Molecular Basis of Sperm-Egg Binding in Frogs

尽管在各种生物系统中详细描述受体-配体的相互作用已经取得了很大的进展，但关于介导细胞-细胞黏附的这种相互作用的知识还很少，即“受体”在一个细胞上，而“配体”在另一个细胞上。受精是一种特殊的细胞-细胞黏附，它有一个明确而深刻的终点，即形成受精卵。在这方面得到广泛研究的两个系统是海胆和老鼠。在这些系统中需要解决的一个中心问题是，在细胞外被层相互作用后，精子如何进入下一阶段，与卵子质膜相互作用并融合。值得注意的是，尽管青蛙被广泛而有效地用于研究与早期发育相关的基本现象，但在受精过程中配子相互作用的机制方面却相对较少受到关注。青蛙系统可能比正在研究的任何其他生物系统都要好，它有可能有助于阐明精子和卵子之间的阶段性相互作用，最终导致两个配子的融合。这是因为1)精子与蛙细胞外被膜中一种被称为卵黄膜(VE)的糖蛋白的最初相互作用已经被表征，2)蛙的VE与质膜之间的关系已经被他人从超微结构上进行了很好的研究。在这项研究中，我们将进一步研究VE中发现的一种参与精子结合的糖蛋白(gp69/)的多肽链和寡糖链结构。将研究这种糖蛋白的每个结构元素在精子结合中的作用以及这一过程是如何发育调节的。已有初步证据表明，质膜上存在一种与抗VE糖蛋白抗体gp69/发生交叉反应的大分子糖蛋白。将对VIE结合到质膜结合的转变以及后者的分子基础进行研究。为此，将研究去除部分VE以暴露质膜“窗口”的卵子，以及完全剥离VE的卵子的精子结合情况。最后，将研究精子和卵子的质膜融合，并检查交叉反应的卵子质膜蛋白在这一事件中的可能作用。预计这些研究将为精子和卵子之间的多步骤相互作用带来新的见解，最终导致受精。

Although great progress has been made in describing in detail receptor-ligand interactions in a wide variety of biological systems, knowledge about such interactions that mediate cell-cell adhesion, where the "receptor" is present on one cell and the "ligand" is on another, is much less developed. Fertilization is a specialized case of cell-cell adhesion that has a well-defined and profound endpoint, formation of a zygote. Two systems that have been studied extensively in this respect are the sea urchin and the mouse. A central issue to be resolved in these systems is how, following interaction at the extracellular coat, the sperm moves to the next stage, interacts and fuses with the egg plasma membrane. Remarkably, the frog, although used very extensively and productively to study basic phenomena related to early development, has received relatively little attention with respect to the mechanism of gamete interaction during fertilization. The frog system, perhaps better than any of the other biological systems under study, has the potential to lead to elucidation of the staged interactions of the sperm and the egg that culminate in fusion of the two gametes. This is the case because 1 ) the initial interaction of the sperm with a glycoprotein in the frog extracellular coat known as the vitelline envelope (VE) has been characterized and 2) the relationship between the VE and the plasma membrane in the frog has been well studied ultrastructurally by others. In this proposal a glycoprotein (gp69/64) found in the VE and shown to be involved in sperm binding will be further studied with respect to the structure of its polypeptide chain and its oligosaccharide chains. The role of each structural element of this glycoprotein in sperm binding and how this process is developmentally regulated will be studied. Preliminary evidence has been obtained for the presence of a high molecular weight glycoprotein in the plasma membrane that cross reacts with antibody to the VE glycoprotein , gp69/64. A study of the transition from VIE binding to plasma membrane binding and the molecular basis of the latter event will be under taken. To do this, eggs from which a portion of the VE has been removed to expose a "window" of the plasma membrane, as well as eggs completely denuded of their VE, will be studied for sperm binding. Finally, fusion of the plasma membrane of the sperm and egg will be investigated and the possible role of the cross-reactive egg plasma membrane protein in this event will be examined. It is anticipated that these studies will bring new insight into the multistep interactions between sperm and egg that culminate in fertilization.