The Role Of Cytoskeleton-Associated Protein 5 During Mouse Oocyte Meiotic Maturation And Early Embryo Development

Cytoskeleton-Associated Protein 5 (CKAP5) belongs to the XMAP215 (Xenopus Microtubule-Associated Protein 215) family of MAPs (Microtubule-Associated Proteins) which regulates microtubule dynamics during mitosis. However, less is known about its role in mouse oocyte meiotic maturation and early embryo development. We employed qRT-PCR, western blot and immunofluorescence assay to examine CKAP5 mRNA, protein expression and subcellular localization in different stages (GV, GVBD, MI, AI-TI, MII, AII-TII, PN, late 1-cell and 2-cell). Both CKAP5 mRNA and protein were found from GV to the 2-cell stage and the mRNA expression pattern was similar to that of protein expression. CKAP5 expression levels were low at the GV and GVBD stage, and increased at MI stage followed by relatively highest abundance from AI-TI to PN stage. Then a decline occurred at late 1-cell and 2-cell stage. Immunostaining results showed CKAP5 localized at the spindle area and its distribution displayed in a cell cycle-dependent manner. The depletion of CKAP5 caused the failure of the first polar body (PB1) extrusion and severe defects in spindle assembly and chromosome alignment. Moreover, the lack of CKAP5 resulted in dramatic decrease of embryo development rate and cleavage, thus impairing the embryo developmental potential. To identify the relationship between CKAP5 and microtubule, oocytes were treated with spindle-perturbing agents nocodazole or taxol to alter microtubule dynamics. These two drugs did not affect the protein level, but rearranged the distribution of CKAP5. In addition, the correlation between CKAP5 and CLTC (Clathrin Heavy Chain 1) was evaluated. They could co-immunoprecipitated from oocytes extract. Depletion of CKAP5 caused mislocalization of CLTC and vice versa. Taken together, our study demonstrate that CKAP5 is involved in spindle assembly and chromosome congression through regulating spindle microtubule dynamics in mouse oocyte maturation and early embryo development.