Ca2+ oscillation-inducing phospholipase C zeta expressed in mouse eggs is accumulated to the pronucleus during egg activation.
Ca2+ oscillation-inducing phospholipase C zeta expressed in mouse eggs is accumulated to the pronucleus during egg activation.
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DOI：
10.1016/j.ydbio.2003.12.028
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发表时间：
2004-04
影响因子：
2.7
通讯作者：
A. Yoda;S. Oda;T. Shikano;Z. Kouchi;T. Awaji;H. Shirakawa;K. Kinoshita;S. Miyazaki
中科院分区：
文献类型：
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作者：
A. Yoda;S. Oda;T. Shikano;Z. Kouchi;T. Awaji;H. Shirakawa;K. Kinoshita;S. Miyazaki
精子特异性磷脂酶czeta （PLCζ）已知通过注射编码PLCζ的RNA在小鼠卵子中表达时，可诱导细胞内Ca2+振荡和卵子活化。我们通过监测与PLCζ融合的黄色荧光蛋白“金星”的荧光，实时研究了PLCζ在卵子中的表达水平和空间分布，以及与Ca2+振荡的起始和终止有关。在RNA注射后40-50 min，表达的PLCζ达到10 - 40 × 10 - 15g时，诱导与受精时相似的Ca2+振荡。plc - ζ - venus在3 h时增加，在4-5 h时达到稳定水平。有趣的是，plc - ζ - venus在5-6 h时积累到原核（PN），并在那里持续增加。在开始积累之前，Ca2+振荡在大多数卵子中停止。缺乏三个EF手结构域的PLCζ变体在诱导Ca2+振荡方面的效果要差得多，并且在原核中很少积累，表明这些结构域的关键作用。聚积到原核的能力使PLCζ成为Ca2+振荡诱导精子因子的强候选物，该因子在精卵融合时被引入卵浆，在诱导卵子激活后被浓缩到原核。
Sperm-specific phospholipase C zeta (PLCζ) is known to induce intracellular Ca2+oscillations and egg activation when expressed in mouse eggs by injection of RNA encoding PLCζ. We investigated the expression level and spatial distribution of PLCζ in the egg in real time and in relation to the initiation and termination of Ca2+oscillations by monitoring fluorescence of a yellow fluorescent protein ‘Venus’ fused with PLCζ. Ca2+oscillations similar to those at fertilization were induced at 40–50 min after RNA injection, when expressed PLCζ reached 10–40 × 10−15g in the egg. PLCζ–Venus increased up to 3 h and attained a steady level at 4–5 h. Interestingly, PLCζ–Venus is accumulated to the pronucleus (PN) formed at 5–6 h and continuously increased there. Ca2+oscillations stopped in most eggs before initiation of the accumulation. A variant of PLCζ that lacks three EF hand domains was much less effective in induction of Ca2+oscillations and little accumulated in the pronucleus, indicating a critical role of those domains. The ability of the accumulation to the pronucleus qualifies PLCζ for a strong candidate of the Ca2+oscillation-inducing sperm factor, which is introduced into the ooplasm upon sperm–egg fusion and concentrated to the pronucleus after inducing egg activation.