A new crustin homologue (SpCrus6) involved in the antimicrobial and antiviral innate immunity in mud crab, Scylla paramamosain
A new crustin homologue (SpCrus6) involved in the antimicrobial and antiviral innate immunity in mud crab, Scylla paramamosain
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一种新的壳蛋白同源物 (SpCrus6) 参与青蟹的抗菌和抗病毒先天免疫
DOI：
10.1016/j.fsi.2018.10.072
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发表时间：
2019-01-01
影响因子：
4.7
通讯作者：
Li, Xin-cang
中科院分区：
文献类型：
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作者：
Du, Zhi-qiang;Wang, Yue;Li, Xin-cang
在甲壳动物的天然免疫系统中，甲壳蛋白在抵御细菌方面发挥着重要作用。在本研究中，我们在青蟹中发现了一个crustin基因，命名为SpCrus6。SpCrus6的ORF在N端有一个信号肽序列(SPS)，在C端有一个WAP结构域。在SpCrus6的SPS和WAP结构域之间有5个Pro残基、5个甘氨酸残基和4个半胱氨酸残基。这些特征表明SpCrus6是crustin家族的新成员。细菌或病毒攻击后，SpCrus6基因转录明显上调。这些变化表明SpCrus6参与了副青蟹的抗微生物和抗病毒反应。重组SpCrus6(RSpCrus6)对革兰氏阳性菌(巨型芽孢杆菌、金黄色葡萄球菌和枯草芽孢杆菌)有很强的抑制作用。但对4种革兰氏阴性菌(副溶血性弧菌、溶藻弧菌、哈维氏弧菌和大肠杆菌)和2种真菌(巴氏毕赤酵母和白色念珠菌)的抑制作用不强。此外，rSpCrus6能与两种革兰氏阳性菌(枯草杆菌和巨大芽孢杆菌)和三种革兰氏阴性菌(溶藻弧菌、副溶血弧菌和哈维氏弧菌)强结合。对金黄色葡萄球菌和两种真菌(巴氏葡萄球菌和白色念珠菌)的结合水平较弱。多糖结合实验结果表明，rSpCrus6对脂多糖、LTA、PGN和β-葡聚糖具有较好的结合活性。通过凝集试验，我们发现rSpCrus6对3种革兰氏阳性菌(金黄色葡萄球菌、枯草杆菌和巨大芽孢杆菌)有很好的凝集作用。对革兰氏阴性杆菌和真菌无凝集活性。在抗病毒功能方面，rSpCrus6能与对虾白斑综合征病毒(WSSV)的重组囊膜蛋白26(RVP26)特异结合，但不能与重组囊膜蛋白28(RVP28)结合，而GST蛋白不能与rVP26或rVP28结合。此外，rSpCrus6还能在一定程度上抑制WSSV的复制。综上所述，SpCrus6是副伤寒沙门氏菌天然免疫防御反应中的多功能免疫效应因子。
Crustins play important roles in defending against bacteria in the innate immunity system of crustaceans. In present study, we identified a crustin gene in Scylla paramamosain, which was named as SpCrus6. The ORF of SpCrus6 possessed a signal peptide sequence (SPS) at the N-terminus and a WAP domain at the C-terminus. And there were 5 Proline residues, 5 Glycine and 4 Cysteine residues between SPS and WAP domain in SpCrus6. These features indicated that SpCrus6 was a new member of crustin family. The SpCrus6 mRNA transcripts were up-regulated obviously after bacteria or virus challenge. These changes showed that SpCrus6 was involved in the antimicrobial and antiviral responses of Scylla paramamosain. Recombinant SpCrus6 (rSpCrus6) showed strong inhibitory abilities against Gram-positive bacteria (Bacillus megaterium, Staphylococcus aureus, and Bacillus subtilis). But the inhibitory abilities against four Gram-negative bacteria (Vibrio parahemolyticus, Vibrio alginolyticus, Vibrio harveyi and Escherichia colt) and two fungi (Pichia pastor-is and Candida albicans) were not strong enough. Besides, rSpCrus6 could strongly bind to two Gram-positive bacteria (B. subtilis and B. megaterium) and three Gram-negative bacteria (V. alginolyticus, V. parahemolyticus, and V. harveyi). And the binding levels to S. aureus and two fungi (P. pastoris and C. albicans) were weak. The polysaccharides binding assays' results showed rSpCrus6 had superior binding activities to LPS, LTA, PGN and beta-glucan. Through agglutinating assays, we found rSpCrus6 could agglutinate well three Gram-positive bacteria (S. aureus, B. subtilis and B. megaterium). And the agglutinating activities to Gram-negative bacteria and fungi were not found. In the aspect of antiviral functions, rSpCrus6 could bind specifically to the recombinant envelop protein 26 (rVP26) of white spot syndrome virus (WSSV) but not to recombinant envelop protein 28 (rVP28), whereas GST protein could not bind to rVP26 or rVP28. Besides, rSpCrus6 could suppress WSSV reproduction to some extent. Taken together, SpCrus6 was a multifunctional immunity effector in the innate immunity defending response of S. paramamosain.