Chen Xiaofeng, Shen Xiaoya, Liu Shijun, Li Wenjuan, Wang He, Li Jiale, Bai Zhiyi
Key Laboratory of Freshwater Aquatic Genetic Resources, Ministry of Agriculture and Rural Affairs, Shanghai Ocean University, Shanghai, China.
Science and Technology Service Center, Shanghai MugaoBiotechnology co., Ltd., Shanghai, China.
Front Immunol. 2025 Jan 14;15:1530732. doi: 10.3389/fimmu.2024.1530732. eCollection 2024.
Shell and pearl formation in bivalves is a sophisticated biomineralization process that encompasses immunological and mineralization aspects, particularly during shell repair and the initial stages of pearl cultivation when a nucleus is inserted. Here, we describe a novel C-type lectin, HcLec1, isolated and characterized from the freshwater pearl mussel Lea.
Immune challenge, RNA interference (RNAi) experiments, ELISA, and antibacterial assays were employed to investigate the role of HcLec1 in innate immunity. We also established shell damage repair and pearl nucleus insertion models to examine the impact of on the biomineralization process in Lea. In vitro calcium carbonate crystallization assays were conducted to explore the direct role of in calcium carbonate crystal formation.
The gene sequence is a full-length cDNA of 1552 bp, encoding 240 amino acids. comprises an N-terminal signal peptide and a carbohydrate-recognition domain (CRD), with QPD (Gln-Pro-Asp) and MND (Met-Asn-Asp) motifs for polysaccharide binding. Tissue expression analysis showed that is predominantly expressed in the gill tissue of Lea under normal conditions, and its expression is significantly elevated in both gill and pearl sac tissues following nucleus insertion for pearl cultivation ( < 0.05). After immune stimulation with and lipopolysaccharides (LPS), expression levels significantly increased in both cases ( < 0.01), indicating a role in bivalve innate immunity. RNA interference (RNAi)-mediated knockdown of led to a significant decrease in the expression levels of immune-related genes (, , and ) and mineralization-related genes (, , , and ) ( < 0.05). In animal models for shell damage and nucleus insertion in pearl cultivation, showed a consistent expression pattern, with an initial significant decrease followed by a marked increase, peaking at day 14 ( < 0.05). This suggests a role for in pearl formation and shell repair. The recombinant HcLec1 protein demonstrated binding affinity to LPS and PGN, a robust ability to agglutinate , , , and , and significantly inhibited bacterial growth ( < 0.05). Moreover, rHcLec1 promoted calcite crystal formation in saturated calcium carbonate solutions and altered crystal morphology.
The gene plays a pivotal role in both innate immunity and biomineralization in the triangle sail mussel. This study enhances our understanding of the functional diversity of C-type lectins and provides a foundation for future studies on shell repair and pearl growth.
双壳贝类的贝壳和珍珠形成是一个复杂的生物矿化过程，涉及免疫和矿化方面，特别是在贝壳修复和珍珠养殖初期插入珠核时。在此，我们描述了一种从淡水珍珠蚌丽蚌中分离并鉴定的新型C型凝集素HcLec1。
采用免疫挑战、RNA干扰（RNAi）实验、酶联免疫吸附测定（ELISA）和抗菌测定来研究HcLec1在先天免疫中的作用。我们还建立了贝壳损伤修复和珍珠核插入模型，以研究其对丽蚌生物矿化过程的影响。进行了体外碳酸钙结晶测定，以探讨其在碳酸钙晶体形成中的直接作用。
HcLec1基因序列是一个1552 bp的全长cDNA，编码240个氨基酸。HcLec1包含一个N端信号肽和一个碳水化合物识别结构域（CRD），具有用于多糖结合的QPD（谷氨酰胺-脯氨酸-天冬氨酸）和MND（甲硫氨酸-天冬酰胺-天冬氨酸）基序。组织表达分析表明，在正常条件下，HcLec1主要在丽蚌的鳃组织中表达，在插入珠核进行珍珠养殖后，其在鳃和珍珠囊组织中的表达均显著升高（P<0.05）。在用嗜水气单胞菌和脂多糖（LPS）进行免疫刺激后，两种情况下HcLec1的表达水平均显著增加（P<0.01），表明其在双壳贝类先天免疫中发挥作用。RNA干扰（RNAi）介导的HcLec1敲低导致免疫相关基因（Tnf、Il-1β和Il-6）和矿化相关基因（Col1、Col3、Mmp9和Timp1）的表达水平显著降低（P<0.05）。在贝壳损伤和珍珠养殖核插入的动物模型中，HcLec1呈现出一致的表达模式，最初显著下降，随后显著增加，在第14天达到峰值（P<0.05）。这表明HcLec1在珍珠形成和贝壳修复中发挥作用。重组HcLec1蛋白表现出对LPS和肽聚糖的结合亲和力，对嗜水气单胞菌、大肠杆菌、金黄色葡萄球菌和藤黄微球菌具有强大的凝集能力，并显著抑制细菌生长（P<0.05）。此外，重组HcLec1促进了饱和碳酸钙溶液中方解石晶体的形成并改变了晶体形态。
HcLec1基因在三角帆蚌的先天免疫和生物矿化中都起着关键作用。本研究增进了我们对C型凝集素功能多样性的理解，并为未来贝壳修复和珍珠生长的研究奠定了基础。
