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. 2021 Apr 9;12:661823. doi: 10.3389/fimmu.2021.661823. eCollection 2021.
C-type lectin (CTL), a well-known immune-related molecule, has received more and more attention due to its diverse functions, especially its important role in development and host defense of vertebrate and invertebrate. Since the research on crab CTLs is still lack, we screened a new CTL homolog, named SpCTL6 from mud crab . The full-length cDNA sequence of SpCTL6 was 738 bp with a 486 bp of ORF, and the deduced amino acids were 161 aa. SpCTL6 was predicted to have a 17 aa signal peptide and its mature peptide was 144 aa (MW 16.7 kDa) with pI value of 5.22. It had typical CTL structural characteristics, such as a single C-type lectin-like domain, 4 conserved cysteines, similar tertiary structure to that of vertebrate CTLs and a mutated Ca binding motif Gln-Pro-Thr (QPT), clustering into the same branch as the crustacean CTLs. SpCTL6 was highly expressed in the entire zoeal larval stages and widely distributed in adult crab tissues with the highest transcription level in testis. During the molting process of juvenile crabs, the expression level of SpCTL6 was remarkably increased after molting. SpCTL6 could be significantly upregulated in two larval stages (Z1 and megalopa) and adult crab testis under immune challenges. Recombinant SpCTL6 (rSpCTL6) was successfully obtained from eukaryotic expression system. rSpCTL6 exhibited binding activity with PAMPs (LPS, lipoteichoic acid, peptidoglycan, and glucan) and had a broad spectrum bacterial agglutination activity in a Ca-dependent manner. In addition, rSpCTL6 could enhance the encapsulation activity of hemocytes and has no cytotoxic effect on hemocytes. Although rSpCTL6 had no bactericidal activity on , rSpCTL6 treatment could significantly reduce the bacterial endotoxin level and greatly improved the survival of under infection . The immunoprotective effect of rSpCTL6 might be due to the regulatory role of rSpCTL6 in immune-related genes and immunological parameters. Our study provides new information for understanding the immune defense of mud crabs and would facilitate the development of effective strategies for mud crab aquaculture disease control.
C 型凝集素（CTL）是一种功能多样的免疫相关分子，因其在脊椎动物和无脊椎动物的发育和宿主防御中的重要作用而受到越来越多的关注。由于对蟹类 CTLs 的研究仍然缺乏，我们从泥蟹中筛选到了一种新的 CTL 同源物，命名为 SpCTL6。SpCTL6 的全长 cDNA 序列为 738 bp，ORF 为 486 bp，推导的氨基酸为 161 aa。SpCTL6 预测有 17 aa 的信号肽，其成熟肽为 144 aa（MW 16.7 kDa），pI 值为 5.22。它具有典型的 CTL 结构特征，如单个 C 型凝集素样结构域、4 个保守半胱氨酸、与脊椎动物 CTLs 相似的三级结构和一个突变的 Ca 结合基序 Gln-Pro-Thr（QPT），聚类到与甲壳类 CTLs 相同的分支中。SpCTL6 在整个幼体阶段高表达，广泛分布于成年蟹组织中，在睾丸中的转录水平最高。在幼蟹蜕皮过程中，蜕皮后 SpCTL6 的表达水平显著增加。SpCTL6 可在两种幼虫阶段（Z1 和大眼幼体）和成年蟹睾丸中被免疫挑战显著上调。从真核表达系统中成功获得了重组 SpCTL6（rSpCTL6）。rSpCTL6 与 PAMPs（LPS、脂磷壁酸、肽聚糖和葡聚糖）具有结合活性，并以 Ca 依赖性方式具有广谱的细菌凝集活性。此外，rSpCTL6 可增强血细胞的包被活性，对血细胞无细胞毒性。尽管 rSpCTL6 对 没有杀菌活性，但 rSpCTL6 处理可显著降低 的内毒素水平，并显著提高 的存活率 感染 。rSpCTL6 的免疫保护作用可能归因于 rSpCTL6 对免疫相关基因和免疫参数的调节作用。我们的研究为了解泥蟹的免疫防御提供了新的信息，并将有助于开发有效的泥蟹水产养殖疾病控制策略。
