The Role Of Clathrin Heavy Chain 1 During Sheep Oocyte Meiotic Maturation And Early Embryo Development

Clathrin is not only a cytosolic protein for vesicular transport to mediate endocytosis,but also has a vital role in stabilizing microtubule and chromosome alignment.Clathrin is a triskelion consisting of three heavy chains each with an associated light chain.Clathrin heavy chain 1(CLTC)is one chain of clathrin and the N-terminal domain of each chain can bind to the mitotic spindle.In this study,we investigated the function of CLTC in sheep oocyte meiotic maturation and early embryo development.CLTC expression was maintained at a high level at sheep oocyte maturation by Western Blot.Although CLTC has no specific localization in the cytoplasm in the germinal vesicle(GV)and germinal vesicle breakdown(GVBD)stages,CLTC is localized in the spindle region in the metaphase Ⅰ(MⅠ)and metaphase Ⅱ(MⅡ)stages.We used treatments by taxol,nocodazole and cold for the relationship between CLTC and β-tubulin(TUBB)in sheep oocyte meiotic maturation,and the treatments did not affect CLTC expression but led to a rearrangement of CLTC.CLTC specific Morpholino(MO)is used to knock down CLTC in the sheep oocyte by microinjection.Knockdown of CLTC disrupted oocyte meiotic spindle assembly and chromosome alignment and impaired first polar body(PB1)emission rate.The subcellular localization of CLTC was examined during sheep early embryo development stages by immunofluorescence staining.Knockdown of CLTC impaired sheep early embryo development in the MII stage.Taken together,our results demonstrate that CLTC plays vital roles in sheep oocyte meiotic maturation and early embryo development.