scholarly journals Random chromosome distribution in the first meiosis of F1 disomic substitution line 2R(2D) x rye hybrids (ABDR, 4× = 28) occurs without bipolar spindle assembly

2020 ◽  
Vol 14 (4) ◽  
pp. 453-482
Author(s):  
Dina B. Loginova ◽  
Anastasia A. Zhuravleva ◽  
Olga G. Silkova
Keyword(s):  
Genome Structure ◽  
Spindle Assembly ◽  
Wheat Chromosome ◽  
Cell Plate ◽  
Spindle Formation ◽  
Bipolar Spindle ◽  
Chromosome Distribution ◽  
Bipolar Spindles ◽  
Mitosis And Meiosis ◽  
Spindle Structure

The assembly of the microtubule-based spindle structure in plant meiosis remains poorly understood compared with our knowledge of mitotic spindle formation. One of the approaches in our understanding of microtubule dynamics is to study spindle assembly in meiosis of amphyhaploids. Using immunostaining with phH3Ser10, CENH3 and α-tubulin-specific antibodies, we studied the chromosome distribution and spindle organisation in meiosis of F1 2R(2D)xR wheat-rye hybrids (genome structure ABDR, 4× = 28), as well as in wheat and rye mitosis and meiosis. At the prometaphase of mitosis, spindle assembly was asymmetric; one half of the spindle assembled before the other, with simultaneous chromosome alignment in the spindle mid-zone. At diakinesis in wheat and rye, microtubules formed a pro-spindle which was subsequently disassembled followed by a bipolar spindle assembly. In the first meiosis of hybrids 2R(2D)xR, a bipolar spindle was not found and the kinetochore microtubules distributed the chromosomes. Univalent chromosomes are characterised by a monopolar orientation and maintenance of sister chromatid and centromere cohesion. Presence of bivalents did not affect the formation of a bipolar spindle. Since the central spindle was absent, phragmoplast originates from “interpolar” microtubules generated by kinetochores. Cell plate development occurred with a delay. However, meiocytes in meiosis II contained apparently normal bipolar spindles. Thus, we can conclude that: (1) cohesion maintenance in centromeres and between arms of sister chromatids may negatively affect bipolar spindle formation in the first meiosis; (2) 2R/2D rye/wheat chromosome substitution affects the regulation of the random chromosome distribution in the absence of a bipolar spindle.

scholarly journals Bimodal Interaction of Mammalian Polo-Like Kinase 1 and a Centrosomal Scaffold, Cep192, in the Regulation of Bipolar Spindle Formation

2015 ◽  
Vol 35 (15) ◽  
pp. 2626-2640 ◽  
Author(s):  
Lingjun Meng ◽  
Jung-Eun Park ◽  
Tae-Sung Kim ◽  
Eun Hye Lee ◽  
Suk-Youl Park ◽  
...  
Keyword(s):  
Xenopus Laevis ◽  
Spindle Assembly ◽  
Human Cells ◽  
Mitotic Progression ◽  
Spindle Formation ◽  
Content Type ◽  
Bipolar Spindle ◽  
Egg Extracts ◽  
Cultured Human Cells ◽  
Bipolar Spindles

Serving as microtubule-organizing centers, centrosomes play a key role in forming bipolar spindles. The mechanism of how centrosomes promote bipolar spindle assembly in various organisms remains largely unknown. A recent study withXenopus laevisegg extracts suggested that the Plk1 ortholog Plx1 interacts with the phospho-T46 (p-T46) motif ofXenopusCep192 (xCep192) to form an xCep192-mediated xAurA-Plx1 cascade that is critical for bipolar spindle formation. Here, we demonstrated that in cultured human cells, Cep192 recruits AurA and Plk1 in a cooperative manner, and this event is important for the reciprocal activation of AurA and Plk1. Strikingly, Plk1 interacted with Cep192 through either the p-T44 (analogous toXenopusp-T46) or the newly identified p-S995 motif via its C-terminal noncatalytic polo-box domain. The interaction between Plk1 and the p-T44 motif was prevalent in the presence of Cep192-bound AurA, whereas the interaction of Plk1 with the p-T995 motif was preferred in the absence of AurA binding. Notably, the loss of p-T44- and p-S995-dependent Cep192-Plk1 interactions induced an additive defect in recruiting Plk1 and γ-tubulin to centrosomes, which ultimately led to a failure in proper bipolar spindle formation and mitotic progression. Thus, we propose that Plk1 promotes centrosome-based bipolar spindle formation by forming two functionally nonredundant complexes with Cep192.

scholarly journals Chromosomes initiate spindle assembly upon experimental dissolution of the nuclear envelope in grasshopper spermatocytes.

1995 ◽  
Vol 131 (5) ◽  
pp. 1125-1131 ◽  
Author(s):  
D Zhang ◽  
R B Nicklas
Keyword(s):  
Nuclear Envelope ◽  
Essential Role ◽  
Spindle Assembly ◽  
Microtubule Assembly ◽  
Chromosomal Protein ◽  
Spindle Microtubule ◽  
Late Prophase ◽  
Spindle Formation ◽  
Bipolar Spindle

Chromosomes are known to enhance spindle microtubule assembly in grasshopper spermatocytes, which suggested to us that chromosomes might play an essential role in the initiation of spindle formation. Chromosomes might, for example, activate other spindle components such as centrosomes and tubulin subunits upon the breakdown of the nuclear envelope. We tested this possibility in living grasshopper spermatocytes. We ruptured the nuclear envelope during prophase, which prematurely exposed the centrosomes to chromosomes and nuclear sap. Spindle assembly was promptly initiated. In contrast, assembly of the spindle was completely inhibited if the nucleus was mechanically removed from a late prophase cell. Other experiments showed that the trigger for spindle assembly is associated with the chromosomes; other constituents of the nucleus cannot initiate spindle assembly in the absence of the chromosomes. The initiation of spindle assembly required centrosomes as well as chromosomes. Extracting centrosomes from late prophase cells completely inhibited spindle assembly after dissolution of the nuclear envelope. We conclude that the normal formation of a bipolar spindle in grasshopper spermatocytes is regulated by chromosomes. A possible explanation is an activator, perhaps a chromosomal protein (Yeo, J.-P., F. Alderuccio, and B.-H. Toh. 1994a. Nature (Lond.). 367: 288-291), that promotes and stabilizes the assembly of astral microtubules and thus promotes assembly of the spindle.

scholarly journals Kinesin-5 Is Dispensable for Bipolar Spindle Formation and Elongation in Candida albicans, but Simultaneous Loss of Kinesin-14 Activity Is Lethal

mSphere ◽  
2019 ◽  
Vol 4 (6) ◽  
Author(s):  
Irsa Shoukat ◽  
Corey Frazer ◽  
John S. Allingham
Keyword(s):  
Candida Albicans ◽  
Mitotic Spindle ◽  
Fungal Pathogens ◽  
Spindle Assembly ◽  
Spindle Pole ◽  
Content Type ◽  
Bipolar Spindle ◽  
Spindle Elongation ◽  
Simultaneous Loss ◽  
Bipolar Spindles

ABSTRACT Mitotic spindles assume a bipolar architecture through the concerted actions of microtubules, motors, and cross-linking proteins. In most eukaryotes, kinesin-5 motors are essential to this process, and cells will fail to form a bipolar spindle without kinesin-5 activity. Remarkably, inactivation of kinesin-14 motors can rescue this kinesin-5 deficiency by reestablishing the balance of antagonistic forces needed to drive spindle pole separation and spindle assembly. We show that the yeast form of the opportunistic fungus Candida albicans assembles bipolar spindles in the absence of its sole kinesin-5, CaKip1, even though this motor exhibits stereotypical cell-cycle-dependent localization patterns within the mitotic spindle. However, cells lacking CaKip1 function have shorter metaphase spindles and longer and more numerous astral microtubules. They also show defective hyphal development. Interestingly, a small population of CaKip1-deficient spindles break apart and reform two bipolar spindles in a single nucleus. These spindles then separate, dividing the nucleus, and then elongate simultaneously in the mother and bud or across the bud neck, resulting in multinucleate cells. These data suggest that kinesin-5-independent mechanisms drive assembly and elongation of the mitotic spindle in C. albicans and that CaKip1 is important for bipolar spindle integrity. We also found that simultaneous loss of kinesin-5 and kinesin-14 (CaKar3Cik1) activity is lethal. This implies a divergence from the antagonistic force paradigm that has been ascribed to these motors, which could be linked to the high mitotic error rate that C. albicans experiences and often exploits as a generator of diversity. IMPORTANCE Candida albicans is one of the most prevalent fungal pathogens of humans and can infect a broad range of niches within its host. This organism frequently acquires resistance to antifungal agents through rapid generation of genetic diversity, with aneuploidy serving as a particularly important adaptive mechanism. This paper describes an investigation of the sole kinesin-5 in C. albicans, which is a major regulator of chromosome segregation. Contrary to other eukaryotes studied thus far, C. albicans does not require kinesin-5 function for bipolar spindle assembly or spindle elongation. Rather, this motor protein associates with the spindle throughout mitosis to maintain spindle integrity. Furthermore, kinesin-5 loss is synthetically lethal with loss of kinesin-14—canonically an opposing force producer to kinesin-5 in spindle assembly and anaphase. These results suggest a significant evolutionary rewiring of microtubule motor functions in the C. albicans mitotic spindle, which may have implications in the genetic instability of this pathogen.

scholarly journals Kinetochore-mediated outward force promotes spindle pole separation in fission yeast

2019 ◽  
Vol 30 (22) ◽  
pp. 2802-2813 ◽  
Author(s):  
Yutaka Shirasugi ◽  
Masamitsu Sato
Keyword(s):  
Fission Yeast ◽  
Motor Proteins ◽  
Spindle Assembly ◽  
Spindle Pole ◽  
Double Knockout ◽  
Bipolar Spindle ◽  
Spindle Poles ◽  
Bipolar Spindles ◽  
Meiosis I

Bipolar spindles are organized by motor proteins that generate microtubule-­dependent forces to separate the two spindle poles. The fission yeast Cut7 (kinesin-5) is a plus-end-directed motor that generates the outward force to separate the two spindle poles, whereas the minus-end-directed motor Pkl1 (kinesin-14) generates the inward force. Balanced forces by these antagonizing kinesins are essential for bipolar spindle organization in mitosis. Here, we demonstrate that chromosomes generate another outward force that contributes to the bipolar spindle assembly. First, it was noted that the cut7 pkl1 double knockout failed to separate spindle poles in meiosis I, although the mutant is known to succeed it in mitosis. It was assumed that this might be because meiotic kinetochores of bivalent chromosomes joined by cross-overs generate weaker tensions in meiosis I than the strong tensions in mitosis generated by tightly tethered sister kinetochores. In line with this idea, when meiotic mono-oriented kinetochores were artificially converted to a mitotic bioriented layout, the cut7 pkl1 mutant successfully separated spindle poles in meiosis I. Therefore, we propose that spindle pole separation is promoted by outward forces transmitted from kinetochores to spindle poles through microtubules.

scholarly journals Centriole and PCM cooperatively recruit CEP192 to spindle poles to promote bipolar spindle assembly

2021 ◽  
Vol 220 (2) ◽  
Author(s):  
Takumi Chinen ◽  
Kaho Yamazaki ◽  
Kaho Hashimoto ◽  
Ken Fujii ◽  
Koki Watanabe ◽  
...  
Keyword(s):  
A549 Cells ◽  
Spindle Assembly ◽  
Spindle Pole ◽  
Scaffold Proteins ◽  
Spindle Formation ◽  
Bipolar Spindle ◽  
Spindle Poles ◽  
Pericentriolar Material ◽  
Mitotic Cells

The pericentriolar material (PCM) that accumulates around the centriole expands during mitosis and nucleates microtubules. Here, we show the cooperative roles of the centriole and PCM scaffold proteins, pericentrin and CDK5RAP2, in the recruitment of CEP192 to spindle poles during mitosis. Systematic depletion of PCM proteins revealed that CEP192, but not pericentrin and/or CDK5RAP2, was crucial for bipolar spindle assembly in HeLa, RPE1, and A549 cells with centrioles. Upon double depletion of pericentrin and CDK5RAP2, CEP192 that remained at centriole walls was sufficient for bipolar spindle formation. In contrast, through centriole removal, we found that pericentrin and CDK5RAP2 recruited CEP192 at the acentriolar spindle pole and facilitated bipolar spindle formation in mitotic cells with one centrosome. Furthermore, the perturbation of PLK1, a critical kinase for PCM assembly, efficiently suppressed bipolar spindle formation in mitotic cells with one centrosome. Overall, these data suggest that the centriole and PCM scaffold proteins cooperatively recruit CEP192 to spindle poles and facilitate bipolar spindle formation.

scholarly journals Spindle assembly and cytokinesis in the absence of chromosomes during Drosophila male meiosis

2003 ◽  
Vol 160 (7) ◽  
pp. 993-999 ◽  
Author(s):  
Elisabetta Bucciarelli ◽  
Maria Grazia Giansanti ◽  
Silvia Bonaccorsi ◽  
Maurizio Gatti
Keyword(s):  
Chromosome Segregation ◽  
Spindle Assembly ◽  
Male Meiosis ◽  
Aurora B ◽  
Aurora B Kinase ◽  
Spindle Formation ◽  
Central Spindle ◽  
Morphological Transformations ◽  
Bipolar Spindles ◽  
Drosophila Mutants

Alarge body of work indicates that chromosomes play a key role in the assembly of both acentrosomal and centrosome-containing spindles. In animal systems, the absence of chromosomes either prevents spindle formation or allows the assembly of a metaphase-like spindle that fails to evolve into an ana-telophase spindle. Here, we show that Drosophila secondary spermatocytes can assemble morphologically normal spindles in the absence of chromosomes. The Drosophila mutants fusolo and solofuso are severely defective in chromosome segregation and produce secondary spermatocytes that are devoid of chromosomes. The centrosomes of these anucleated cells form robust asters that give rise to bipolar spindles that undergo the same ana-telophase morphological transformations that characterize normal spindles. The cells containing chromosome-free spindles are also able to assemble regular cytokinetic structures and cleave normally. In addition, chromosome-free spindles normally accumulate the Aurora B kinase at their midzones. This suggests that the association of Aurora B with chromosomes is not a prerequisite for its accumulation at the central spindle, or for its function during cytokinesis.

scholarly journals Kinesin-5 forms a stable bipolar spindle in a fast, irreversible snap

2018 ◽  
Author(s):  
Allen Leary ◽  
Elena Nazarova ◽  
Shannon Sim ◽  
Kristy Shulist ◽  
Paul Francois ◽  
...  
Keyword(s):  
Mitotic Spindle ◽  
Electron Tomography ◽  
Maximal Extension ◽  
Spindle Formation ◽  
Bipolar Spindle ◽  
Spindle Poles ◽  
Equilibrium Length ◽  
Bipolar Spindles

SUMMARYGRAPHICAL ABSTRACTSeparation of duplicated spindle poles is the first step in forming the mitotic spindle. Kinesin-5 crosslinks and slides anti-parallel microtubules, but it is unclear how these two activities contribute to the first steps in spindle formation. In this study we report that in monopolar spindles, the duplicated spindle poles snap apart in a fast and irreversible step that produces a nascent bipolar spindle. Using mutations in Kinesin-5 that inhibit microtubule sliding, we show crosslinking alone drives the fast, irreversible pole separation. Electron tomography revealed microtubule pairs in monopolar spindles have short overlaps that intersect at high angles and are unsuited for ensemble Kinesin-5 sliding. However, maximal extension of a subset of microtubule pairs approaches the length of nascent bipolar spindles and is consistent with a Kinesin-5 crosslinking driven transition. Finally, stochastic microtubule sliding by Kinesin-5 stabilizes the nascent spindle and sets a stereotyped equilibrium length.

scholarly journals OsMTOPVIB is required for meiotic bipolar spindle assembly

2019 ◽  
Vol 116 (32) ◽  
pp. 15967-15972 ◽  
Author(s):  
Zhihui Xue ◽  
Changzhen Liu ◽  
Wenqing Shi ◽  
Yongjie Miao ◽  
Yi Shen ◽  
...  
Keyword(s):  
Arabidopsis Thaliana ◽  
Chromosome Segregation ◽  
Spindle Assembly ◽  
Strand Break ◽  
Microtubule Assembly ◽  
Multipolar Spindle ◽  
Bipolar Spindle ◽  
Multipolar Spindles ◽  
Bipolar Spindles ◽  
Plant Meiosis

The organization of microtubules into a bipolar spindle is essential for chromosome segregation. Both centrosome and chromatin-dependent spindle assembly mechanisms are well studied in mouse, Drosophila melanogaster, and Xenopus oocytes; however, the mechanism of bipolar spindle assembly in plant meiosis remains elusive. According to our observations of microtubule assembly in Oryza sativa, Zea mays, Arabidopsis thaliana, and Solanum lycopersicum, we propose that a key step of plant bipolar spindle assembly is the correction of the multipolar spindle into a bipolar spindle at metaphase I. The multipolar spindles failed to transition into bipolar ones in OsmtopVIB with the defect in double-strand break (DSB) formation. However, bipolar spindles were normally assembled in several other mutants lacking DSB formation, such as Osspo11-1, pair2, and crc1, indicating that bipolar spindle assembly is independent of DSB formation. We further revealed that the mono-orientation of sister kinetochores was prevalent in OsmtopVIB, whereas biorientation of sister kinetochores was frequently observed in Osspo11-1, pair2, and crc1. In addition, mutations of the cohesion subunit OsREC8 resulted in biorientation of sister kinetochores as well as bipolar spindles even in the background of OsmtopVIB. Therefore, we propose that biorientation of the kinetochore is required for bipolar spindle assembly in the absence of homologous recombination.

scholarly journals The Kinesinlike Protein Subito Contributes to Central Spindle Assembly and Organization of the Meiotic Spindle in Drosophila Oocytes

2005 ◽  
Vol 16 (10) ◽  
pp. 4684-4694 ◽  
Author(s):  
J. K. Jang ◽  
T. Rahman ◽  
K. S. McKim
Keyword(s):  
Meiotic Spindle ◽  
Spindle Formation ◽  
Present Evidence ◽  
Homologous Chromosomes ◽  
Central Spindle ◽  
Bipolar Spindle ◽  
Passenger Proteins ◽  
Bipolar Spindles ◽  
Meiosis I

In the oocytes of many species, bipolar spindles form in the absence of centrosomes. Drosophila melanogaster oocyte chromosomes have a major role in nucleating microtubules, which precedes the bundling and assembly of these microtubules into a bipolar spindle. Here we present evidence that a region similar to the anaphase central spindle functions to organize acentrosomal spindles. Subito mutants are characterized by the formation of tripolar or monopolar spindles and nondisjunction of homologous chromosomes at meiosis I. Subito encodes a kinesinlike protein and associates with the meiotic central spindle, consistent with its classification in the Kinesin 6/MKLP1 family. This class of proteins is known to be required for cytokinesis, but our results suggest a new function in spindle formation. The meiotic central spindle appears during prometaphase and includes passenger complex proteins such as AurB and Incenp. Unlike mitotic cells, the passenger proteins do not associate with centromeres before anaphase. In the absence of Subito, central spindle formation is defective and AurB and Incenp fail to properly localize. We propose that Subito is required for establishing and/or maintaining the central spindle in Drosophila oocytes, and this substitutes for the role of centrosomes in organizing the bipolar spindle.

scholarly journals Nek2-mediated GAS2L1 phosphorylation and centrosome-linker disassembly induce centrosome disjunction

2020 ◽  
Vol 219 (5) ◽  
Author(s):  
Franco K.C. Au ◽  
Bill K.T. Hau ◽  
Robert Z. Qi
Keyword(s):  
Chromosome Segregation ◽  
Binding Protein ◽  
Spindle Assembly ◽  
Actin Binding ◽  
Spindle Formation ◽  
Bipolar Spindle ◽  
Actin Binding Protein ◽  
Calponin Homology

Centrosome disjunction occurs in late G2 to facilitate bipolar spindle formation and is mediated by the NIMA-related kinase Nek2. Here, we show that GAS2L1, a microtubule- and F-actin–binding protein required for centrosome disjunction, undergoes Nek2-mediated phosphorylation at Ser352 in G2/M. The phosphorylation is essential for centrosome disjunction in late G2 and for proper spindle assembly and faithful chromosome segregation in mitosis. GAS2L1 contains a calponin-homology (CH) domain and a GAS2-related (GAR) domain, which bind to F-actin and microtubules, respectively. Notably, the CH and GAR domains bind to each other to inhibit the functions of both domains, and Ser352 phosphorylation disrupts the interaction between the two domains and relieves the autoinhibition. We dissected the roles of the GAS2L1 phosphorylation and of centrosome-linker disassembly, which is another Nek2-mediated event, and found that these events together trigger centrosome disjunction. Therefore, our findings demonstrate the concerted Nek2 actions that split the centrosomes in late G2.

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