Bioscience and Biotechnology Center, Nagoya University, Chikusa, Nagoya 464-8601, Japan.
Graduate School of Bioagricultural Sciences, Nagoya University, Chikusa, Nagoya 464-8601, Japan.
Biomolecules. 2023 Jan 10;13(1):146. doi: 10.3390/biom13010146.
Alveolin is a cortical alveolus proteinase that is secreted in the perivitelline space (PVS) at fertilization to act on the chorion. Purified alveolin is known to induce chorion hardening in vitro by processing zona pellucida B (ZPB), a major chorion component. However, in vivo function of alveolin remains unclear; thus, in this study, the effects of efficiency () at the organism level were investigated using the medaka, . The fertilized eggs were mechanically fragile; however, they developed normally and left offspring as long as they were carefully handled before hatching. A mechanical press test showed that the fertilized eggs were six times more fragile than the wild-type eggs. They were 35% larger owing to the enlarged PVS, 34% thinner, and permeable to even 10 kDa FITC-dextran. These results are consistent with the transmission electron microscopy observation that the periphery of the inner layers was highly porous in the chorion. In chorion hardening, the alveolin-mediated processing of ZPB and the transglutaminase (TGase)-mediated crosslinking of chorion components are the key steps. This study was the first to show that alveolin also processed TGase concomitantly with ZPB, which greatly facilitated the crosslinking. Thus, alveolin was concluded to be the primary trigger for chorion hardening in vivo. Furthermore, fertilization in a balanced salt solution could partially improve the impaired chorion hardening of the eggs fertilized in water, probably through an alveolin-independent mechanism.
肺泡蛋白水解酶是一种皮质肺泡蛋白水解酶，在受精时分泌到卵黄周隙（PVS）中，作用于卵膜。已知纯化的肺泡蛋白水解酶能够通过处理主要卵膜成分 ZPB 来诱导卵膜变硬。然而，肺泡蛋白水解酶的体内功能尚不清楚；因此，在这项研究中，使用 来研究其在生物体水平上的效应。受精的 卵机械上很脆弱；然而，只要在孵化前小心处理，它们就能正常发育并留下后代。机械压力测试表明， 受精的卵比野生型卵脆弱六倍。由于 PVS 增大，它们的体积增大了 35%，厚度变薄了 34%，甚至 10 kDa 的 FITC-葡聚糖也能渗透。这些结果与透射电子显微镜观察结果一致，即 卵膜内层的周边高度多孔。在卵膜变硬过程中，肺泡蛋白水解酶介导的 ZPB 处理和转谷氨酰胺酶（TGase）介导的卵膜成分交联是关键步骤。这项研究首次表明，肺泡蛋白水解酶还同时处理 TGase 和 ZPB，这极大地促进了交联。因此，肺泡蛋白水解酶被认为是体内卵膜变硬的主要触发因素。此外，在平衡盐溶液中受精可以部分改善在水中受精的 卵受损的卵膜变硬，可能是通过一种与肺泡蛋白水解酶无关的机制。
