Department of Biological Sciences, Stanford University, Hopkins Marine Station, Pacific Grove, CA 93950.
Proc Natl Acad Sci U S A. 1985 Apr;82(8):2389-93. doi: 10.1073/pnas.82.8.2389.
We have followed the sequestration of Ca(2+) by intracellular compartments in sea urchin embryos through the first cell cycles. To gain biochemical access to these compartments, the embryos were permeabilized by brief exposure to an intense electric field. Sequestration was determined as the retention of tracer, (45)Ca, after filtration of aliquots on Millipore filters. The permeabilized cells sequester Ca(2+) at a constant rate for at least 20 min, with the following characteristics: (i) ATP is required. (ii) Sequestration occurs at Ca(2+) levels corresponding to those estimated in vivo. (iii) The Ca(2+) concentration dependence of sequestration and its insensitivity to mitochondrial poisons imply that the activity derives from a single, nonmitochondrial transport system. The Ca(2+)-sequestering activities of embryos that are permeabiized at successive stages of the first cell cycle (one-cell stage) progressively increase to 5 times the initial level. The rate of sequestration is maximal during telophase and, in some populations of zygotes, is nearly as great throughout prophase. Over the course of the second cell cycle (two-cell stage), the activity undergoes a 2-fold oscillation that bears the same temporal relationship to mitosis as the previous fluctuation.
我们通过海胆胚胎的第一个细胞周期，观察了细胞内区室对 Ca(2+)的螯合作用。为了获得这些区室的生化方法，胚胎通过短暂暴露在强电场中来实现通透性。通过将等分试样过滤在 Millipore 滤器上，来确定（45）Ca 的示踪剂的保留情况，以此来确定螯合作用。通透性细胞以恒定的速率螯合 Ca(2+)至少 20 分钟，具有以下特征：（i）需要 ATP。（ii）螯合作用发生在与体内估计值相对应的 Ca(2+)水平。（iii）Ca(2+)浓度依赖性螯合作用及其对线粒体毒物的不敏感性表明，该活性源自单个非线粒体转运系统。在第一个细胞周期（单细胞阶段）的连续阶段通透性的胚胎的 Ca(2+)-螯合活性逐渐增加到初始水平的 5 倍。在末期，螯合作用的速度达到最大值，并且在一些受精卵群体中，在整个前期几乎与有丝分裂一样大。在第二个细胞周期（双细胞阶段）期间，活性经历了 2 倍的波动，其与有丝分裂的时间关系与前一个波动相同。
