Sumoylation regulates central spindle protein dynamics during chromosome segregation in oocytes

Meiotic spindles in most species lack centrosomes and the mechanisms that underlie faithful chromosome segregation in acentrosomal meiotic spindles are not well understood. In C. elegans oocytes, spindle microtubules exert a poleward force on chromosomes dependent on the microtubule-stabilising protein CLS-2CLASP. The kinase BUB-1Bub1 and CLS-2CLASP localise in the central-spindle and display a dynamic localisation pattern throughout anaphase but the signals regulating their anaphase-specific localisation remains unknown. We have shown that SUMO regulates BUB-1 localisation during metaphase I. Here, we found that SUMO modification of BUB-1Bub1 is regulated by the SUMO E3 ligase GEI-17 and the SUMO protease ULP-1. SUMO is required for BUB-1 localisation in between segregating chromosomes during early anaphase I, and SUMO depletion partially phenocopies BUB-1Bub1 depletion. We also show that CLS-2CLASP is subject to SUMO-mediated regulation. Over-all, we provide evidence for a novel, SUMO-mediated control of protein dynamics during early anaphase I in oocytes.

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Central-spindle microtubules are strongly coupled to chromosomes during both anaphase A and anaphase B

Molecular Biology of the Cell ◽

10.1091/mbc.e19-01-0074 ◽

2019 ◽

Vol 30 (19) ◽

pp. 2503-2514 ◽

Cited By ~ 11

Author(s):

Che-Hang Yu ◽

Stefanie Redemann ◽

Hai-Yin Wu ◽

Robert Kiewisz ◽

Tae Yeon Yoo ◽

...

Keyword(s):

Chromosome Segregation ◽

Tomographic Reconstruction ◽

Mitotic Spindles ◽

Strongly Coupled ◽

Central Spindle ◽

Spindle Poles ◽

Meiotic Spindles ◽

Spindle Microtubules ◽

Anaphase B ◽

Chromosome Motion

Spindle microtubules, whose dynamics vary over time and at different locations, cooperatively drive chromosome segregation. Measurements of microtubule dynamics and spindle ultrastructure can provide insight into the behaviors of microtubules, helping elucidate the mechanism of chromosome segregation. Much work has focused on the dynamics and organization of kinetochore microtubules, that is, on the region between chromosomes and poles. In comparison, microtubules in the central-spindle region, between segregating chromosomes, have been less thoroughly characterized. Here, we report measurements of the movement of central-spindle microtubules during chromosome segregation in human mitotic spindles and Caenorhabditis elegans mitotic and female meiotic spindles. We found that these central-spindle microtubules slide apart at the same speed as chromosomes, even as chromosomes move toward spindle poles. In these systems, damaging central-spindle microtubules by laser ablation caused an immediate and complete cessation of chromosome motion, suggesting a strong coupling between central-spindle microtubules and chromosomes. Electron tomographic reconstruction revealed that the analyzed anaphase spindles all contain microtubules with both ends between segregating chromosomes. Our results provide new dynamical, functional, and ultrastructural characterizations of central-spindle microtubules during chromosome segregation in diverse spindles and suggest that central-spindle microtubules and chromosomes are strongly coupled in anaphase.

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Central spindle microtubules are strongly coupled to chromosomes during both anaphase A and anaphase B

10.1101/537290 ◽

2019 ◽

Cited By ~ 2

Author(s):

Che-Hang Yu ◽

Stefanie Redemann ◽

Hai-Yin Wu ◽

Robert Kiewisz ◽

Tae Yeon Yoo ◽

...

Keyword(s):

Chromosome Segregation ◽

Tomographic Reconstruction ◽

Mitotic Spindles ◽

Strongly Coupled ◽

Central Spindle ◽

Spindle Poles ◽

Meiotic Spindles ◽

Spindle Microtubules ◽

Anaphase B ◽

Chromosome Motion

AbstractSpindle microtubules, whose dynamics vary over time and at different locations, cooperatively drive chromosome segregation. Measurements of microtubule dynamics and spindle ultrastructure can provide insight into the behaviors of microtubules, helping elucidate the mechanism of chromosome segregation. Much work has focused on the dynamics and organization of kinetochore microtubules, i.e. on the region between chromosomes and poles. In comparison, microtubules in the central spindle region, between segregating chromosomes, have been less thoroughly characterized. Here, we report measurements of the movement of central spindle microtubules during chromosome segregation in human mitotic spindles, and Caenorhabditis elegans mitotic and female meiotic spindles. We found that these central spindle microtubules slide apart at the same speed as chromosomes, even as chromosomes move towards spindle poles. In these systems, damaging central spindle microtubules by laser ablation caused an immediate and complete cessation of chromosome motion, suggesting a strong coupling between central spindle microtubules and chromosomes. Electron tomographic reconstruction revealed that the analyzed anaphase spindles all contain microtubules with both ends between segregating chromosomes. Our results provide new dynamical, functional, and ultrastructural characterizations of central spindle microtubules during chromosome segregation in diverse spindles, and suggest that central spindle microtubules and chromosomes are strongly coupled in anaphase.

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ZYG-9ch-TOG promotes the stability of acentrosomal poles via regulation of spindle microtubules in C. elegans oocyte meiosis

10.1101/2022.01.04.474888 ◽

2022 ◽

Author(s):

Gabriel Cavin-Meza ◽

Timothy J. Mullen ◽

Ian D. Wolff ◽

Emily R. Czajkowski ◽

Nikita Santosh Divekar ◽

...

Keyword(s):

Fluorescence Recovery After Photobleaching ◽

Structural Role ◽

C Elegans ◽

Oocyte Meiosis ◽

Pole Structure ◽

Meiotic Spindles ◽

Spindle Microtubules ◽

Per Se ◽

The Stability ◽

Bipolar Spindles

During mitosis, centrosomes serve as microtubule organizing centers that guide the formation of a bipolar spindle. However, oocytes of many species lack centrosomes; how meiotic spindles establish and maintain these acentrosomal poles remains poorly understood. Here, we show that the microtubule polymerase ZYG-9ch-TOG is required to maintain acentrosomal pole integrity in C. elegans oocyte meiosis; following acute depletion of ZYG-9 from pre-formed spindles, the poles split apart and an unstable multipolar structure forms. Depletion of TAC-1, a protein known to interact with ZYG-9 in mitosis, caused loss of proper ZYG-9 localization and similar spindle phenotypes, further demonstrating that ZYG-9 is required for pole integrity. However, depletion of ZYG-9 surprisingly did not affect the assembly or stability of monopolar spindles, suggesting that ZYG-9 is not required for acentrosomal pole structure per se. Moreover, fluorescence recovery after photobleaching (FRAP) revealed that ZYG-9 turns over rapidly at acentrosomal poles, displaying similar turnover dynamics to tubulin itself, suggesting that ZYG-9 does not play a static structural role at poles. Together, these data support a global role for ZYG-9 in regulating the stability of bipolar spindles and demonstrate that the maintenance of acentrosomal poles requires factors beyond those acting to organize the pole structure itself.

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Male meiotic spindle features that efficiently segregate paired and lagging chromosomes

eLife ◽

10.7554/elife.50988 ◽

2020 ◽

Vol 9 ◽

Cited By ~ 4

Author(s):

Gunar Fabig ◽

Robert Kiewisz ◽

Norbert Lindow ◽

James A Powers ◽

Vanessa Cota ◽

...

Keyword(s):

Chromosome Segregation ◽

Electron Tomography ◽

Male Meiosis ◽

Meiotic Spindle ◽

Sperm Production ◽

C Elegans ◽

Unpaired Chromosome ◽

Meiotic Spindles ◽

Anaphase B ◽

Distance Change

Chromosome segregation during male meiosis is tailored to rapidly generate multitudes of sperm. Little is known about mechanisms that efficiently partition chromosomes to produce sperm. Using live imaging and tomographic reconstructions of spermatocyte meiotic spindles in Caenorhabditis elegans, we find the lagging X chromosome, a distinctive feature of anaphase I in C. elegans males, is due to lack of chromosome pairing. The unpaired chromosome remains tethered to centrosomes by lengthening kinetochore microtubules, which are under tension, suggesting that a ‘tug of war’ reliably resolves lagging. We find spermatocytes exhibit simultaneous pole-to-chromosome shortening (anaphase A) and pole-to-pole elongation (anaphase B). Electron tomography unexpectedly revealed spermatocyte anaphase A does not stem solely from kinetochore microtubule shortening. Instead, movement of autosomes is largely driven by distance change between chromosomes, microtubules, and centrosomes upon tension release during anaphase. Overall, we define novel features that segregate both lagging and paired chromosomes for optimal sperm production.

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CENP-32 is required to maintain centrosomal dominance in bipolar spindle assembly

Molecular Biology of the Cell ◽

10.1091/mbc.e14-09-1366 ◽

2015 ◽

Vol 26 (7) ◽

pp. 1225-1237 ◽

Cited By ~ 3

Author(s):

Shinya Ohta ◽

Laura Wood ◽

Iyo Toramoto ◽

Ken-Ichi Yagyu ◽

Tatsuo Fukagawa ◽

...

Keyword(s):

Chromosome Segregation ◽

Additional Data ◽

Weak Interactions ◽

Metaphase Plate ◽

Spindle Pole ◽

Animal Cells ◽

Central Spindle ◽

Spindle Poles ◽

Spindle Microtubules ◽

Bipolar Spindles

Centrosomes nucleate spindle formation, direct spindle pole positioning, and are important for proper chromosome segregation during mitosis in most animal cells. We previously reported that centromere protein 32 (CENP-32) is required for centrosome association with spindle poles during metaphase. In this study, we show that CENP-32 depletion seems to release centrosomes from bipolar spindles whose assembly they had previously initiated. Remarkably, the resulting anastral spindles function normally, aligning the chromosomes to a metaphase plate and entering anaphase without detectable interference from the free centrosomes, which appear to behave as free asters in these cells. The free asters, which contain reduced but significant levels of CDK5RAP2, show weak interactions with spindle microtubules but do not seem to make productive attachments to kinetochores. Thus CENP-32 appears to be required for centrosomes to integrate into a fully functional spindle that not only nucleates astral microtubules, but also is able to nucleate and bind to kinetochore and central spindle microtubules. Additional data suggest that NuMA tethers microtubules at the anastral spindle poles and that augmin is required for centrosome detachment after CENP-32 depletion, possibly due to an imbalance of forces within the spindle.

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The structural basis for Ulp2 recruitment to the kinetochore

10.1101/2020.12.30.424835 ◽

2021 ◽

Author(s):

Yun Quan ◽

Stephen M. Hinshaw ◽

Pang-Che Wang ◽

Stephen C. Harrison ◽

Huilin Zhou

Keyword(s):

Cell Cycle ◽

Cell Division ◽

Chromosome Segregation ◽

Structural Basis ◽

Centromeric Dna ◽

Step Process ◽

Sumo Protease ◽

Daughter Cells ◽

Sister Chromatids ◽

Spindle Microtubules

ABSTRACTThe step-by-step process of chromosome segregation defines the stages of the cell division cycle. In eukaryotes, signaling pathways that control these steps converge upon the kinetochore, a multiprotein assembly that connects spindle microtubules to the centromere of each chromosome. Kinetochores control and adapt to major chromosomal transactions, including replication of centromeric DNA, biorientation of sister centromeres on the metaphase spindle, and transit of sister chromatids into daughter cells during anaphase. Although the mechanisms that ensure tight microtubule coupling at anaphase are at least partly understood, kinetochore adaptations that support other cell cycle transitions are not. We report here a mechanism that enables regulated control of kinetochore sumoylation. A conserved surface of the Ctf3/CENP-I kinetochore protein provides a binding site for the SUMO protease, Ulp2. Ctf3 mutations that disable Ulp2 recruitment cause elevated inner kinetochore sumoylation and defective chromosome segregation. The location of the site within the assembled kinetochore suggests coordination between sumoylation and other cell cycle-regulated processes.

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Sumoylation regulates protein dynamics during meiotic chromosome segregation in C. elegans oocytes

Journal of Cell Science ◽

10.1242/jcs.232330 ◽

2019 ◽

Vol 132 (14) ◽

pp. jcs232330 ◽

Cited By ~ 13

Author(s):

Federico Pelisch ◽

Laura Bel Borja ◽

Ellis G. Jaffray ◽

Ronald T. Hay

Keyword(s):

Protein Dynamics ◽

Chromosome Segregation ◽

Meiotic Chromosome ◽

C Elegans

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Spindle Self-organization and Cytokinesis During Male Meiosis in asterless Mutants of Drosophila melanogaster

The Journal of Cell Biology ◽

10.1083/jcb.142.3.751 ◽

1998 ◽

Vol 142 (3) ◽

pp. 751-761 ◽

Cited By ~ 115

Author(s):

Silvia Bonaccorsi ◽

Maria Grazia Giansanti ◽

Maurizio Gatti

Keyword(s):

Drosophila Melanogaster ◽

Chromosome Segregation ◽

Spindle Assembly ◽

Male Meiosis ◽

Self Organization ◽

Wild Type ◽

Female Meiosis ◽

Classical View ◽

Central Spindle ◽

Spindle Microtubules

While Drosophila female meiosis is anastral, both meiotic divisions in Drosophila males exhibit prominent asters. We have identified a gene we call asterless (asl) that is required for aster formation during male meiosis. Ultrastructural analysis showed that asl mutants have morphologically normal centrioles. However, immunostaining with antibodies directed either to γ tubulin or centrosomin revealed that these proteins do not accumulate in the centrosomes, as occurs in wild-type. Thus, asl appears to specify a function required for the assembly of centrosomal material around the centrioles. Despite the absence of asters, meiotic cells of asl mutants manage to develop an anastral spindle. Microtubules grow from multiple sites around the chromosomes, and then focus into a peculiar bipolar spindle that mediates chromosome segregation, although in a highly irregular way. Surprisingly, asl spermatocytes eventually form a morphologically normal ana–telophase central spindle that has full ability to stimulate cytokinesis. These findings challenge the classical view on central spindle assembly, arguing for a self-organization of this structure from either preexisting or newly formed microtubules. In addition, these findings strongly suggest that the asters are not required for signaling cytokinesis.

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