Genome-Wide Identification of the Collectin Family in Silver Pomfret (Pampus argenteus) and Innate Immune Functions of a Novel Truncated Member PaCL-11b against Vibrio parahaemolyticus.
Jul 2026· Fish and Shellfish Immunology· Vol 177, pp.
111601
· 0 citations· 66 references
Medicine
TL;DR
The systematic identification of the Collectin family in silver pomfret is presented and the multifunctional immune roles of PaCL-11b in defense against V. parahaemolyticus are revealed, providing a theoretical basis for the prevention and control of bacterial diseases in silver pomfret aquaculture.
Abstract
The Collectin family, an important member of the C-type lectin superfamily, participates in innate immune defense through Ca2+-dependent recognition of glycan ligands on pathogen surfaces. In this study, five Collectin family members were identified in the silver pomfret (Pampus argenteus) genome, and their gene structures, conserved motifs, and chromosomal localizations were systematically characterized. Notably, a novel member, collectin-11b (PaCL-11b) was found to encode a truncated isoform containing only the neck region and the carbohydrate recognition domain (CRD). It exhibited the highest expression level in the liver of healthy silver pomfret and was significantly downregulated in the liver, kidney, and gills following Vibrio parahaemolyticus challenge. Functional characterization of the recombinant protein rPaCL-11b demonstrated Ca2+-dependent binding, agglutination, direct antimicrobial, and antibiofilm activities against both Gram-negative (V. parahaemolyticus and Escherichia coli) and Gram-positive (Staphylococcus aureus) bacteria, with the most potent inhibitory effects observed against V. parahaemolyticus. This study presents a systematic identification of the Collectin family in silver pomfret and reveals the multifunctional immune roles of PaCL-11b in defense against V. parahaemolyticus infection, providing a theoretical basis for the prevention and control of bacterial diseases in silver pomfret aquaculture.
C-type lectins (CTLs) are important pattern recognition receptors (PRRs) involved in innate immune defense in crustaceans. In this study, a novel C-type lectin, designated LvCTL, was identified and functionally characterized from Litopenaeus vannamei. Sequence analysis revealed that the open reading frame (ORF) of LvCTL is 495 bp in length and encodes a 164-amino acid polypeptide containing a typical C-type lectin-like domain but lacking the classical carbohydrate-binding motifs. Notably, two novel EPF motifs were identified in the CTLD region, suggesting a potential non-canonical ligand recognition pattern. Phylogenetic analysis revealed that LvCTL clustered with other crustacean CTLs. Tissue expression analysis revealed that LvCTL exhibited the highest transcript level in the hepatopancreas. After immune stimulation, LvCTL expression in the hepatopancreas was significantly regulated by LPS, Vibrio parahaemolyticus, Staphylococcus aureus, WSSV and Poly(I: C), indicating its involvement in antimicrobial immune responses. RNA interference assays showed that knockdown of LvCTL significantly increased the bacterial burden and reduced the survival rate of shrimp after V. parahaemolyticus infection. Furthermore, recombinant LvCTL displayed broad bacterial binding activity to Gram-negative and Gram-positive bacteria and induced calcium-dependent agglutination of V. parahaemolyticus and S. aureus. Taken together, these results demonstrate that LvCTL functions as an important immune recognition molecule and contributes to antibacterial defense against V. parahaemolyticus in L. vannamei. These findings broaden our understanding of the functional diversity of shrimp CTLs and their contributions to crustacean innate immunity.
Ning Fu, Mengxin Xing, Yuyu Wang et al.· Fishes· 0 citations
Ladderlectin, a member of the C-type lectin (CTL) family, has historically been identified only in fish and shrimp, where it is primarily involved in innate immune defense. In teleosts, ladderlectin plays a crucial dual role in both innate immunity against pathogens and reproductive regulation. However, the molecular characterization and immunomodulatory functions of ladderlectin homolog (ladderlectin-like) in Hexagrammos otakii remain poorly understood. In this study, a novel ladderlectin-like gene (HoLL-like) was identified and cloned from Hexagrammos otakii based on genomic data. Sequence analysis and domain prediction revealed that HoLL-like contains a 534-bp open reading frame encoding a 177-aa polypeptide, with an N-terminal signal peptide and a single C-type lectin-like domain (CTLD). The CTLD retains the WSD, WND, and WIGG motifs and is rich in cysteine residues. Phylogenetic analysis showed that HoLL-like forms a distinct, relatively independent clade within the teleost ladderlectin-like family. Quantitative RT-PCR analysis demonstrated that Holl-like is expressed in multiple tissues, with the highest transcript levels in the liver. Besides, it exhibited a time-dependent transcriptional response in the liver, spleen, skin, intestine, and kidney following Vibrio harveyi infection. After heterologous expression, purification, and refolding, the recombinant protein rHoLL-like displayed characteristic ladder-like oligomeric bands in immunoblotting. Functional assays revealed that rHoLL-like binds and agglutinates multiple Gram-positive and Gram-negative bacteria in a Ca2+-independent manner and binds pathogen-associated molecular patterns (PAMPs), including lipopolysaccharide (LPS), peptidoglycan (PGN), and poly(I:C) in a dose-dependent manner, showing higher affinity for LPS. Further analyses indicated that rHoLL-like did not directly agglutinate or lyse erythrocytes, but enhanced serum hemolytic and bactericidal activities under Ca2+-containing conditions. These effects were sensitive to EDTA and serum heat inactivation, suggesting that rHoLL-like may modulate heat-labile, Ca2+-dependent serum effector mechanisms, potentially associated with complement activity. Collectively, these findings suggest that HoLL-like plays a crucial role in the innate immune response of H. otakii against bacterial and other pathogen infections. This study expands our understanding of the functional repertoire of teleost ladderlectin-like proteins and provides new insights into the molecular strategies to prevent and control bacterial diseases in marine aquaculture, as well as for the development of immunoactive agents.
Yuyoumin Wen, Xu Wei, Tingting He et al.· Fish and Shellfish Immunolog...· 0 citations
Golden pompano (Trachinotus ovatus) is an economically important marine fish in southern China, but intensive aquaculture has increased the risk of bacterial diseases, especially streptococcosis caused by Streptococcus agalactiae. As a key component of innate immunity, the NLRP3 inflammasome (NOD-like receptor family, pyrin domain containing 3 inflammasome) regulates inflammatory responses through caspase-1 (Cysteine-aspartic acid protease 1) activation and the maturation of IL-1β (Interleukin-1β) and IL-18 (Interleukin-18). In this study, five core genes associated with the NLRP3 inflammasome pathway in T. ovatus including ToNLRP3, ToASC, ToCaspase-1, ToIL-1β, and ToIL-18 were cloned and characterized. Sequence analysis showed that these genes contain conserved functional domains and share evolutionary conservation with teleost homologs. Tissue distribution analysis revealed broad expression, with relatively high levels in immune-related and mucosal tissues, including the spleen, head kidney, gill, intestine, and skin. After S. agalactiae challenge, all five genes showed inducible and tissue-dependent expression patterns, supporting their involvement in the antibacterial immune response. Fluorescence-based assays showed that ToNLRP3 was mainly localized in the cytoplasm and that ToASC interacted with ToCaspase-1, indicating the potential for inflammasome-like complex assembly. These findings provide a basis for understanding inflammasome-related immunity in golden pompano.
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C-type lectins (CTLs) are vital pattern-recognition receptors (PRRs) that mediate innate immune responses in mollusks, yet their characterization in Caenogastropoda, the largest gastropod group, remains limited. This study characterizes LitCTL1, a novel secreted single-domain C-type lectin from the common periwinkle, Littorina littorea. The 199-amino acid polypeptide contains a conserved carbohydrate recognition domain with canonical QPD and WND motifs and is predicted to form a homodimer. Uniquely, LitCTL1 was localized in both circulating hemocytes and mucus-secreting epithelial cells of the foot, mantle, and hypobranchial gland - the first report of such dual localization for a molluscan lectin, linking systemic and mucosal defense. Expression analysis revealed that LitCTL1 is constitutively expressed in hemocytes. Functional assays with recombinant LitCTL1 demonstrated its role as a potent opsonin with hemagglutinating activity, significantly enhancing hemocyte spreading and the phagocytosis of zymosan. Genomic analysis reveals that LitCTL1 belongs to a rapidly diversifying, genus-specific expansion distinct from conserved perlucin-like lineages. These results identify LitCTL1 as a key effector molecule in both systemic and mucosal innate immunity, likely reflecting an evolutionary adaptation to the microbial challenges of the intertidal environment.
Alexander Gorbushin, E. Borisova· Fish and Shellfish Immunolog...· 0 citations
Toll-like receptor (TLR) family proteins act as pattern recognition receptors with pivotal roles in immune defense; however, the function of TLRs in marine mollusks requires further investigation. In this study, we identified a novel TLR gene (CfTLR-Like-1 [CfTLR-L1]) in Chlamys farreri and analyzed its immune functions. CfTLR-L1 has a coding sequence of 1,785 bp and encodes 594 amino acids. It includes typical protein domains; the amino acid sequences of the protein domains are conserved. Phylogenetic analyses demonstrated a close evolutionary relationship among proteins in the mollusk TLR subfamily. CfTLR-L1 is predominantly located on the cell membrane. CfTLR-L1 mRNA is expressed in all tissues, particularly in the hepatopancreas and mantle. Pathogenic stressors, including lipopolysaccharide, peptidoglycan, polyinosinic-polycytidylic acid, Vibrio anguillarum, Vibrio alginolyticus, Vibrio splendidus, and acute viral necrosis virus, significantly increase the levels of CfTLR-L1 mRNA transcripts. The Toll/interleukin 1 receptor (TIR) domain of CfTLR-L1 interacts with the MyD88 family proteins of scallops (including CfMyD88-1, CfMyD88-3, and CfMyD88-4), implying that multiple unique TLR signal transduction mechanisms exist in marine mollusks. CfTLR-L1-TIR interacts with CfMyD88-1-P3, which contains only the TIR structural domain. Overexpression of CfTLR-L1-TIR in HEK293T cells activated reporter genes, including IFNβ, IFNγ, and STAT3. Similarly, overexpression of CfTLR-L1 significantly affected mitogen-activated protein kinase phosphorylation, specifically of p38 and c-Jun N-terminal kinase. Our findings expand the knowledge of the role of TLR in mollusk immunity and may provide evidence for selecting disease-resistant scallops for cultivation.
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SiTRAF3 is identified as a pivotal scaffold protein in the echinoid innate immune system, as it significantly drove the promoter activities of nuclear factor kappa-B, activator protein-1, signal transducer and activator of transcription 3, along with multiple inflammatory cytokines and interferon-related elements.
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