Intracellular Ca2+ increase induces post-fertilization events via MAP kinase dephosphorylation in eggs of the hydrozoan jellyfish Cladonema pacificum
Intracellular Ca2+ increase induces post-fertilization events via MAP kinase dephosphorylation in eggs of the hydrozoan jellyfish Cladonema pacificum
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DOI：
10.1016/j.ydbio.2006.02.005
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发表时间：
2006-05-01
影响因子：
2.7
通讯作者：
Deguchi, R
中科院分区：
文献类型：
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作者：
Kondoh, E;Tachibana, K;Deguchi, R
太平洋枝藻水母的自然产卵停留在类G(1)原核阶段，直到受精。Cladonema的受精卵经历了一系列受精后的事件，包括精子吸引能力的丧失，卵子表面黏附物质的表达，以及导致DNA合成和分裂的细胞周期的启动。在这里，我们研究这些事件是否受细胞内钙离子浓度和有丝分裂原激活蛋白激酶(MAP)活性变化的调节。我们发现MAP激酶在未受精卵中以磷酸化的形式存在。精子诱导的钙升高是受精的第一个迹象，在受精后几分钟内，MAP激酶立即去磷酸化。受精卵通常会在5分钟后停止吸引精子，并在这段时间变得粘稠。在类似于受精后1h，它们进一步进行胞质分裂，产生2-细胞胚胎，在此之前，DNA合成以BrdU掺入细胞核为证据。将肌醇1，4，5-三磷酸(IP3)注射到未受精卵中，产生与受精时相似的钙离子增加，触发了MAPK去磷酸化和上述受精后的事件。反之，受精卵内注射BAPTA/Ca~(2+)浓度约为10~(-10)S时，Ca~(2+)浓度升高，立即降低升高的Ca~(2+)水平，并抑制随后的受精后事件。用MAT激酶(MEK)抑制剂U0126处理后，在未受精卵中触发了受精后事件，MAP激酶去磷酸化而不是钙离子增加。相反，预先注射维持MAP激酶磷酸化状态的谷胱甘肽S转移酶融合蛋白(MOS)，可以阻止受精卵中受精后的事件，但不能阻止钙离子的增加。这些结果有力地表明，受精后的三个事件，即精子吸引力的停止、表面黏附的表达和细胞周期的进展，都位于由Ca~(2+)增加触发的MAP激酶去磷酸化的下游。(C)2006 Elsevier Inc.保留所有权利。
Naturally spawned eggs of the hydrozoan jellyfish Cladonema pacificum are arrested at G(1)-like pronuclear stage until fertilization. Fertilized eggs of Cladonema undergo a series of post-fertilization events, including loss of sperm-attracting ability, expression of adhesive materials on the egg surface, and initiation of cell cycle leading to DNA synthesis and cleavage. Here, we investigate whether these events are regulated by changes in intracellular Ca2+ concentration and mitogen-activated protein kinase (MAP kinase) activity in Cladonema eggs. We found that MAP kinase is maintained in the phosphorylated form in unfertilized eggs. Initiation of sperm-induced Ca2+ increase, which is the first sign of fertilization, was immediately followed by MAP kinase dephosphorylation within a few minutes of fertilization. The fertilized eggs typically stopped sperm attraction by an additional 5 min and became sticky around this time. They further underwent cytokinesis yielding 2-cell embryos at similar to 1 h post-fertilization, which was preceded by DNA synthesis evidenced by BrdU incorporation into the nuclei. Injection of inositol 1,4,5-trisphosphate (IP3) into unfertilized eggs, which produced a Ca2+ increase similar to that seen at fertilization, triggered MAP kinase dephosphorylation and the above post-fertilization events without insemination. Conversely, injection of BAPTA/Ca2+ into fertilized eggs at similar to 10 s after the initiation of Ca2+ increase immediately lowered the elevating Ca2+ level and inhibited the subsequent post-fertilization events. Treatment with U0126, an inhibitor of MAT kinase kinase (MEK), triggered the post-fertilization events in unfertilized eggs, where MAP kinase dephosphorylation but not Ca2+ increase was generated. Conversely, preinjection of the glutathione S-transferase (GST) fusion protein of MAP kinase kinase kinase (Mos), which maintained the phosphorylated state of MAP kinase, blocked the post-fertilization events in fertilized eggs without preventing a Ca2+ increase. These results strongly suggest that all of the three post-fertilization events, cessation of sperm attraction, expression of surface adhesion, and progression of cell cycle, lie downstream of MAP kinase dephosphorylation that is triggered by a Ca2+ increase. (c) 2006 Elsevier Inc. All rights reserved.