Phospholipase Cζ mRNA Expression and its Potency During Spermatogenesis for Activation of Quail Oocyte as a Sperm Factor

This study was conducted to investigate the role of a sperm-borne compound in oocyte activation in special reference to the time when oocyte activation is required by testicular cells during spermatogenesis in quail. First, effects of a microinjection of quail sperm extract (SE) and quail phospholipase C zeta (PLC zeta C) cRNA into quail oocytes were assessed by observation of pronuclear formation and cytoplasmic segmentation, respectively. Secondly, the effects of a microinjection of round spermatids with or without PLC zeta cRNA into quail oocytes were studied by observation of development. When the oocytes were injected with SE at 0.13mg protein/ml, both pronuclear formation and cytoplasmic segmentation were optimally induced. However, pronuclear formation was blocked when SE was pretreated with heat or when the oocyte was pretreated with BAPTA (a Ca2+ chelator) before SE injection. On the other hand, when the oocytes were injected with PLC zeta cRNA at 60 mu g/ml, not only pronuclear formation but also cytoplasmic segmentation were optimally induced. However, PLC zeta cRNA-induced pronuclear formation was blocked by pretreatment with cycloheximide (an inhibitor of protein synthesis) or with BAPTA. Most interestingly, round spermatids alone cannot induce blastodermal development but microinjection of a round spermatid with PLC zeta cRNA can induce development. In addition, RT-PCR revealed that PLC zeta mRNA is expressed in elongated spermatids and testicular sperm but not in round spermatids. It is concluded that PLC zeta is a functional sperm factor for oocyte activation to initiate resumption of meiotic division in quail and its potency is acquired after elongated spermatid formation during the spermatogenesis.