Meiotic spindles are critical to ensure chromosome segregation during gamete formation. Oocytes lack centrosomes and use alternative microtubule-nucleation mechanisms for spindle building. How these mechanisms are regulated is still unknown. Aurora kinase A (AURKA) is essential for mouse oocyte meiosis because in pro-metaphase I it triggers microtubule organizing-center fragmentation and its expression compensates for the loss of the two other Aurora kinases (AURKB/AURKC). Although knockout mouse models were useful for foundational studies, AURK spatial and temporal functions are not yet resolved. We provide high-resolution analyses of AURKA/AURKC requirements during meiotic spindle-building and identify the subcellular populations that carry out these functions: 1) AURKA is required in early spindle assembly and later for spindle stability, whereas 2) AURKC is required in late pro-metaphase, and 3) Targeted AURKA constructs expressed in triple AURK knockout oocytes reveal that spindle pole-localized AURKA is the most important population controlling spindle building and stability mechanisms.

• Keywords
• cell biology,
• Publication type
• Journal Article MeSH

OBJECTIVE: Miscarriages affect 10% of women aged 25-29, and 53% of women over 45. The primary cause of miscarriage is aneuploidy that originated in eggs. The Aurora kinase family has three members that regulate chromosome segregation. Therefore, distinguishing the roles of these isoforms is important to understand aneuploidy etiology. In meiosis, Aurora kinase A (AURKA) localizes to spindle poles, where it binds TPX2. Aurora kinase C (AURKC) localizes on chromosomes, where it replaces AURKB as the primary AURK in the chromosomal passenger complex (CPC) via INCENP binding. Although AURKA compensates for CPC function in oocytes lacking AURKB/C, it is unknown whether AURKA binds INCENP in wild type mouse oocytes. ZINC08918027 (ZC) is an inhibitor that prevents the interaction between AURKB and INCENP in mitotic cells. We hypothesized that ZC would block CPC function of any AURK isoform. RESULTS: ZC treatment caused defects in meiotic progression and spindle building. By Western blotting and immunofluorescence, we observed that activated AURKA and AURKC levels in ZC-treated oocytes decreased compared to controls. These results suggest there is a population of AURKA-CPC in mouse oocytes. These data together suggest that INCENP-dependent AURKA and AURKC activities are needed for spindle bipolarity and meiotic progression.

• Keywords
• Aurora kinase, Chromosomal passenger complex, Meiosis, Oocyte,
• MeSH
• Spindle Apparatus metabolism   MeSH
• Aurora Kinase B genetics   metabolism   MeSH
• Meiosis * MeSH
• Mice MeSH
• Oocytes * metabolism   MeSH
• Protein Isoforms genetics   MeSH
• Chromosome Segregation MeSH
• Animals MeSH
• Check Tag
• Mice MeSH
• Female MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Names of Substances
• Aurora Kinase B MeSH
• Protein Isoforms MeSH

The Aurora protein kinases are well-established regulators of spindle building and chromosome segregation in mitotic and meiotic cells. In mouse oocytes, there is significant Aurora kinase A (AURKA) compensatory abilities when the other Aurora kinase homologs are deleted. Whether the other homologs, AURKB or AURKC can compensate for loss of AURKA is not known. Using a conditional mouse oocyte knockout model, we demonstrate that this compensation is not reciprocal because female oocyte-specific knockout mice are sterile, and their oocytes fail to complete meiosis I. In determining AURKA-specific functions, we demonstrate that its first meiotic requirement is to activate Polo-like kinase 1 at acentriolar microtubule organizing centers (aMTOCs; meiotic spindle poles). This activation induces fragmentation of the aMTOCs, a step essential for building a bipolar spindle. We also show that AURKA is required for regulating localization of TACC3, another protein required for spindle building. We conclude that AURKA has multiple functions essential to completing MI that are distinct from AURKB and AURKC.

• MeSH
• Spindle Apparatus genetics   MeSH
• Aurora Kinase A genetics   MeSH
• Aurora Kinase B genetics   MeSH
• Aurora Kinase C genetics   MeSH
• Cell Nucleus Division genetics   MeSH
• Fetal Proteins genetics   MeSH
• Humans MeSH
• Meiosis genetics   MeSH
• Mice MeSH
• Oocytes growth & development   metabolism   MeSH
• Microtubule-Organizing Center metabolism   MeSH
• Polo-Like Kinase 1 MeSH
• Spindle Poles genetics   MeSH
• Protein Serine-Threonine Kinases genetics   MeSH
• Microtubule-Associated Proteins genetics   MeSH
• Cell Cycle Proteins genetics   MeSH
• Proto-Oncogene Proteins genetics   MeSH
• Chromosome Segregation genetics   MeSH
• Gene Expression Regulation, Developmental genetics   MeSH
• Animals MeSH
• Check Tag
• Humans MeSH
• Mice MeSH
• Female MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Research Support, N.I.H., Extramural MeSH
• Names of Substances
• Aurkb protein, mouse MeSH Browser
• Aurkc protein, mouse MeSH Browser
• Aurora Kinase A MeSH
• Aurora Kinase B MeSH
• Aurora Kinase C MeSH
• Fetal Proteins MeSH
• Protein Serine-Threonine Kinases MeSH
• Microtubule-Associated Proteins MeSH
• Cell Cycle Proteins MeSH
• Proto-Oncogene Proteins MeSH
• TACC3 protein, mouse MeSH Browser

Formation of the hatching mouse blastocyst marks the end of preimplantation development, whereby previous cell cleavages culminate in the formation of three distinct cell lineages (trophectoderm, primitive endoderm and epiblast). We report that dysregulated expression of Wwc2, a genetic paralog of Kibra/Wwc1 (a known activator of Hippo-signaling, a key pathway during preimplantation development), is specifically associated with cell autonomous deficits in embryo cell number and cell division abnormalities. Division phenotypes are also observed during mouse oocyte meiotic maturation, as Wwc2 dysregulation blocks progression to the stage of meiosis II metaphase (MII) arrest and is associated with spindle defects and failed Aurora-A kinase (AURKA) activation. Oocyte and embryo cell division defects, each occurring in the absence of centrosomes, are fully reversible by expression of recombinant HA-epitope tagged WWC2, restoring activated oocyte AURKA levels. Additionally, clonal embryonic dysregulation implicates Wwc2 in maintaining the pluripotent epiblast lineage. Thus, Wwc2 is a novel regulator of meiotic and early mitotic cell divisions, and mouse blastocyst cell fate.

• Keywords
• blastocyst cell number, cell division, cell lineage decision, cell-fate, oocyte maturation, preimplantation mouse embryo,
• Publication type
• Journal Article MeSH

Homologous chromosome segregation during meiosis I (MI) in mammalian oocytes is carried out by the acentrosomal MI spindles. Whereas studies in human oocytes identified Ran GTPase as a crucial regulator of the MI spindle function, experiments in mouse oocytes questioned the generality of this notion. Here, we use live-cell imaging with fluorescent probes and Förster resonance energy transfer (FRET) biosensors to monitor the changes in Ran and importin β signaling induced by perturbations of Ran in mouse oocytes while examining the MI spindle dynamics. We show that unlike RanT24N employed in previous studies, a RanT24N, T42A double mutant inhibits RanGEF without perturbing cargo binding to importin β and disrupts MI spindle function in chromosome segregation. Roles of Ran and importin β in the coalescence of microtubule organizing centers (MTOCs) and MI spindle assembly are further supported by the use of the chemical inhibitor importazole, whose effects are partially rescued by the GTP hydrolysis-resistant RanQ69L mutant. These results indicate that RanGTP is essential for MI spindle assembly and function both in humans and mice.

• Keywords
• RanGTP, importazole, importin β, meiosis I, oocyte,
• MeSH
• Spindle Apparatus physiology   MeSH
• beta Karyopherins genetics   metabolism   MeSH
• Nuclear Proteins genetics   metabolism   MeSH
• Meiosis physiology   MeSH
• Microtubules metabolism   MeSH
• Mutation MeSH
• Mice MeSH
• Oocytes cytology   metabolism   MeSH
• Cell Cycle Proteins genetics   metabolism   MeSH
• ran GTP-Binding Protein genetics   metabolism   MeSH
• Chromosome Segregation MeSH
• Guanine Nucleotide Exchange Factors genetics   metabolism   MeSH
• Animals MeSH
• Check Tag
• Mice MeSH
• Female MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Research Support, N.I.H., Extramural MeSH
• Names of Substances
• beta Karyopherins MeSH
• Nuclear Proteins MeSH
• Cell Cycle Proteins MeSH
• ran GTP-Binding Protein MeSH
• Rcc1 protein, mouse MeSH Browser
• Guanine Nucleotide Exchange Factors MeSH