Abstract

Mammalian oocyte meiotic maturation is the precondition of early embryo development. Lots of microtubules (MT)-associated proteins participate in oocyte maturation process. Cytoskeleton-associated protein 5 (CKAP5) is a member of the XMAP215 family that regulates microtubule dynamics during mitosis. However, its role in meiosis has not been fully studied. Here, we investigated the function of CKAP5 in mouse oocyte meiotic maturation and early embryo development. Western blot showed that CKAP5 expression increased from GVBD, maintaining at high level at metaphase, and decreased after late 1-cell stage. Confocal microscopy showed there is no specific accumulation of CKAP5 at interphase (GV, PN or 2-cell stage). However, once cells enter into meiotic or mitotic division, CKAP5 was localized at the whole spindle apparatus. Treatment of oocytes with the tubulin-disturbing reagents nocodazole (induces MTs depolymerization) or taxol (prevents MTs depolymerization) did not affect CKAP5 expression but led to a rearrangement of CKAP5. Further, knock-down of CKAP5 resulted in a failure of first polar body extrusion, serious defects in spindle assembly, and failure of chromosome alignment. Loss of CKAP5 also decreased early embryo development potential. Furthermore, co-immunoprecipitation showed that CKAP5 bound to clathrin heavy chain 1 (CLTC). Taken together, our results demonstrate that CKAP5 is important in oocyte maturation and early embryo development, and CKAP5 might work together with CLTC in mouse oocyte maturation.

Highlights

  • Spindles are highly dynamic biomechanical structures assembled by microtubules (MTs) during mitosis and meiosis, and they play a crucial role in maintaining genome stability

  • We further demonstrated a relationship between Cytoskeleton-associated protein 5 (CKAP5) and another component of inter-MT bridges, clathrin heavy chain 1 (CLTC), and their role in regulating spindle assembly and chromosome congression

  • Our observation of CKAP5 localization patterns in oocytes differs from reports that other family members, such as Msps in Drosophila and colonic and hepatic tumor overexpressed gene (ch-tumor overexpressed gene (TOG)) in HeLa cells, are mainly localized to the centrosome and contribute to spindle pole integrity and organization in mitotic cells [32, 33]

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Summary

INTRODUCTION

Spindles are highly dynamic biomechanical structures assembled by microtubules (MTs) during mitosis and meiosis, and they play a crucial role in maintaining genome stability. Members in this family include XMAP215 in Xenopus [9], DdCP224 in Dictyostelium [10], STU2 in budding yeast [11], Dis and its second homologue Alp in fission yeast [12, 13], ZYG-9 in Caenorhabditis elegans www.impactjournals.com/oncotarget [14], Msps in Drosophila [15], ch-TOG in humans, and MOR1 in Arabidopsis [16, 17] These proteins have highly conserved structures and are characterized by tumor overexpressed gene (TOG)-containing domains with HEAT motifs in the N-terminal [18, 19]. We investigated the expression and function of CKAP5 during oocyte meiotic maturation and early embryo development and explored the relationship between CKAP5 and CLTC in meiotic spindle assembly and organization

RESULTS
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