Division of Animal Sciences, University of Missouri-Columbia, Columbia, MO 65201, USA.
Dev Biol. 2012 Jul 15;367(2):154-62. doi: 10.1016/j.ydbio.2012.04.028. Epub 2012 May 4.
During fertilization in mammals, a series of oscillations in the oocyte's intracellular free Ca(2+) concentration is responsible for oocyte activation and stimulation of embryonic development. The oscillations are associated with influx of Ca(2+) across the plasma membrane that is probably triggered by the depletion of the intracellular stores, a mechanism known as store-operated Ca(2+) entry. Recently, STIM1 has been identified in oocytes as a key component of the machinery that generates the Ca(2+) influx after store depletion. In this study, the involvement of STIM1 in the sperm-induced Ca(2+) oscillations and its significance in supporting subsequent embryo development were investigated. Downregulation of STIM1 levels in pig oocytes by siRNA completely inhibited the repetitive Ca(2+) signal triggered by the fertilizing sperm. In addition, a significantly lower percentage of oocytes cleaved or formed blastocysts when STIM1 was downregulated prior to fertilization compared to the control groups. Restoring STIM1 levels after fertilization in such oocytes by means of mRNA injection could not rescue embryonic development that in most cases was arrested at the 2-cell stage. On the other hand, STIM1 overexpression prior to fertilization did not alter the pattern of sperm-induced Ca(2+) oscillations and development of these fertilized oocytes up to the blastocyst stage was also similar to that registered in the control group. Finally, downregulation of STIM1 had no effect on oocyte activation when activation was stimulated artificially by inducing a single large elevation in the oocyte's intracellular free Ca(2+) concentration. These findings suggest that STIM1 is essential for normal fertilization as it is involved in the maintenance of the long-lasting repetitive Ca(2+) signal.
在哺乳动物的受精过程中，卵母细胞内游离 Ca(2+) 浓度的一系列波动负责卵母细胞的激活和胚胎发育的刺激。这些波动与 Ca(2+) 通过质膜的内流有关，这种机制可能是由细胞内储存的耗竭触发的，这种机制被称为储存操纵的 Ca(2+) 内流。最近，STIM1 在卵母细胞中被鉴定为产生储存耗竭后 Ca(2+) 内流的机器的关键组成部分。在这项研究中，研究了 STIM1 在精子诱导的 Ca(2+) 波动中的作用及其在支持随后胚胎发育中的意义。通过 siRNA 下调猪卵母细胞中的 STIM1 水平完全抑制了由受精精子触发的重复 Ca(2+) 信号。此外，与对照组相比，在受精前下调 STIM1 水平的卵母细胞中，卵裂或形成囊胚的比例显著降低。通过 mRNA 注射在这种卵母细胞中恢复 STIM1 水平后，不能挽救胚胎发育，大多数情况下胚胎发育在 2 细胞阶段停滞。另一方面，在受精前过表达 STIM1 并没有改变精子诱导的 Ca(2+) 波动模式，并且这些受精卵母细胞的发育也与对照组中记录的模式相似。最后，当通过诱导卵母细胞内游离 Ca(2+) 浓度的单次大幅升高来人工刺激卵母细胞激活时，STIM1 的下调对卵母细胞的激活没有影响。这些发现表明，STIM1 对于正常受精是必不可少的，因为它参与维持持久的重复 Ca(2+) 信号。
