Zona pellucida (ZP) modules mediate extracellular protein–protein interactions and contribute to important biological processes including syngamy and cellular morphogenesis. Although some biomedically relevant ZP modules are well studied, little is known about the protein family’s broad-scale diversity and evolution. The increasing availability of sequenced genomes from “nonmodel” systems provides a valuable opportunity to address this issue and to use comparative approaches to gain new insights into ZP module biology. Here, through phylogenetic and structural exploration of ZP module diversity across the nematode phylum, I report evidence that speaks to two important aspects of ZP module biology. First, I show that ZP-C domains—which in some modules act as regulators of ZP-N domain-mediated polymerization activity, and which have never before been found in isolation—can indeed be found as standalone domains. These standalone ZP-C domain proteins originated in independent (paralogous) lineages prior to the diversification of extant nematodes, after which they evolved under strong stabilizing selection, suggesting the presence of ZP-N domain-independent functionality. Second, I provide a much-needed phylogenetic perspective on disulfide bond variability, uncovering evidence for both convergent evolution and disulfide-bond reshuffling. This result has implications for our evolutionary understanding and classification of ZP module structural diversity and highlights the usefulness of phylogenetics and diverse sampling for protein structural biology. All told, these findings set the stage for broad-scale (cross-phyla) evolutionary analysis of ZP modules and position Caenorhabditis elegans and other nematodes as important experimental systems for exploring the evolution of ZP modules and their constituent domains.

中文翻译：

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线虫Zona Pellucida（ZP）模块的分子进化分析揭示了二硫键的改组和独立的ZP-C域。

透明带（ZP）模块介导细胞外蛋白质之间的相互作用，并促进重要的生物学过程，包括合酶和细胞形态发生。尽管已经对一些生物医学相关的ZP模块进行了深入研究，但对蛋白质家族的广泛多样性和进化知之甚少。来自“非模型”系统的测序基因组的可用性不断提高，为解决此问题并使用比较方法获得对ZP模块生物学的新见解提供了宝贵的机会。在这里，通过对整个线虫门的ZP模块多样性进行系统发育和结构探索，我报告了与ZP模块生物学的两个重要方面相关的证据。首先，我展示了ZP-C域-在某些模块中充当ZP-N域介导的聚合活性的调节剂，并且以前从未被孤立地发现过-确实可以作为独立域找到。这些独立的ZP-C结构域蛋白在现存线虫多样化之前起源于独立的（旁系）谱系，之后它们在强烈的稳定选择下进化，表明存在ZP-N域独立的功能。其次，我提供了关于二硫键变异性的急需的系统发育观点，为融合进化和二硫键改组提供了证据。该结果对我们对ZP模块结构多样性的进化理解和分类具有重要意义，并突出了系统发育学和多样性采样对蛋白质结构生物学的有用性。总而言之，秀丽隐杆线虫和其他线虫是探索ZP模块及其组成域进化的重要实验系统。