoogle ScholarPubMed
Wittmann, T, Hyman, A & Desai, A (2001). The spindle: A dynamic assembly of microtubules and motors. Nat Cell Biol 3, 28–34.CrossRefGoogle ScholarPubMed
Zhang, T, Zaal, KJ, Sheridan, J, Mehta, A, Gundersen, GG & Ralston, E (2009). Microtubule plus-end binding protein EB1 is necessary for muscle cell differentiation, elongation and fusion. J Cell Sci 122(Pt 9), 1401–1409.CrossRefGoogle ScholarPubMed

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