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Chromokinesin Kif4 promotes proper anaphase in mouse oocyte meiosis. Heath CM et al. (2019) Oocytes of many species lack centrioles, and therefore form acentriolar spindles. Despite the necessity of oocyte meiosis for successful reproduction, how these spindles mediate accurate chromosome segregation is poorly understood. We have gained insight into this process through studies of the kinesin-4 family member Kif4 in mouse oocytes. We found that Kif4 localizes to chromosomes through metaphase and then largely re-distributes to the spindle midzone during anaphase, transitioning from stretches along microtubules to distinct ring-like structures; these structures then appear to fuse together by telophase. Kif4's binding partner PRC1 and MgcRacGAP, a component of the centralspindlin complex, have a similar localization pattern, demonstrating dynamic spindle midzone organization in oocytes. Kif4 knockdown results in defective midzone formation and longer spindles, revealing new anaphase roles for Kif4 in mouse oocytes. Moreover, inhibition of Aurora B/C kinases results in Kif4 mislocalization and causes anaphase defects. Taken together, our work reveals essential roles for Kif4 during the meiotic divisions, furthering our understanding of mechanisms promoting accurate chromosome segregation in acentriolar oocytes. Movie S1 Movie S1 Movie S1 is a z-stack animation of an Anaphase I spindle in side view, with a midzone in the "ring stage" labeled with Kif4 (red), microtubules (green), DNA (blue) and corresponds to Figure 4A, row 1. Microtubule channel blinks in and out for clarity. Bar = 5 μm. Movie S2 Movie S2 Movie S2 is a z-stack animation of an Anaphase II spindle in end-on view, with a midzone in the "ring stage" labeled with Kif4 (red), microtubules (green), and DNA (blue). Bar = 5 μm. Movie S3 Movie S3 Movie S3 is a z-stack animation of an Anaphase I spindle in side view, with a midzone in the "plate stage" labeled with Kif4 (red), microtubules (green), and DNA (blue) and corresponds to Figure 4A, row 2. Bar = 5 μm. Movie S4 Movie S4 Movie S4 is a z-stack animation of an Anaphase I spindle in an angled view, with a midzone in the "plate stage" labeled with Kif4 (red), microtubules (green), and DNA (blue). Bar = 5 μm. Movie S5 Movie S5 Movie S5 is a z-stack animation of an Anaphase I spindle in side view, with a midzone in the "transition stage" labeled with Kif4 (red), microtubules (green), and DNA (blue) and corresponds to Figure 4B. Microtubule channel blinks in and out for clarity. Bar = 5 μm. Movie S6 Movie S6 Movie S6 is a z-stack animation of an Anaphase I spindle in end-on view, with a midzone in the "ring stage" labeled with MgcRacGAP (red), microtubules (green), and DNA (blue) and corresponds to Figure 5B, row 1. Microtubule channel blinks in and out for clarity. Bar = 5 μm. Movie S7 Movie S7 Movie S7 is a z-stack animation of an Anaphase II spindle in side view, with a midzone in the "transition stage" labeled with PRC1 (red), microtubules (green), DNA (blue) and corresponds to Supplemental Figure 5A, row 1. Microtubule channel blinks in and out for clarity. Bar = 5 μm.//////////////////
Involvement of Kif4a in Spindle Formation and Chromosome Segregation in Mouse Oocytes. Tang F et al. (2018) Kif4a, a member of the kinesin superfamily, has been reported to participate in a series of cellular processes such as chromosome condensation and cytokinesis during mitosis. However, the roles of KIF4a in meiosis are still unknown. In present study we found that the Kif4a protein expression decreased in maternal aged mouse oocytes. We then explored the roles of Kif4a in mouse oocyte meiosis by knockdown analysis. Kif4a was enriched at the spindle during mouse oocyte maturation. By specific knock down of the Kif4a using morpholino microinjection, we found that the disruption of Kif4a caused the failure of polar body extrusion. Further analysis indicated that Kif4a might affect the spindle morphology and chromosome alignment in the mouse oocytes, and this might be due to the regulation of tubulin acetylation. Moreover, our results showed that an increased proportion of aneuploidy in the Kif4a knock down oocytes, and this might be due to the loss of kinetochore-microtubule attachment. Taken together, these results suggested that Kif4a possibly regulated mouse oocyte meiosis through its effects on the spindle organization and accurate chromosome segregation, and the loss of Kif4a might be related with aneuploidy of aging oocytes.//////////////////
Kif4 Is Essential for Mouse Oocyte Meiosis. Camlin NJ et al. (2017) Progression through the meiotic cell cycle must be strictly regulated in oocytes to generate viable embryos and offspring. During mitosis, the kinesin motor protein Kif4 is indispensable for chromosome condensation and separation, midzone formation and cytokinesis. Additionally, the bioactivity of Kif4 is dependent on phosphorylation via Aurora Kinase B and Cdk1, which regulate Kif4 function throughout mitosis. Here, we examine the role of Kif4 in mammalian oocyte meiosis. Kif4 localized in the cytoplasm throughout meiosis I and II, but was also observed to have a dynamic subcellular distribution, associating with both microtubules and kinetochores at different stages of development. Co-localization and proximity ligation assays revealed that the kinetochore proteins, CENP-C and Ndc80, are potential Kif4 interacting proteins. Functional analysis of Kif4 in oocytes via antisense knock-down demonstrated that this protein was not essential for meiosis I completion. However, Kif4 depleted oocytes displayed enlarged polar bodies and abnormal metaphase II spindles, indicating an essential role for this protein for correct asymmetric cell division in meiosis I. Further investigation of the phosphoregulation of meiotic Kif4 revealed that Aurora Kinase and Cdk activity is critical for Kif4 kinetochore localization and interaction with Ndc80 and CENP-C. Finally, Kif4 protein but not gene expression was found to be upregulated with age, suggesting a role for this protein in the decline of oocyte quality with age.//////////////////
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