Gamma-tubulin reorganization during mouse fertilization and early development

1993 ◽  
Vol 104 (2) ◽  
pp. 383-389 ◽  
Author(s):  
M.J. Palacios ◽  
H.C. Joshi ◽  
C. Simerly ◽  
G. Schatten
Keyword(s):  
Early Development ◽  
Basal Body ◽  
Spindle Pole ◽  
Meiotic Spindle ◽  
Cytoplasmic Microtubule ◽  
Mouse Oocytes ◽  
Spindle Pole Bodies ◽  
Preimplantation Stage ◽  
Pericentriolar Material ◽  
Gamma Tubulin

gamma-Tubulin, a component of spindle pole bodies in fungal cells and pericentriolar material in vertebrate cells, is thought to play a role in the nucleation of microtubule growth and to define their polarity. In contrast to the adult somatic cells, microtubules are nucleated in the absence of centrioles in mammalian oocytes and early embryos. By studying acentriolar mouse oocytes and their early development following fertilization, we show that gamma-tubulin antibody crossreacts with a 50,000 M(r) protein in unfertilized mouse oocytes and demonstrate that gamma-tubulin distribution is rearranged dramatically during fertilization. In unfertilized mouse oocytes, gamma-tubulin is concentrated in the broad spindle poles of meiotic spindle (MII) and as the distinct foci which form the centers of the cytoplasmic microtubule asters (cytasters). The integrity of these gamma-tubulin foci and their cytoplasmic location is maintained during the drug- or cold-induced depolymerization of microtubules. gamma-Tubulin is also found in the basal body of the mouse sperm. During fertilization, the gamma-tubulin is found at the cytastral centers as well as in the incorporated sperm basal body complex, and the gamma-tubulin foci coalesce at the perinuclear microtubule organizing regions of the two pronuclei at the first mitotic prophase. During mitosis, gamma-tubulin is found associated with broad bands that form the poles of the first mitotic spindle. By the late preimplantation stage, when newly generated centrioles have been reported to arise, gamma-tubulin remains localized at the centrosome of mitotic cells.(ABSTRACT TRUNCATED AT 250 WORDS)

scholarly journals Spatial signals link exit from mitosis to spindle position

eLife ◽  
2016 ◽  
Vol 5 ◽  
Author(s):  
Jill Elaine Falk ◽  
Dai Tsuchiya ◽  
Jolien Verdaasdonk ◽  
Soni Lacefield ◽  
Kerry Bloom ◽  
...  
Keyword(s):  
Spindle Pole ◽  
Mitotic Exit ◽  
Zone Model ◽  
Cytoplasmic Microtubule ◽  
Binucleate Cells ◽  
Spindle Pole Bodies ◽  
Mitotic Exit Network ◽  
Bud Neck ◽  
Competing Models ◽  
Spindle Position

In budding yeast, if the spindle becomes mispositioned, cells prevent exit from mitosis by inhibiting the mitotic exit network (MEN). The MEN is a signaling cascade that localizes to spindle pole bodies (SPBs) and activates the phosphatase Cdc14. There are two competing models that explain MEN regulation by spindle position. In the 'zone model', exit from mitosis occurs when a MEN-bearing SPB enters the bud. The 'cMT-bud neck model' posits that cytoplasmic microtubule (cMT)-bud neck interactions prevent MEN activity. Here we find that 1) eliminating cMT– bud neck interactions does not trigger exit from mitosis and 2) loss of these interactions does not precede Cdc14 activation. Furthermore, using binucleate cells, we show that exit from mitosis occurs when one SPB enters the bud despite the presence of a mispositioned spindle. We conclude that exit from mitosis is triggered by a correctly positioned spindle rather than inhibited by improper spindle position.

The role of gamma-tubulin in mitotic spindle formation and cell cycle progression in Aspergillus nidulans

1997 ◽  
Vol 110 (5) ◽  
pp. 623-633 ◽  
Author(s):  
M.A. Martin ◽  
S.A. Osmani ◽  
B.R. Oakley
Keyword(s):  
Cell Cycle ◽  
Mitotic Spindle ◽  
Cell Cycle Progression ◽  
Spindle Pole ◽  
Microtubule Assembly ◽  
Cytoplasmic Microtubule ◽  
Cycle Progression ◽  
Spindle Microtubules ◽  
Gamma Tubulin

gamma-Tubulin has been hypothesized to be essential for the nucleation of the assembly of mitotic spindle microtubules, but some recent results suggest that this may not be the case. To clarify the role of gamma-tubulin in microtubule assembly and cell-cycle progression, we have developed a novel variation of the gene disruption/heterokaryon rescue technique of Aspergillus nidulans. We have used temperature-sensitive cell-cycle mutations to synchronize germlings carrying a gamma-tubulin disruption and observe the phenotypes caused by the disruption in the first cell cycle after germination. Our results indicate that gamma-tubulin is absolutely required for the assembly of mitotic spindle microtubules, a finding that supports the hypothesis that gamma-tubulin is involved in spindle microtubule nucleation. In the absence of functional gamma-tubulin, nuclei are blocked with condensed chromosomes for about the length of one cell cycle before chromatin decondenses without nuclear division. Our results indicate that gamma-tubulin is not essential for progression from G1 to G2, for entry into mitosis nor for spindle pole body replication. It is also not required for reactivity of spindle pole bodies with the MPM-2 antibody which recognizes a phosphoepitope important to mitotic spindle formation. Finally, it does not appear to be absolutely required for cytoplasmic microtubule assembly but may play a role in the formation of normal cytoplasmic microtubule arrays.

scholarly journals CEP215 and AURKA regulate spindle pole focusing and aMTOC organization in mouse oocytes

Reproduction ◽  
2020 ◽  
Vol 159 (3) ◽  
pp. 261-274
Author(s):  
Xiaotian Wang ◽  
Claudia Baumann ◽  
Rabindranath De La Fuente ◽  
Maria M Viveiros
Keyword(s):  
Critical Role ◽  
Super Resolution ◽  
Aurora Kinase ◽  
Spindle Pole ◽  
Meiotic Spindle ◽  
Dependent Manner ◽  
Aurora Kinase A ◽  
Mouse Oocytes ◽  
Kinase A

Acentriolar microtubule-organizing centers (aMTOCs) play a critical role in stable meiotic spindle assembly in oocytes, necessary for accurate chromosome segregation. Yet, there is a limited understanding of the essential regulatory components of these unique MTOCs. In somatic cells, CEP215 (Centrosomal Protein 215) serves as an important regulator of centrosome maturation and spindle organization. Here, we assessed whether it has a similar function in mouse oocytes. CEP215 was detected in oocyte lysates and specifically localized to aMTOCs throughout the progression of meiosis in a pericentrin-dependent manner. Super-resolution microscopy revealed CEP215 co-localization with pericentrin and a unique pore/ring-like structural organization of aMTOCs. Interestingly, inhibition of Aurora Kinase A in either MI or MII-stage oocytes resulted in a striking loss of the ring-like aMTOC organization and pronounced CEP215 clustering at spindle poles, as well as shorter spindles with highly focused poles. In vitro siRNA-mediated transcript knockdown effectively reduced CEP215 in approximately 85% of the oocytes. Maturation rates to MII were similar in the Cep215 siRNA and injected controls; however, a high percentage (~40%) of the Cep215-knockdown oocytes showed notable variations in spindle pole focusing. Surprisingly, pericentrin and γ-tubulin localization and fluorescence intensity at aMTOCs were unaltered in knockdown oocytes, contrasting with mitotic cells where CEP215 depletion reduced γ-tubulin at centrosomes. Our results demonstrate that CEP215 is a functional component of oocyte aMTOCs and participates in the regulation of meiotic spindle pole focusing. Moreover, these studies reveal a vital role for Aurora Kinase A activity in the maintenance of aMTOC organization in oocytes.

Nek2 and its substrate, centrobin/Nip2, are required for proper meiotic spindle formation of the mouse oocytes

Zygote ◽  
2010 ◽  
Vol 19 (1) ◽  
pp. 15-20 ◽  
Author(s):  
Seongkeun Sonn ◽  
Goo Taeg Oh ◽  
Kunsoo Rhee
Keyword(s):  
Spindle Assembly ◽  
Early Embryogenesis ◽  
Mouse Oocyte ◽  
Meiotic Spindle ◽  
Biological Functions ◽  
Spindle Formation ◽  
Mouse Oocytes ◽  
Early Embryos ◽  
Pericentriolar Material ◽  
Meiotic Spindles

SummaryA typical centrosome consists of a pair of centrioles embedded in a proteinous matrix called pericentriolar material. However, the centrosomes in the mouse oocytes and early embryos lack centrioles, but consist of the γ-tubulin-enriched vesicle aggregates. We previously revealed that Nek2 and centrobin/Nip2, a centrosomal substrate of Nek2, is critical for the mouse early embryogenesis, especially at the step of spindle assembly during mitosis. In order to expand our understanding of the biological functions of Nek2, we examined expression and knockdown phenotypes of Nek2 and its substrates, centrobin and C-Nap1, in the mouse oocyte. Nek2, centrobin and C-Nap1 in the mouse oocytes were also centrosomal. Suppression of Nek2 and its substrates did not affect meiotic resumption of the oocytes. However, meiosis of the Nek2- and centrobin-suppressed oocytes was not completed, but arrested with defects in spindle assembly. No visible phenotype was observed in the C-Nap1-suppressed oocytes. These results indicate that Nek2 is critical for proper assembly of the meiotic spindles. Centrobin may be a possible substrate of Nek2 responsible for the meiotic spindle assembly in the mouse oocytes.

scholarly journals Cytoplasmic Microtubule Organizing Centers Regulate Meiotic Spindle Positioning in Mouse Oocyte

2020 ◽  
Author(s):  
Daniela Londono Vasquez ◽  
Katherine Rodriguez-Lukey ◽  
Susanta K. Behura ◽  
Ahmed Z. Balboula
Keyword(s):  
Polar Body ◽  
Spindle Pole ◽  
Meiotic Spindle ◽  
Cytoplasmic Microtubule ◽  
Spindle Positioning ◽  
Nuclear Envelope Breakdown ◽  
Oocyte Meiosis ◽  
Polar Bodies ◽  
Polar Body Extrusion

ABSTRACTDuring oocyte meiosis, migration of the spindle and its positioning must be tightly regulated to ensure elimination of the polar bodies and provide developmentally competent euploid eggs. Although the role of F-actin in regulating these critical processes has been studied extensively, little is known whether microtubules (MTs) participate in regulating these processes. Here, we characterize a pool of MTOCs in the oocyte that does not contribute to spindle assembly but instead remains free in the cytoplasm during metaphase I (metaphase cytoplasmic MTOCs; mcMTOCs). In contrast to spindle pole MTOCs, which primarily originate from the perinuclear region in prophase I, the mcMTOCs are found near the cortex of the oocyte. At nuclear envelope breakdown, they exhibit robust nucleation of MTs, which diminishes during polar body extrusion before returning robustly during metaphase II. The asymmetric positioning of the mcMTOCs provides the spindle with a MT-based anchor line to the cortex opposite the site of polar body extrusion. Depletion of mcMTOCs, by laser ablation, or manipulating their numbers, through autophagy inhibition, revealed that the mcMTOCs are required to regulate the timely migration and positioning of the spindle in meiosis. We discuss how forces exerted by F-actin in mediating movement of the spindle to the oocyte cortex are balanced by MT-mediated forces from the mcMTOCs to ensure spindle positioning and timely spindle migration.

The spindle pole body cycle, meiosis, and basidial cytology of the smut fungus Microbotryum violaceum

1991 ◽  
Vol 69 (8) ◽  
pp. 1795-1803 ◽  
Author(s):  
Mary L. Berbee ◽  
Robert Bauer ◽  
F. Oberwinkler
Keyword(s):  
Spindle Pole Body ◽  
Freeze Substitution ◽  
Ustilago Maydis ◽  
Spindle Pole ◽  
Meiotic Spindle ◽  
Smut Fungus ◽  
Microbotryum Violaceum ◽  
Pole Body ◽  
Spindle Pole Bodies ◽  
Middle Piece

Freeze-substituted basidia of the smut fungus Microbotryum violaceum (Ustilaginales, Basidiomycotina) were examined electron microscopically with particular attention to the meiotic spindle pole body cycle and cytoplasmic characters of phylogenetic significance. Prophase basidia contained a subapical cluster of vesicles and tubules. During prophase, the spindle pole body consisted of two globular elements connected by a middle piece. The spindle pole body had an electron-opaque layer near the nucleus, and each globular element was bisected by an electron-opaque disk. The meiosis I spindle extended between two monoglobular, disc-containing spindle pole bodies. During interphase I and II, septa lacking pores divided the basidium between daughter nuclei. In interphase I, a putative new spindle pole body appeared between the nuclear envelope and the monoglobular spindle pole body residual from the first division. In meiosis II, a spindle was again established between two monoglobular spindle pole bodies, each of which again contained an electron-opaque disc. The cytoplasmic characters of M. violaceum are compared with those of Ustilago maydis and Sphacelotheca polygoni-serrulati. Key words: Microbotryum violaceum, basidiomycete, Ustilaginales, spindle pole body, freeze-substitution, ultrastructure.

scholarly journals A highly divergent gamma-tubulin gene is essential for cell growth and proper microtubule organization in Saccharomyces cerevisiae.

1995 ◽  
Vol 131 (6) ◽  
pp. 1775-1788 ◽  
Author(s):  
S G Sobel ◽  
M Snyder
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Cell Growth ◽  
Yeast Cell ◽  
Spindle Pole Body ◽  
Spindle Pole ◽  
Predicted Amino Acid Sequence ◽  
Nuclear Staining ◽  
Microtubule Organization ◽  
Cytoplasmic Microtubule ◽  
Gamma Tubulin

A Saccharomyces cerevisiae gamma-tubulin-related gene, TUB4, has been characterized. The predicted amino acid sequence of the Tub4 protein (Tub4p) is 29-38% identical to members of the gamma-tubulin family. Indirect immunofluorescence experiments using a strain containing an epitope-tagged Tub4p indicate that Tub4p resides at the spindle pole body throughout the yeast cell cycle. Deletion of the TUB4 gene indicates that Tub4p is essential for yeast cell growth. Tub4p-depleted cells arrest during nuclear division; most arrested cells contain a large bud, replicated DNA, and a single nucleus. Immunofluorescence and nuclear staining experiments indicate that cells depleted of Tub4p contain defects in the organization of both cytoplasmic and nuclear microtubule arrays; such cells exhibit nuclear migration failure, defects in spindle formation, and/or aberrantly long cytoplasmic microtubule arrays. These data indicate that the S. cerevisiae gamma-tubulin protein is an important SPB component that organizes both cytoplasmic and nuclear microtubule arrays.

scholarly journals Basal Body Duplication and Maintenance Require One Member of the Tetrahymena thermophila Centrin Gene Family

2005 ◽  
Vol 16 (8) ◽  
pp. 3606-3619 ◽  
Author(s):  
Alexander J. Stemm-Wolf ◽  
Garry Morgan ◽  
Thomas H. Giddings ◽  
Erin A. White ◽  
Robb Marchione ◽  
...  
Keyword(s):  
Basal Body ◽  
Calcium Binding ◽  
Tetrahymena Thermophila ◽  
Essential Gene ◽  
Expression Patterns ◽  
Spindle Pole ◽  
The Other ◽  
Basal Bodies ◽  
Spindle Pole Bodies ◽  
Ef Hand

Centrins, small calcium binding EF-hand proteins, function in the duplication of a variety of microtubule organizing centers. These include centrioles in humans, basal bodies in green algae, and spindle pole bodies in yeast. The ciliate Tetrahymena thermophila contains at least four centrin genes as determined by sequence homology, and these have distinct localization and expression patterns. CEN1's role at the basal body was examined more closely. The Cen1 protein localizes primarily to two locations: one is the site at the base of the basal body where duplication is initiated. The other is the transition zone between the basal body and axoneme. CEN1 is an essential gene, the deletion of which results in the loss of basal bodies, which is likely due to defects in both basal body duplication and basal body maintenance. Analysis of the three other centrins indicates that two of them function at microtubule-rich structures unique to ciliates, whereas the fourth is not expressed under conditions examined in this study, although when artificially expressed it localizes to basal bodies. This study provides evidence that in addition to its previously known function in the duplication of basal bodies, centrin is also important for the integrity of these organelles.

scholarly journals Stable Preanaphase Spindle Positioning Requires Bud6p and an Apparent Interaction between the Spindle Pole Bodies and the Neck

2007 ◽  
Vol 6 (5) ◽  
pp. 797-807 ◽  
Author(s):  
Brian K. Haarer ◽  
Astrid Hoes Helfant ◽  
Scott A. Nelson ◽  
John A. Cooper ◽  
David C. Amberg
Keyword(s):  
Close Association ◽  
Spindle Pole Body ◽  
Genetic Material ◽  
Neck Region ◽  
Spindle Pole ◽  
Positive Signal ◽  
Cytoplasmic Microtubule ◽  
Pole Body ◽  
Spindle Pole Bodies ◽  
Bud Neck

ABSTRACT Faithful partitioning of genetic material during cell division requires accurate spatial and temporal positioning of nuclei within dividing cells. In Saccharomyces cerevisiae, nuclear positioning is regulated by an elegant interplay between components of the actin and microtubule cytoskeletons. Regulators of this process include Bud6p (also referred to as the actin-interacting protein Aip3p) and Kar9p, which function to promote contacts between cytoplasmic microtubule ends and actin-delimited cortical attachment points. Here, we present the previously undetected association of Bud6p with the cytoplasmic face of yeast spindle pole bodies, the functional equivalent of metazoan centrosomes. Cells lacking Bud6p show exaggerated movements of the nucleus between mother and daughter cells and display reduced amounts of time a given spindle pole body spends in close association with the neck region of budding cells. Furthermore, overexpression of BUD6 greatly enhances interactions between the spindle pole body and mother-bud neck in a spindle alignment-defective dynactin mutant. These results suggest that association of either spindle pole body with neck components, rather than simply entry of a spindle pole body into the daughter cell, provides a positive signal for the progression of mitosis. We propose that Bud6p, through its localization at both spindle pole bodies and at the mother-bud neck, supports this positive signal and provides a regulatory mechanism to prevent excessive oscillations of preanaphase nuclei, thus reducing the likelihood of mitotic delays and nuclear missegregation.

Sign in / Sign up

Export Citation Format

Share Document