Molecular mechanism of cytokinesis

1)认为某些信号从有丝分裂器传递到细胞皮质，以便在适当的位置形成收缩环。然而，其作用机制尚不清楚。我使用荧光素标记的小麦胚芽凝集素(WGA)受体研究了WGA受体在这一过程中的作用。中期，WGA受体分布于卵子表面。它们在后期聚集成簇，后期变成纤维状。在末期，它们聚集在预期的沟槽区域形成环状。上面观察到的肌动蛋白细丝的分布与荧光鬼笔花素染色显示的肌动蛋白细丝非常相似。已有研究表明，WGA受体与肌动蛋白在质膜上的微丝锚定部位有密切关系。有一种可能性被否认，即受体首先聚集在预期的沟槽区域，然后诱导肌动蛋白细丝组装。2)当未受精的海胆卵与海绵毒素Calyculin A孵育时，可诱导卵裂。这些卵子既没有形成有丝分裂器官，也没有形成二倍体。肌动蛋白、肌球蛋白和WGA受体聚集在沟沟区。这一过程被蛋白激酶抑制剂抑制。提示蛋白质磷酸化参与了收缩环的形成。此外，染色体被浓缩，组蛋白1激酶活性以非细胞周期蛋白B依赖的方式增加。3)去年，我报道了使用含有洗涤剂的培养液分离出Clypester Japan卵的卵裂沟。我用奇异的Scaphechinus卵延长了隔离时间。我还用改良的培养液从青海花蝇和大眼球蛾的卵中分离出了沟沟。我们在这些鸡蛋的二维凝胶中发现了几种常见的蛋白质，现在正在确定部分肽序列。

1)It has been considered that some signal is transfered from the mitotic apparatus to the cell cortex in order to form the contractile ring at a proper position. However, its mechanism has not been clarified yet. I investigated role of wheat germ agglutinin (WGA) receptors in this process, using fluorescein-labeled WGA. WGA receptors distributed on the surface of the egg during metaphase. They were aggregated into clusters during anaphase, which became fibrous in late anaphase. At telophase, they accumulated in the anticipated furrow region forming a ring. The distribution observed above was quite similar to that of actin filaments visualized by fluorescent phalloidin staining. It has been shown that the WGA receptors have intimate relationship to actin filament-anchoring sites on the plasma membrane. A possibility was denied that the receptors accumulate in the anticipated furrow region at first and then induce actin filament assembly. 2) When the unfertilized sea urchin eggs were incubated with calyculin A, a sponge toxin which inhibits activities of protein phopsphatases, cleavage was induced. Neither mitotic apparatus nor aster was formed in these eggs. Actin, myosin, and WGA receptors were accumulated in the furrow region. This process was inhibited by protein kinase inhibitors. It is suggested that protein phosphorylation is involved in the contractile ring formation. In addition, chromosomes were condensed and histone 1 kinase activity was increased in a cyclin Bindependent manner. 3) Last year, I reported isolation of cleavage furrows from Clypeaster japonicus eggs using a detergent-containing medium. I extended the isolation with Scaphechinus mirabilis eggs. I also isolated clevage furrows from eggs of Anthocidaris crassispina and Hemicentrotus pulcherrimus using modified medium. We found several proteins in 2-D gels common to these eggs and now determining partial peptide sequences.