Aug 2026· Fish and Shellfish Immunology· Vol 178, pp.
111649
· 0 citations· 66 references
Medicine
TL;DR
Subcellular localization revealed that SsTFPI-1 predominantly localizes in the cytoplasm of EPC cells, and its C-terminus undergoes endogenous cleavage, which underscores the potential of TS20 as a promising antimicrobial agent.
Abstract
Tissue factor pathway inhibitor (TFPI) is a serine protease inhibitor existing in two isoforms, TFPI-1 and TFPI-2. Peptides derived from the TFPI C-terminus exhibit potent antimicrobial activity. In this study, a TFPI-1 homolog from Sebastes schlegelii (SsTFPI-1) was identified, and its sequence characteristics, immune response profile, and the antimicrobial activity of its C-terminal peptide TS20 were systematically characterized. The open reading frame of SsTFPI-1 is 867 bp in length, encoding 288 amino acids that comprise a signal peptide, three Kunitz domains, and a low-complexity region. SsTFPI-1 was ubiquitously expressed across all examined tissues, with the highest expression in blood, and its transcript levels were significantly upregulated in the spleen or head kidney following stimulation with Listonella anguillarum, Edwardsiella piscicida, or poly(I:C). Subcellular localization revealed that SsTFPI-1 predominantly localizes in the cytoplasm of EPC cells, and its C-terminus undergoes endogenous cleavage. Antimicrobial assays demonstrated that TS20 exerts pronounced antibacterial activity against Micrococcus luteus with favorable thermal stability. Mechanistically, TS20 disrupts the bacterial cell membrane, enters the cells, binds to DNA, and progressively degrades it. Furthermore, TS20 significantly inhibits SVCV infection in EPC cells by targeting both viral particles and host cells, thereby reducing viral adhesion and entry, while simultaneously downregulating the expression of type I interferon (IFN-I) related genes. In vivo experiments confirmed that TS20 significantly reduced viral loads of SVCV and RBIV-C1 in common carp and turbot, respectively. Collectively, these findings elucidate the immunological role of SsTFPI-1 in S. schlegelii and underscore the potential of TS20 as a promising antimicrobial agent.
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.
Xiaolong Chu, Fengchen Liu, Yingying Liu et al.· Fish and Shellfish Immunolog...· 0 citations
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.
Wenxiaoman Wang, Meng Xu, Yadong Xue et al.· Fish and Shellfish Immunolog...· 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
Nesfatin-1 is an 82-amino acid polypeptide derived from the precursor protein nucleobindin 2 (NUCB2), which is a conserved multifunctional peptide in vertebrates that plays a key role in metabolic regulation, inflammatory response and ferroptosis. Although Nesfatin-1 has been characterized in various vertebrates, from mammals to teleosts, its structural and functional characteristics in primitive vertebrates remain unclear. In this study, Nesfatin-1 orthologs from the most primitive jawless vertebrate lamprey (Lethenteron camtschaticum) were cloned and identified, designated as Lc-Nesfatin-1. Bioinformatic analysis revealed that Nesfatin-1 of lamprey is relatively conserved in both sequence similarity and three-dimensional (3D) structure compared to that of higher vertebrates. Lc-Nesfatin-1 was significantly upregulated under lipopolysaccharide (LPS) stimulation, which also markedly induced the expression of pro-inflammatory cytokines, the key signaling molecule NF-κB, and anti-inflammatory cytokines. Notably, Flow cytometry analysis showed that Lc-Nesfatin-1 did not significantly inhibit LPS-induced intracellular ROS production. Furthermore, Nesfatin-1 serves as an effective negative regulator that significantly inhibits ferroptosis. These results reveal that Lc-Nesfatin-1 plays a critical role in modulating both LPS-induced inflammatory responses and ferroptosis. This study has successfully elucidated the characteristics and functions of Nesfatin-1 in primitive vertebrates and provided valuable insights into the investigation of inflammatory responses and ferroptosis in jawless vertebrates.
Mengqian He, Yaocen Wang, Yitong Li et al.· Fish and Shellfish Immunolog...· 0 citations
This study provides a new perspective for understanding the evolutionary conservation and species specificity of molluscan innate immune signaling pathways and offers important theoretical support for further comprehension of the molecular mechanism of scallop immune defense and promoting the green control of aquatic diseases in marine farming.
Xiaoxue Lin, Zihao Zhang, Xiaolong Chu et al.· Fish and Shellfish Immunolog...· 0 citations
Mycobacterium tuberculosis
is extremely dependent upon lipid-hydrolyzing enzymes to utilize the resources of host lipids and survive within the cell. Rv1063c is a conserved hypothetical protein predicted to share sequence homology with the patatin-like family.
In the present study, the
rv1063c
gene was cloned and heterologously expressed in Escherichia coli BL21 (DE3). Functional enzyme was obtained by solubilizing inclusion bodies in 8 mol·L
−1
urea, refolding via gradient dialysis, and purifying through Ni-affinity chromatography.
Bioinformatic homology analysis indicates Rv1063c is homologous to members of the patatin-like family containing a putative Ser52-Asp166 catalytic dyad, though this residue pair has not been experimentally validated in the present study. The recombinant Rv1063c was mainly expressed as inclusion bodies without significant soluble cytoplasmic expression. Biochemical assays demonstrated the recombinant protein exclusively hydrolyzes medium-chain pnitrophenyl esters (C8–12), with maximum hydrolytic activity toward p-NP-C8 (caprylate), followed by C10 and C12, and exhibited the highest activity at 40 °C and pH 7.0. The enzyme exhibited moderate catalytic efficiency but poor thermal stability. All catalytic data in this paper were obtained from artificial p-nitrophenyl ester substrates, which cannot fully reflect natural lipid substrate preference of mycobacteria in vivo.
This work biochemically characterizes Rv1063c as a medium-chain-specific esterase encoded by a protein homologous to the patatin superfamily. Its physiological and pathogenic roles remain undetermined, and further multi-level experiments are required to clarify its biological functions.
Yuming Song, Songsong Dong, Rumeng Zhai et al.· Frontiers in Microbiology· 0 citations