A cyclic adenosine 3′,5′-monophosphate-dependent protein kinase C activation is involved in the hyperactivation of boar spermatozoa
A cyclic adenosine 3′,5′-monophosphate-dependent protein kinase C activation is involved in the hyperactivation of boar spermatozoa
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DOI：
10.1002/mrd.20460
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发表时间：
2006-09-01
影响因子：
2.5
通讯作者：
Miyake, Masashi
中科院分区：
文献类型：
--
作者：
Harayama, Hiroshi;Miyake, Masashi
细胞内cAMP PKA信号通路在哺乳动物精子可复性的表达中起着关键作用。本研究的目的是揭示猪精子中丝氨酸/苏氨酸蛋白激酶在cAMP PKA信号作用下的生物学功能。将射出的精子与cEiMPS（细胞可渗透的cAMP类似物）在38.5 ℃下孵育长达180分钟，然后将它们用于通过蛋白质印迹和间接免疫荧光进行PKC的生化分析，并用于评估鞭毛运动。精子与cBiMPS的孵育逐渐激活连接段中的PKC。精子PKCs的激活伴随着PKA介导的丝氨酸/苏氨酸磷酸化引起的电泳迁移率的变化。与PKC激活相一致，cBiMPS孵育的精子能够表现出鞭毛运动的过度激活。此外，cBiMPS诱导的超活化通过向精子悬浮液中加入特异性PKC抑制剂（Ro-32-0432和双吲哚马来酰亚胺1）而显著抑制。另一方面，使用钙缺乏培养基的实验表明，cBiMPS诱导的鞭毛运动的过度激活和PKC的激活需要细胞外钙。基于所获得的数据，我们得出结论，cAMP-PKA信号可以诱导激活的钙敏感性蛋白激酶C，导致鞭毛运动在公猪精子的超激活。此外，cAMP可能在猪精子的可复性表达过程中作为PKC的上调因子而发挥独特的作用。
An intracellular cAMP-PKA signaling plays a pivotal role in the expression of fertilizing ability in mammalian spermatozoa. The aim of this study is to disclose biological function of serine/ threonine protein kinases that are activated by the action of the cAMP-PKA signaling in boar spermatozoa. Ejaculated spermatozoa were incubated with cEiMPS (a cell-permeable cAMP analog) at 38.5 degrees C up to 180 min, and then they were used for biochemical analyses of PKCs by Western blotting and indirect immunofluorescence and for assessment of flagellar movement. The incubation of spermatozoa with cBiMPS gradually activated PKCs in the connecting piece. The activation of sperm PKCs was accompanied with changes of their electrophoretic mobility by the PKA-mediated serine/threonine phosphorylation. In coincidence with the PKC activation, the cBiMPS-incubated spermatozoa were capable of exhibiting hyperactivation of flagellar movement. Moreover, the cBiMPS-induced hyperactivation was dramatically suppressed by the addition of either of specific PKC inhibitors (Ro-32-0432 and bisindolylmaleimide 1) to the sperm suspensions. On the other hand, experiments using a calcium-deficient medium showed that the cBiMPS-induced hyperactivation of flagellar movement and activation of PKCs required the extracellular calcium. Based on the obtained data, we have concluded that a cAMP-PKA signaling can induce activation of calcium-sensitive PKCs that is leading to the hyperactivation of flagellar movement in boar spermatozoa. Moreover, the cAMP may have a unique role as the up-regulator of PKCs during the expression of fertilizing ability in boar spermatozoa.