Jones K T, Matsuda M, Parrington J, Katan M, Swann K
Department of Anatomy, University College, Gower Street, London, WC1E 6BT, UK.
Biochem J. 2000 Mar 15;346 Pt 3(Pt 3):743-9.
A soluble phospholipase C (PLC) from boar sperm generates InsP(3) and hence causes Ca(2+) release when added to sea urchin egg homogenate. This PLC activity is associated with the ability of sperm extracts to cause Ca(2+) oscillations in mammalian eggs following fractionation. A sperm PLC may, therefore, be responsible for causing the observed Ca(2+) oscillations at fertilization. In the present study we have further characterized this boar sperm PLC activity using sea urchin egg homogenate. Consistent with a sperm PLC acting on egg PtdIns(4,5)P(2), the ability of sperm extracts to release Ca(2+) was blocked by preincubation with the PLC inhibitor U73122 or by the addition of neomycin to the homogenate. The Ca(2+)-releasing activity was also detectable in sperm from other species and in whole testis extracts. However, activity was not observed in extracts from other tissues. Moreover recombinant PLCbeta1, -gamma1, -gamma2, -delta1, all of which had higher specific activities than boar sperm extracts, were not able to release Ca(2+) in the sea urchin egg homogenate. In addition these PLCs were not able to cause Ca(2+) oscillations following microinjection into mouse eggs. These results imply that the sperm PLC possesses distinct properties that allow it to hydrolyse PtdIns(4,5)P(2) in eggs.
公猪精子中的一种可溶性磷脂酶C（PLC）能产生肌醇三磷酸（InsP(3)），因此添加到海胆卵匀浆中时会引起钙离子（Ca(2+)）释放。这种PLC活性与精子提取物在分级分离后引起哺乳动物卵中钙离子振荡的能力相关。因此，精子PLC可能是受精时观察到的钙离子振荡的原因。在本研究中，我们使用海胆卵匀浆进一步表征了这种公猪精子PLC活性。与作用于卵磷脂酰肌醇-4,5-二磷酸（PtdIns(4,5)P(2)）的精子PLC一致，精子提取物释放钙离子的能力可通过与PLC抑制剂U73122预孵育或向匀浆中添加新霉素来阻断。在其他物种的精子和整个睾丸提取物中也可检测到钙离子释放活性。然而，在其他组织的提取物中未观察到活性。此外，重组PLCβ1、-γ1、-γ2、-δ1，其比公猪精子提取物具有更高的比活性，但均不能在海胆卵匀浆中释放钙离子。此外，将这些PLC显微注射到小鼠卵中后也不能引起钙离子振荡。这些结果表明，精子PLC具有独特的特性，使其能够水解卵中的PtdIns(4,5)P(2)。
