scholarly journals C. elegans CLASP/CLS-2 negatively regulates membrane ingression throughout the oocyte cortex and is required for polar body extrusion

2020 ◽  
Author(s):  
Aleesa J. Schlientz ◽  
Bruce Bowerman
Keyword(s):  
Polar Body ◽  
Spindle Assembly ◽  
Meiotic Spindle ◽  
Contractile Apparatus ◽  
Contractile Ring ◽  
C Elegans ◽  
Cell Divisions ◽  
Polar Bodies ◽  
Polar Body Extrusion ◽  
The Relationship

AbstractThe requirements for oocyte meiotic cytokinesis during polar body extrusion are not well understood. In particular, the relationship between the oocyte meiotic spindle and polar body contractile ring dynamics remains largely unknown. We have used live cell imaging and spindle assembly defective mutants lacking the function of CLASP/CLS-2, kinesin-12/KLP-18, or katanin/MEI-1 to investigate the relationship between meiotic spindle structure and polar body extrusion in C. elegans oocytes. We show that spindle bipolarity and chromosome segregation are not required for polar body contractile ring formation and chromosome extrusion in klp-18 mutants, but oocytes with severe spindle assembly defects due to loss of CLS-2 or MEI-1 have penetrant and distinct polar body extrusion defects: CLS-2 is required early for contractile ring assembly or stability, while MEI-1 is required later for contractile ring constriction. We also show that CLS-2 negatively regulates membrane ingression throughout the oocyte cortex during meiosis I, and we explore the relationship between global cortical dynamics and oocyte meiotic cytokinesis.Author SummaryThe precursor cells that produce gametes—sperm and eggs in animals—have two copies of each chromosome, one from each parent. These precursors undergo specialized cell divisions that leave each gamete with only one copy of each chromosome; defects that produce incorrect chromosome number cause severe developmental abnormalities. In oocytes, these cell divisions are highly asymmetric, with extra chromosomes discarded into small membrane bound polar bodies, leaving one chromosome set within the much larger oocyte. How oocytes assemble the contractile apparatus that pinches off polar bodies remains poorly understood. To better understand this process, we have used the nematode Caenorhabditis elegans to investigate the relationship between the bipolar structure that separates oocyte chromosomes, called the spindle, and assembly of the contractile apparatus that pinches off polar bodies. We used a comparative approach, examining this relationship in three spindle assembly defective mutants. Bipolar spindle assembly and chromosome separation were not required for polar body extrusion, as it occurred normally in mutants lacking a protein called KLP-18. However, mutants lacking the protein CLS-2 failed to assemble the contractile apparatus, while mutants lacking the protein MEI-1 assembled a contractile apparatus that failed to fully constrict. We also found that CLS-2 down-regulates membrane ingression throughout the oocyte surface, and we explored the relationship between oocyte membrane dynamics and polar body extrusion.

scholarly journals ALADIN is Required for the Production of Fertile Mouse Oocytes.

2016 ◽  
Author(s):  
Sara Carvalhal ◽  
Michelle Stevense ◽  
Katrin Koehler ◽  
Ronald Naumann ◽  
Angela Huebner ◽  
...  
Keyword(s):  
Polar Body ◽  
Spindle Assembly ◽  
Cell Periphery ◽  
Cell Stage ◽  
Spindle Positioning ◽  
Asymmetric Cell Divisions ◽  
Cell Divisions ◽  
Polar Bodies ◽  
Polar Body Extrusion

Asymmetric cell divisions depend upon the precise placement of the mitotic spindle. In mammalian oocytes, spindles assemble close to the cell center but chromosome segregation takes place at the cell periphery where half of the chromosomes are expelled into small, non-developing polar bodies at anaphases. By dividing so asymmetrically, most of the cytoplasmic content within the oocyte is preserved, which is critical for successful fertilization and early development. Recently, we determined that the nucleoporin ALADIN participates in spindle assembly in somatic cells, and we have also shown that female mice homozygous deficient for ALADIN are sterile. In this study we show that this protein is involved in specific meiotic stages including meiotic resumption, spindle assembly, and spindle positioning. In the absence of ALADIN, polar body extrusion is impaired in a majority of oocytes due to problems in spindle orientation prior to the first meiotic anaphase. Those few oocytes that can mature far enough to be fertilized in vitro are unable to support embryonic development beyond the two-cell stage. Overall, we find that ALADIN is critical for oocyte maturation and appears to be far more essential for this process than for somatic cell divisions.

scholarly journals ALADIN is required for the production of fertile mouse oocytes

2017 ◽  
Vol 28 (19) ◽  
pp. 2470-2478 ◽  
Author(s):  
Sara Carvalhal ◽  
Michelle Stevense ◽  
Katrin Koehler ◽  
Ronald Naumann ◽  
Angela Huebner ◽  
...  
Keyword(s):  
Polar Body ◽  
Spindle Assembly ◽  
Cell Periphery ◽  
Cell Stage ◽  
Spindle Positioning ◽  
Asymmetric Cell Divisions ◽  
Cell Divisions ◽  
Polar Bodies ◽  
Polar Body Extrusion

Asymmetric cell divisions depend on the precise placement of the spindle apparatus. In mammalian oocytes, spindles assemble close to the cell’s center, but chromosome segregation takes place at the cell periphery where half of the chromosomes are expelled into small, nondeveloping polar bodies at anaphase. By dividing so asymmetrically, most of the cytoplasmic content within the oocyte is preserved, which is critical for successful fertilization and early development. Recently we determined that the nucleoporin ALADIN participates in spindle assembly in somatic cells, and we have also shown that female mice homozygously null for ALADIN are sterile. In this study we show that this protein is involved in specific meiotic stages, including meiotic resumption, spindle assembly, and spindle positioning. In the absence of ALADIN, polar body extrusion is compromised due to problems in spindle orientation and anchoring at the first meiotic anaphase. ALADIN null oocytes that mature far enough to be fertilized in vitro are unable to support embryonic development beyond the two-cell stage. Overall, we find that ALADIN is critical for oocyte maturation and appears to be far more essential for this process than for somatic cell divisions.

scholarly journals A casein kinase 1 prevents expulsion of the oocyte meiotic spindle into a polar body by regulating cortical contractility

2017 ◽  
Vol 28 (18) ◽  
pp. 2410-2419 ◽  
Author(s):  
Jonathan R. Flynn ◽  
Francis J. McNally
Keyword(s):  
Polar Body ◽  
Spindle Pole ◽  
Casein Kinase ◽  
Meiotic Spindle ◽  
Contractile Ring ◽  
Direct Inhibition ◽  
Casein Kinase 1 ◽  
Asymmetric Cell Divisions ◽  
Polar Bodies ◽  
Cortical Contractility

During female meiosis, haploid eggs are generated from diploid oocytes. This reduction in chromosome number occurs through two highly asymmetric cell divisions, resulting in one large egg and two small polar bodies. Unlike mitosis, where an actomyosin contractile ring forms between the sets of segregating chromosomes, the meiotic contractile ring forms on the cortex adjacent to one spindle pole, then ingresses down the length of the spindle to position itself at the exact midpoint between the two sets of segregating chromosomes. Depletion of casein kinase 1 gamma (CSNK-1) in Caenorhabditis elegans led to the formation of large polar bodies that contain all maternal DNA, because the contractile ring ingressed past the spindle midpoint. Depletion of CSNK-1 also resulted in the formation of deep membrane invaginations during meiosis, suggesting an effect on cortical myosin. Both myosin and anillin assemble into dynamic rho-dependent cortical patches that rapidly disassemble in wild-type embryos. CSNK-1 was required for disassembly of both myosin patches and anillin patches. Disassembly of anillin patches was myosin independent, suggesting that CSNK-1 prevents expulsion of the entire meiotic spindle into a polar body by negatively regulating the rho pathway rather than through direct inhibition of myosin.

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.

scholarly journals Kinetochore Fibers Are Not Involved in the Formation of the First Meiotic Spindle in Mouse Oocytes, but Control the Exit from the First Meiotic M Phase

1999 ◽  
Vol 146 (1) ◽  
pp. 1-12 ◽  
Author(s):  
Stéphane Brunet ◽  
Angélica Santa Maria ◽  
Philippe Guillaud ◽  
Denis Dujardin ◽  
Jacek Z. Kubiak ◽  
...  
Keyword(s):  
Polar Body ◽  
Meiotic Spindle ◽  
Homologous Chromosomes ◽  
Mouse Oocytes ◽  
Sister Chromatids ◽  
M Phase ◽  
Continuous Presence ◽  
Set Up ◽  
Polar Body Extrusion ◽  
Reductional Division

During meiosis, two successive divisions occur without any intermediate S phase to produce haploid gametes. The first meiotic division is unique in that homologous chromosomes are segregated while the cohesion between sister chromatids is maintained, resulting in a reductional division. Moreover, the duration of the first meiotic M phase is usually prolonged when compared with mitotic M phases lasting 8 h in mouse oocytes. We investigated the spindle assembly pathway and its role in the progression of the first meiotic M phase in mouse oocytes. During the first 4 h, a bipolar spindle forms and the chromosomes congress near the equatorial plane of the spindle without stable kinetochore– microtubule end interactions. This late prometaphase spindle is then maintained for 4 h with chromosomes oscillating in the central region of the spindle. The kinetochore–microtubule end interactions are set up at the end of the first meiotic M phase (8 h after entry into M phase). This event allows the final alignment of the chromosomes and exit from metaphase. The continuous presence of the prometaphase spindle is not required for progression of the first meiotic M phase. Finally, the ability of kinetochores to interact with microtubules is acquired at the end of the first meiotic M phase and determines the timing of polar body extrusion.

scholarly journals OLA1 is responsible for normal spindle assembly and SAC activation in mouse oocytes

2019 ◽  
Author(s):  
Di Xie ◽  
Juan Zhang ◽  
JinLi Ding ◽  
Jing Yang ◽  
Yan Zhang
Keyword(s):  
Spindle Assembly Checkpoint ◽  
Polar Body ◽  
Germinal Vesicle ◽  
Spindle Assembly ◽  
Mouse Oocyte ◽  
Immunofluorescent Staining ◽  
Germinal Vesicle Breakdown ◽  
Oocyte Meiosis ◽  
Polar Body Extrusion ◽  
Spread Analysis

Background. OLA1 is a member of the GTPase protein family, unlike other members, it can bind and hydrolyze ATP more efficiently than GTP. OLA1 participates in cell proliferation, oxidative response and tumorigenesis. However, whether OLA1 is also required for oocyte meiosis is still unknown. Methods. In this study, the localization, expression, and functions of OLA1 in the mouse oocyte meiosis were examined. Immunofluorescent and confocal microscopy were used to explore the location pattern of OLA1 in the mouse oocyte. Moreover, nocodazole treatment was used to confirm the spindle-like location of OLA1 during mouse meiosis. Western blot was used to explore the expression pattern of OLA1 in the mouse oocyte. Microinjection of siRNA was used to explore the OLA1 functions in the mouse oocyte meiosis. In addition, chromosome spreading was used to investigate the spindle assembly checkpoint (SAC) activity. Results. Immunofluorescent staining showed that OLA1 evenly distributed in the cytoplasm at germinal vesicle (GV) stage. After meiosis resumption (GVBD), OLA1 co-localized with spindles, which was further identified by nocodazole treatment experiments. Knockdown of OLA1 impaired the germinal vesicle breakdown progression and finally resulted in a lower polar body extrusion rate. Immunofluorescence analysis indicated that knockdown of OLA1 led to abnormal spindle assembly, which was evidenced by multipolar spindles in OLA1-RNAi-oocytes. After 6 h post-GVBD in culture, an increased proportion of oocyte which has precociously entered into anaphase/telephase I (A/TI) was observed in OLA1-knockdown oocytes, suggesting that loss of OLA1 resulted in the premature segregation of homologous chromosomes. In addition, the chromosome spread analysis suggested that OLA1 knockdown induced premature anaphase onset was due to the precocious inactivation of SAC. Taken together, we concluded that OLA1 plays important role in GVBD, spindle assembly and SAC activation maintenance in oocyte meiosis.

scholarly journals Sequential actin-based pushing forces drive meiosis I chromosome migration and symmetry breaking in oocytes

2013 ◽  
Vol 200 (5) ◽  
pp. 567-576 ◽  
Author(s):  
Kexi Yi ◽  
Boris Rubinstein ◽  
Jay R. Unruh ◽  
Fengli Guo ◽  
Brian D. Slaughter ◽  
...  
Keyword(s):  
Symmetry Breaking ◽  
Cytoplasmic Streaming ◽  
Feedback Loop ◽  
Polar Body ◽  
Meiotic Spindle ◽  
Directed Motion ◽  
Family Protein ◽  
Polar Body Extrusion ◽  
Spindle Position ◽  
Meiosis I

Polar body extrusion during oocyte maturation is critically dependent on asymmetric positioning of the meiotic spindle, which is established through migration of the meiosis I (MI) spindle/chromosomes from the oocyte interior to a subcortical location. In this study, we show that MI chromosome migration is biphasic and driven by consecutive actin-based pushing forces regulated by two actin nucleators, Fmn2, a formin family protein, and the Arp2/3 complex. Fmn2 was recruited to endoplasmic reticulum structures surrounding the MI spindle, where it nucleated actin filaments to initiate an initially slow and poorly directed motion of the spindle away from the cell center. A fast and highly directed second migration phase was driven by actin-mediated cytoplasmic streaming and occurred as the chromosomes reach a sufficient proximity to the cortex to activate the Arp2/3 complex. We propose that decisive symmetry breaking in mouse oocytes results from Fmn2-mediated perturbation of spindle position and the positive feedback loop between chromosome signal-induced Arp2/3 activation and Arp2/3-orchestrated cytoplasmic streaming that transports the chromosomes.

scholarly journals Assembly and Positioning of the Oocyte Meiotic Spindle

2018 ◽  
Vol 34 (1) ◽  
pp. 381-403 ◽  
Author(s):  
Binyam Mogessie ◽  
Kathleen Scheffler ◽  
Melina Schuh
Keyword(s):  
Menstrual Cycle ◽  
Recent Work ◽  
Meiotic Division ◽  
Polar Body ◽  
Mitotic Division ◽  
Model System ◽  
Small Cell ◽  
Meiotic Spindle ◽  
Polar Body Extrusion ◽  
Shed Light

Fertilizable eggs develop from diploid precursor cells termed oocytes. Once every menstrual cycle, an oocyte matures into a fertilizable egg in the ovary. To this end, the oocyte eliminates half of its chromosomes into a small cell termed a polar body. The egg is then released into the Fallopian tube, where it can be fertilized. Upon fertilization, the egg completes the second meiotic division, and the mitotic division of the embryo starts. This review highlights recent work that has shed light on the cytoskeletal structures that drive the meiotic divisions of the oocyte in mammals. In particular, we focus on how mammalian oocytes assemble a microtubule spindle in the absence of centrosomes, how they position the spindle in preparation for polar body extrusion, and how the spindle segregates the chromosomes. We primarily focus on mouse oocytes as a model system but also highlight recent insights from human oocytes.

scholarly journals Acentrosomal spindle assembly and stability in C. elegans oocytes requires a kinesin-12 non-motor microtubule interaction domain

2021 ◽  
Author(s):  
Ian Daniel Wolff ◽  
Jeremy Alden Hollis ◽  
Sarah Marie Wignall
Keyword(s):  
Adaptor Protein ◽  
Direct Interaction ◽  
Spindle Assembly ◽  
Meiotic Spindle ◽  
Interaction Domain ◽  
Microtubule Binding ◽  
C Elegans ◽  
Insight Into

During the meiotic divisions in oocytes, microtubules are sorted and organized by motor proteins to generate a bipolar spindle in the absence of centrosomes. In most organisms, kinesin-5 family members crosslink and slide microtubules to generate outward force that promotes acentrosomal spindle bipolarity. However, the mechanistic basis for how other kinesin families act on acentrosomal spindles has not been explored. We investigated this question in C. elegans oocytes, where kinesin-5 is not required to generate outward force. Instead, the kinesin-12 family motor KLP-18 performs this function. KLP-18 acts with adaptor protein MESP-1 (meiotic spindle 1) to sort microtubule minus ends to the periphery of a microtubule array, where they coalesce into spindle poles. If either of these proteins is depleted, outward sorting of microtubules is lost and minus ends converge to form a monoaster. Here we use a combination of in vitro biochemical assays and in vivo mutant analysis to provide insight into the mechanism by which these proteins collaborate to promote acentrosomal spindle assembly. We identify a microtubule binding site on the C-terminal stalk of KLP-18 and demonstrate that a direct interaction between the KLP-18 stalk and MESP-1 activates non-motor microtubule binding. We also provide evidence that this C-terminal domain is required for KLP-18 activity during spindle assembly and show that KLP-18 is continuously required to maintain spindle bipolarity. This study thus provides new insight into the construction and maintenance of the oocyte acentrosomal spindle as well as into kinesin-12 mechanism and regulation.

Sign in / Sign up

Export Citation Format

Share Document