State Key Laboratory of Marine Environmental Science, College of Ocean & Earth Sciences, Xiamen University, Xiamen, China.
State-Province Joint Engineering Laboratory of Marine Bioproducts and Technology, College of Ocean & Earth Sciences, Xiamen University, Xiamen, China.
Front Immunol. 2022 Jul 8;13:946227. doi: 10.3389/fimmu.2022.946227. eCollection 2022.
Crustins are the most abundant class of antimicrobial peptides in crustaceans and are essential for protecting animals from infection. Among them, type II crustins usually exhibit potent antimicrobial activity. Interestingly, in this study, a newly identified type II crustin gene homolog (named SpCrus8) from mud crab , the recombinant proteins of which (rSpCrus8 and rTrx-SpCrus8) showed no obvious antibacterial effects, but could significantly reduce the bacterial load in crab hemolymph and improve the survival rate of crabs infected with . The immune-related function of SpCrus8 and the underlying mechanism deserve further investigation. It was found that the SpCrus8 gene was widely distributed in various tissues of adult crabs. In the hepatopancreas of crabs infected with or , transcripts of the SpCrus8 gene were remarkably induced, indicating that the SpCrus8 gene was involved in the immune response to bacterial infection . In addition, rSpCrus8 and rTrx-SpCrus8 had strong binding activity not only to microbial surface components (lipopolysaccharide, lipoteichoic acid, peptidoglycan, and glucan), but also to the tested bacteria (, and ). Notably, rSpCrus8 and rTrx-SpCrus8 could significantly promote hemocyte phagocytosis. After rSpCrus8 and rTrx-SpCrus8 treatment, a large number of fluorescent microspheres were observed to aggregate into clusters and be phagocytosed by multiple hemocytes, while hemocytes in the control group phagocytosed only individual microspheres, indicating that SpCrus8 played an important role in opsonization. When the SpCrus8 gene was knocked down, the expression levels of the key phagocytosis-related genes SpRab5 and SpRab7 were significantly downregulated, as well as the IMD signaling pathway genes SpIKKβ and SpRelish, and another crustin gene SpCrus5. Correspondingly, all the SpIKKβ, SpRelish and SpCrus5 genes were significantly upregulated after rSpCrus8 treatment, suggesting that SpCrus8 might be involved in the immunomodulation of . Taken together, this study revealed the immune-related functions of the SpCrus8 gene in opsonization and regulation, which will help us further understand the role of the crustin gene family in the immune system of mud crabs and provide new insights into the function of type II crutins.
甲壳质是甲壳类动物中最丰富的一类抗菌肽，对于保护动物免受感染至关重要。其中，II 型甲壳质通常具有很强的抗菌活性。有趣的是，在这项研究中，从泥蟹中鉴定出一种新的 II 型甲壳质基因同源物（命名为 SpCrus8），其重组蛋白（rSpCrus8 和 rTrx-SpCrus8）没有明显的抗菌作用，但可以显著降低蟹血淋巴中的细菌负荷并提高感染 的螃蟹的存活率。SpCrus8 的免疫相关功能及其潜在机制值得进一步研究。研究发现，SpCrus8 基因在成年蟹的各种组织中广泛分布。在感染 或 的蟹的肝胰腺中，SpCrus8 基因的转录物显著诱导，表明 SpCrus8 基因参与了对细菌感染的免疫反应。此外，rSpCrus8 和 rTrx-SpCrus8 不仅与微生物表面成分（脂多糖、脂磷壁酸、肽聚糖和葡聚糖）而且与测试细菌（、 和 ）具有很强的结合活性。值得注意的是，rSpCrus8 和 rTrx-SpCrus8 可以显著促进血细胞吞噬作用。在用 rSpCrus8 和 rTrx-SpCrus8 处理后，观察到大量荧光微球聚集成簇并被多个血细胞吞噬，而对照组中的血细胞仅吞噬单个微球，表明 SpCrus8 在调理作用中发挥重要作用。当 SpCrus8 基因被敲低时，关键吞噬相关基因 SpRab5 和 SpRab7 的表达水平以及 IMD 信号通路基因 SpIKKβ 和 SpRelish 和另一个甲壳质基因 SpCrus5 的表达水平显著下调。相应地，在用 rSpCrus8 处理后，所有 SpIKKβ、SpRelish 和 SpCrus5 基因均显著上调，表明 SpCrus8 可能参与了 的免疫调节。总之，本研究揭示了 SpCrus8 基因在调理和调节中的免疫相关功能，这将有助于我们进一步了解甲壳质基因家族在泥蟹免疫系统中的作用，并为 II 型甲壳质的功能提供新的见解。
