Parkin is an E3 ubiquitinated ligase that mainly participates in mitophagy and plays an essential biological role in organisms. To investigate Parkin's function in fish, a Parkin homolog was cloned from Epinephelus coioides (EcParkin). The open reading frame (ORF) of EcParkin consists of 1461 nucleotides and encodes a protein of 486 amino acids, with a predicted molecular weight of 53.32 kDa. EcParkin was highly expressed in the heart, kidney, and head kidney of healthy groupers, especially in the heart. The expression levels of EcParkin were upregulated after Singapore grouper iridovirus (SGIV) and red-spotted grouper nervous necrosis virus (RGNNV) infection. Intracellular localization studies revealed that EcParkin is distributed in both the cytoplasm and nucleus of GS cells. Overexpression of EcParkin promoted SGIV and RGNNV replication in vitro, while knockdown of EcParkin inhibited SGIV and RGNNV replication. EcParkin suppressed the promoter activities of IFN-β, ISRE, and NF-κB, as well as the expression of interferon-related factors and inflammatory cytokines. EcParkin was found to colocalize and interact with EcMDA5, EcMAVS, EcTBK1, EcIRF3, and EcIRF7. Additionally, EcParkin enhanced LC3-II production in GS cells. These findings suggest that EcParkin may play a crucial role in the antiviral innate immunity and cellular autophagy of fish.
{"title":"Parkin is a critical factor in grouper immune response to virus infection","authors":"Xiaoxia Lei , Siting Wu , Zhuqing Xu , Qiongyue Xu , Helong Cao , Zhouling Zhan , Qiwei Qin , Jingguang Wei","doi":"10.1016/j.dci.2024.105293","DOIUrl":"10.1016/j.dci.2024.105293","url":null,"abstract":"<div><div>Parkin is an E3 ubiquitinated ligase that mainly participates in mitophagy and plays an essential biological role in organisms. To investigate Parkin's function in fish, a Parkin homolog was cloned from <em>Epinephelus coioides</em> (EcParkin). The open reading frame (ORF) of EcParkin consists of 1461 nucleotides and encodes a protein of 486 amino acids, with a predicted molecular weight of 53.32 kDa. EcParkin was highly expressed in the heart, kidney, and head kidney of healthy groupers, especially in the heart. The expression levels of EcParkin were upregulated after Singapore grouper iridovirus (SGIV) and red-spotted grouper nervous necrosis virus (RGNNV) infection. Intracellular localization studies revealed that EcParkin is distributed in both the cytoplasm and nucleus of GS cells. Overexpression of EcParkin promoted SGIV and RGNNV replication <em>in vitro</em>, while knockdown of EcParkin inhibited SGIV and RGNNV replication. EcParkin suppressed the promoter activities of IFN-β, ISRE, and NF-κB, as well as the expression of interferon-related factors and inflammatory cytokines. EcParkin was found to colocalize and interact with EcMDA5, EcMAVS, EcTBK1, EcIRF3, and EcIRF7. Additionally, EcParkin enhanced LC3-II production in GS cells. These findings suggest that EcParkin may play a crucial role in the antiviral innate immunity and cellular autophagy of fish.</div></div>","PeriodicalId":11228,"journal":{"name":"Developmental and comparative immunology","volume":"162 ","pages":"Article 105293"},"PeriodicalIF":2.7,"publicationDate":"2024-11-26","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142746448","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"农林科学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-11-20DOI: 10.1016/j.dci.2024.105292
Miriam Angulo , Carlos Angulo
Trained immunity has been described as the memory capacity of the innate immune system. Several microbial components have been shown to induce trained immunity. Research on the potential of probiotics to trigger these effects has been limited to a few in vitro studies but remains completely unknown in vivo. Components from the probiotic Debaryomyces hansenii CBS 8339 (Dh) have been shown to induce innate immune memory in goat kids and calves. In the present study, stimulating innate immune cells from newborn goats with probiotic Dh increased respiratory burst activity and nitric oxide production, while cell phagocytosis was unaffected. Glucose uptake was enhanced in goat's cells stimulated with Dh, but lactate production was decreased. In newborn goats, after the training scheme (via oral probiotic administration), cell phagocytosis, nitric oxide production and glycolysis — through the upregulation of AKT and HIF1A gene expression, glucose consumption and lactate production— were enhanced. The expression of IL1B gene was similar between the D. hansenii and control groups. Moreover, the potential long-lasting effects were assessed 30 days after initiation of the training scheme. Cell phagocytosis, respiratory burst and myeloperoxidase activity were enhanced, while glycolytic parameters remained unaffected. Altogether, the results of the present study suggest that the immune training scheme may induce trained immunity by the probiotic D. hansenii in newborn goats. However, our findings were not conclusive regarding the long-lasting (one-month) effects of trained immunity by probiotics.
{"title":"Analysis of the potential long-lasting effects of probiotic Debaryomyces hansenii CBS 8339 on trained immunity in newborn goats","authors":"Miriam Angulo , Carlos Angulo","doi":"10.1016/j.dci.2024.105292","DOIUrl":"10.1016/j.dci.2024.105292","url":null,"abstract":"<div><div>Trained immunity has been described as the memory capacity of the innate immune system. Several microbial components have been shown to induce trained immunity. Research on the potential of probiotics to trigger these effects has been limited to a few <em>in vitro</em> studies but remains completely unknown <em>in vivo</em>. Components from the probiotic <em>Debaryomyces hansenii</em> CBS 8339 (Dh) have been shown to induce innate immune memory in goat kids and calves. In the present study, stimulating innate immune cells from newborn goats with probiotic Dh increased respiratory burst activity and nitric oxide production, while cell phagocytosis was unaffected. Glucose uptake was enhanced in goat's cells stimulated with Dh, but lactate production was decreased. In newborn goats, after the training scheme (via oral probiotic administration), cell phagocytosis, nitric oxide production and glycolysis — through the upregulation of <em>AKT</em> and <em>HIF1A</em> gene expression, glucose consumption and lactate production— were enhanced. The expression of <em>IL1B</em> gene was similar between the <em>D. hansenii</em> and control groups. Moreover, the potential long-lasting effects were assessed 30 days after initiation of the training scheme. Cell phagocytosis, respiratory burst and myeloperoxidase activity were enhanced, while glycolytic parameters remained unaffected. Altogether, the results of the present study suggest that the immune training scheme may induce trained immunity by the probiotic <em>D. hansenii</em> in newborn goats. However, our findings were not conclusive regarding the long-lasting (one-month) effects of trained immunity by probiotics.</div></div>","PeriodicalId":11228,"journal":{"name":"Developmental and comparative immunology","volume":"162 ","pages":"Article 105292"},"PeriodicalIF":2.7,"publicationDate":"2024-11-20","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142692558","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"农林科学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
The clonal triploid ginbuna crucian carp Carassius auratus langsdorfii, a naturally occurring gynogenetic fish, is suitable for cell transplantation studies to reveal the roles of stem cells and immune cells. To ensure long-term traceability of donor cells within recipient fish, we have established a transgenic ginbuna line that expresses green fluorescent protein (GFP). The Xenopus laevis ef1a promoter was introduced for regulating GFP expression. Tol2 transposon-based transgenesis to ginbuna embryos resulted in producing a putative founder fish (F0) in a mosaic fluorescent fashion; the frequency of germline transmission was 14.9%. All embryos of GFP-positive offspring (F1)-derived F2 generation expressed GFP widely across the body. The result of Southern blot analysis showed that the transgene was present on a single DNA fragment of equivalent size among F1 and F2 individuals tested, indicating that the transgene was stably transmitted without translocation. Analysis of the fluorescence intensity of organs obtained from F1 and F2 juveniles using fluorescence microscope showed that eyes, brain, skeletal muscle, heart and gonad exhibited a strong GFP fluorescence while gill, spleen and intestine gave a weak signal; no fluorescence was observed in erythrocytes. Flow cytometric analyses of peripheral leukocytes from F1 and F2 adult fish revealed all cell populations expressed GFP. Scale grafts from the transgenic fish to the wild-type fish exhibited persistent engraftment. Together, our transgenic line can be a powerful tool for studying cellular dynamics by cell transplantation and provide a solid basis for further immunological research advances in teleost.
克隆三倍体银鲫（Carassius auratus langsdorfii）是一种天然雌雄同体鱼类，适合用于细胞移植研究，以揭示干细胞和免疫细胞的作用。为了确保受体鱼体内供体细胞的长期可追溯性，我们建立了一个表达绿色荧光蛋白（GFP）的转基因金线鱼品系。为了调节 GFP 的表达，我们引入了 Xenopus laevis ef1a 启动子。基于 Tol2 转座子的转基因技术可在金枪鱼胚胎中以镶嵌荧光方式产生推定的创始鱼（F0），种系传递频率为 14.9%。GFP 阳性后代（F1）产生的 F2 代的所有胚胎都在全身广泛表达 GFP。Southern 印迹分析的结果表明，受测的 F1 和 F2 个体中，转基因存在于大小相当的单个 DNA 片段上，表明转基因是稳定传递的，没有发生易位。利用荧光显微镜对 F1 和 F2 幼体器官的荧光强度进行分析表明，眼睛、大脑、骨骼肌、心脏和性腺显示出较强的 GFP 荧光，而鳃、脾脏和肠道则显示出微弱的信号；红细胞未观察到荧光。对 F1 和 F2 成鱼外周白细胞的流式细胞分析显示，所有细胞群都表达了 GFP。从转基因鱼到野生型鱼的鳞片移植显示出持续的接种。总之，我们的转基因品系可以成为通过细胞移植研究细胞动态的有力工具，并为进一步推进远洋鱼类的免疫学研究奠定坚实的基础。
{"title":"Establishment of a novel clonal GFP-expressing transgenic ginbuna crucian carp","authors":"Ren Uehara , Shinji Takeda , Daichi Oku , Ryo Sasaki , Masaru Murakami , Hajime Shiba , Fumihiko Katakura , Tadaaki Moritomo","doi":"10.1016/j.dci.2024.105290","DOIUrl":"10.1016/j.dci.2024.105290","url":null,"abstract":"<div><div>The clonal triploid ginbuna crucian carp <em>Carassius auratus langsdorfii</em>, a naturally occurring gynogenetic fish, is suitable for cell transplantation studies to reveal the roles of stem cells and immune cells. To ensure long-term traceability of donor cells within recipient fish, we have established a transgenic ginbuna line that expresses green fluorescent protein (GFP). The <em>Xenopus laevis ef1a</em> promoter was introduced for regulating GFP expression. <em>Tol2</em> transposon-based transgenesis to ginbuna embryos resulted in producing a putative founder fish (F0) in a mosaic fluorescent fashion; the frequency of germline transmission was 14.9%. All embryos of GFP-positive offspring (F1)-derived F2 generation expressed GFP widely across the body. The result of Southern blot analysis showed that the transgene was present on a single DNA fragment of equivalent size among F1 and F2 individuals tested, indicating that the transgene was stably transmitted without translocation. Analysis of the fluorescence intensity of organs obtained from F1 and F2 juveniles using fluorescence microscope showed that eyes, brain, skeletal muscle, heart and gonad exhibited a strong GFP fluorescence while gill, spleen and intestine gave a weak signal; no fluorescence was observed in erythrocytes. Flow cytometric analyses of peripheral leukocytes from F1 and F2 adult fish revealed all cell populations expressed GFP. Scale grafts from the transgenic fish to the wild-type fish exhibited persistent engraftment. Together, our transgenic line can be a powerful tool for studying cellular dynamics by cell transplantation and provide a solid basis for further immunological research advances in teleost.</div></div>","PeriodicalId":11228,"journal":{"name":"Developmental and comparative immunology","volume":"162 ","pages":"Article 105290"},"PeriodicalIF":2.7,"publicationDate":"2024-11-19","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142686405","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"农林科学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-11-12DOI: 10.1016/j.dci.2024.105288
Qiongyao Zeng , Yiyang Tang , Yujun Liu , Ye Yang , Pingyuan Li , Zejun Zhou , Qinbo Qin
Secreted phospholipase A2 family protein (sPLA2) is associated with immune response and plays a critical role in the regulation of gut homeostasis. However, whether sPLA2 is involved in innate immunity in teleost is essentially unknown. For this purpose, we reported the identification of a classical sPLA2 in grass carp (CisPLA2) and elucidated its role in the antibacterial immunity in this study. The result of bioinformatics analysis showed that mammalian sPLA2-IIA is the most similar homologue to CisPLA2. CisPLA2 is expressed in a variety of tissues, including liver and gut, and is significantly upregulated in response to Aeromonas hydrophila infection. Recombinant CisPLA2 protein (rCisPLA2) showed significant antibacterial activity against A. hydrophila by enhancing the phagocytosis of host phagocytes in vitro. Moreover, rCisPLA2 induces significant expression of the antimicrobial molecules and tight junctions in the gut during bacterial infection. Fish administered with rCisPLA2 significantly alleviates the gut permeability and apoptosis. In addition, rCisPLA2 preserves the morphology of the gut mucosa and limits the colonization of A. hydrophila in systemic immune organs. These results indicate that CisPLA2 plays a crucial role in the regulation of gut mucosal barrier, and thus has a potential application for antimicrobial immunity in fish.
{"title":"A recombinant sPLA2 protein promotes gut mucosal barrier against bacterial infection in fish","authors":"Qiongyao Zeng , Yiyang Tang , Yujun Liu , Ye Yang , Pingyuan Li , Zejun Zhou , Qinbo Qin","doi":"10.1016/j.dci.2024.105288","DOIUrl":"10.1016/j.dci.2024.105288","url":null,"abstract":"<div><div>Secreted phospholipase A2 family protein (sPLA2) is associated with immune response and plays a critical role in the regulation of gut homeostasis. However, whether sPLA2 is involved in innate immunity in teleost is essentially unknown. For this purpose, we reported the identification of a classical sPLA2 in grass carp (<em>Ci</em>sPLA2) and elucidated its role in the antibacterial immunity in this study. The result of bioinformatics analysis showed that mammalian sPLA2-IIA is the most similar homologue to <em>Ci</em>sPLA2. <em>CisPLA2</em> is expressed in a variety of tissues, including liver and gut, and is significantly upregulated in response to <em>Aeromonas hydrophila</em> infection. Recombinant <em>Ci</em>sPLA2 protein (r<em>Ci</em>sPLA2) showed significant antibacterial activity against <em>A</em>. <em>hydrophila</em> by enhancing the phagocytosis of host phagocytes <em>in vitro</em>. Moreover, r<em>Ci</em>sPLA2 induces significant expression of the antimicrobial molecules and tight junctions in the gut during bacterial infection. Fish administered with r<em>Ci</em>sPLA2 significantly alleviates the gut permeability and apoptosis. In addition, r<em>Ci</em>sPLA2 preserves the morphology of the gut mucosa and limits the colonization of <em>A</em>. <em>hydrophila</em> in systemic immune organs. These results indicate that <em>Ci</em>sPLA2 plays a crucial role in the regulation of gut mucosal barrier, and thus has a potential application for antimicrobial immunity in fish.</div></div>","PeriodicalId":11228,"journal":{"name":"Developmental and comparative immunology","volume":"162 ","pages":"Article 105288"},"PeriodicalIF":2.7,"publicationDate":"2024-11-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142616551","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"农林科学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-11-12DOI: 10.1016/j.dci.2024.105286
Erin Glass , Stephan L. Robinson , Emily E. Rosowski
Pattern recognition receptors (PRRs) such as C-type lectin receptors (CLRs) and Toll-like receptors (TLRs) are used by hosts to recognize pathogen-associated molecular patterns (PAMPs) in microorganisms and to initiate innate immune responses. While PRRs exist across invertebrate and vertebrate species, the functional homology of many of these receptors is still unclear. In this study, we investigate the innate immune response of zebrafish larvae to zymosan, a β-glucan-containing particle derived from fungal cell walls. Macrophages and neutrophils robustly respond to zymosan and are required for zymosan-induced activation of the NF-κB transcription factor. Full activation of NF-κB in response to zymosan depends on Card9/Syk and Myd88, conserved CLR and TLR adaptor proteins, respectively. Two putative CLRs, Clec4c and Sclra, are both required for maximal sensing of zymosan and NF-κB activation but not required for inflammatory gene expression. Altogether, we identify conserved PRRs and PRR signaling pathways in larval zebrafish that promote recognition of fungal PAMPs. These results inform modeling of human fungal infections in zebrafish and increase our knowledge of the evolution and conservation of PRR pathways in vertebrates.
{"title":"Zebrafish use conserved CLR and TLR signaling pathways to respond to fungal PAMPs in zymosan","authors":"Erin Glass , Stephan L. Robinson , Emily E. Rosowski","doi":"10.1016/j.dci.2024.105286","DOIUrl":"10.1016/j.dci.2024.105286","url":null,"abstract":"<div><div>Pattern recognition receptors (PRRs) such as C-type lectin receptors (CLRs) and Toll-like receptors (TLRs) are used by hosts to recognize pathogen-associated molecular patterns (PAMPs) in microorganisms and to initiate innate immune responses. While PRRs exist across invertebrate and vertebrate species, the functional homology of many of these receptors is still unclear. In this study, we investigate the innate immune response of zebrafish larvae to zymosan, a β-glucan-containing particle derived from fungal cell walls. Macrophages and neutrophils robustly respond to zymosan and are required for zymosan-induced activation of the NF-κB transcription factor. Full activation of NF-κB in response to zymosan depends on Card9/Syk and Myd88, conserved CLR and TLR adaptor proteins, respectively. Two putative CLRs, Clec4c and Sclra, are both required for maximal sensing of zymosan and NF-κB activation but not required for inflammatory gene expression. Altogether, we identify conserved PRRs and PRR signaling pathways in larval zebrafish that promote recognition of fungal PAMPs. These results inform modeling of human fungal infections in zebrafish and increase our knowledge of the evolution and conservation of PRR pathways in vertebrates.</div></div>","PeriodicalId":11228,"journal":{"name":"Developmental and comparative immunology","volume":"162 ","pages":"Article 105286"},"PeriodicalIF":2.7,"publicationDate":"2024-11-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142616554","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"农林科学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-11-12DOI: 10.1016/j.dci.2024.105289
D.C.G. Rodrigo , H.M.V. Udayantha , D.S. Liyanage , W.K.M. Omeka , Y.K. Kodagoda , H.A.C.R. Hanchapola , M.A.H. Dilshan , G.A.N.P. Ganepola , W.A.D.L.R. Warnakula , Gaeun Kim , Jeongeun Kim , Jihun Lee , Qiang Wan , Jehee Lee
Peroxiredoxin 5 (Prdx5) is the last recognized member of Prdx family. It is a unique, atypical, 2-Cys antioxidant enzyme, protecting cells from death caused by reactive oxygen species (ROS). In this study, the Prdx5 ortholog of Amphiprion clarkii (AcPrdx5) was identified and characterized to explore its specific structural features and functional properties. The open reading frame of AcPrdx5 is 573 bp long and encodes 190 amino acids containing a mitochondrial targeting sequence, thioredoxin domain, and two conserved cysteine residues responsible for antioxidant function. The predicted molecular weight and theoretical isoelectric point of AcPrdx5 are 20.3 kDa and 9.01, respectively. AcPrdx5 sequences were found to be highly conserved across the other orthologs from various organisms and it distinctively clustered within the fish Prdx5 subclade of the phylogenetic tree. The expression of AcPrdx5 was ubiquitously detected among twelve tested tissues, with the highest level in the brain. Furthermore, the mRNA levels of AcPrdx5 in the blood and head-kidney tissues were significantly (p < 0.05) upregulated following polyinosinic-polycytidylic acid (Poly I:C), lipopolysaccharide (LPS), and Vibrio harveyi immune challenge. A concentration-dependent antioxidant potential of recombinant AcPrdx5 was observed in insulin disulfide bond reduction, heavy metal detoxification, free radical and hydrogen peroxide (H2O2) scavenging assays. Additionally, AcPrdx5 overexpression in fathead minnow (FHM) cells upregulated the antioxidant-associated gene (Rrm1, MAPK, SOD2, and PRDX1) expression after H2O2 treatment, and promoted cell viability upon arsenic (As) exposure. In macrophages, AcPrdx5 overexpression effectively suppressed substantial nitric oxide production under lipopolysaccharide treatment. Collectively, our results suggest that AcPrdx5 may play roles in both antioxidant defense system and innate immune response against pathogenic invasions in A. clarkii.
{"title":"Functional characterization of peroxiredoxin 5 from yellowtail clownfish (Amphiprion clarkii): Immunological expression assessment, antioxidant activities, heavy metal detoxification, and nitrosative stress mitigation","authors":"D.C.G. Rodrigo , H.M.V. Udayantha , D.S. Liyanage , W.K.M. Omeka , Y.K. Kodagoda , H.A.C.R. Hanchapola , M.A.H. Dilshan , G.A.N.P. Ganepola , W.A.D.L.R. Warnakula , Gaeun Kim , Jeongeun Kim , Jihun Lee , Qiang Wan , Jehee Lee","doi":"10.1016/j.dci.2024.105289","DOIUrl":"10.1016/j.dci.2024.105289","url":null,"abstract":"<div><div>Peroxiredoxin 5 (<em>Prdx5</em>) is the last recognized member of Prdx family. It is a unique, atypical, 2-Cys antioxidant enzyme, protecting cells from death caused by reactive oxygen species (ROS). In this study, the Prdx5 ortholog of <em>Amphiprion clarkii</em> (<em>AcPrdx5</em>) was identified and characterized to explore its specific structural features and functional properties. The open reading frame of AcPrdx5 is 573 bp long and encodes 190 amino acids containing a mitochondrial targeting sequence, thioredoxin domain, and two conserved cysteine residues responsible for antioxidant function. The predicted molecular weight and theoretical isoelectric point of AcPrdx5 are 20.3 kDa and 9.01, respectively. AcPrdx5 sequences were found to be highly conserved across the other orthologs from various organisms and it distinctively clustered within the fish <em>Prdx5</em> subclade of the phylogenetic tree. The expression of <em>AcPrdx5</em> was ubiquitously detected among twelve tested tissues, with the highest level in the brain. Furthermore, the mRNA levels of <em>AcPrdx5</em> in the blood and head-kidney tissues were significantly (<em>p < 0.05</em>) upregulated following polyinosinic-polycytidylic acid (Poly I:C), lipopolysaccharide (LPS), and <em>Vibrio harveyi</em> immune challenge<em>.</em> A concentration-dependent antioxidant potential of recombinant AcPrdx5 was observed in insulin disulfide bond reduction, heavy metal detoxification, free radical and hydrogen peroxide (H<sub>2</sub>O<sub>2</sub>) scavenging assays. Additionally, AcPrdx5 overexpression in fathead minnow (FHM) cells upregulated the antioxidant-associated gene (Rrm1, MAPK, SOD2, and PRDX1) expression after H<sub>2</sub>O<sub>2</sub> treatment, and promoted cell viability upon arsenic (As) exposure. In macrophages, AcPrdx5 overexpression effectively suppressed substantial nitric oxide production under lipopolysaccharide treatment. Collectively, our results suggest that AcPrdx5 may play roles in both antioxidant defense system and innate immune response against pathogenic invasions in <em>A</em>. <em>clarkii</em>.</div></div>","PeriodicalId":11228,"journal":{"name":"Developmental and comparative immunology","volume":"162 ","pages":"Article 105289"},"PeriodicalIF":2.7,"publicationDate":"2024-11-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142616552","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"农林科学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-11-08DOI: 10.1016/j.dci.2024.105287
Victoria L. Rhodes , Robert M. Waterhouse , Kristin Michel
Innate immunity in mosquitoes has received much attention due to its potential impact on vector competence for vector-borne disease pathogens, including malaria parasites. The nuclear factor (NF)-κB-dependent Toll pathway is a major regulator of innate immunity in insects. In mosquitoes, this pathway controls transcription of the majority of the known canonical humoral immune effectors, mediates anti-bacterial, anti-fungal and anti-viral immune responses, and contributes to malaria parasite killing. However, besides initial gene annotation of putative Toll pathway members and genetic analysis of the contribution of few key components to immunity, the molecular make-up and function of the Toll pathway in mosquitoes is largely unexplored. To facilitate functional analyses of the Toll pathway in mosquitoes, we report here manually annotated and refined gene models of Toll-like receptors and all putative components of the intracellular signal transduction cascade across 19 anopheline genomes, and in two culicine genomes. In addition, based on phylogenetic analyses, we identified differing levels of evolutionary constraint across the intracellular Toll pathway members, and identified a recent radiation of TOLL1/5 within the Anopheles gambiae complex. Together, this study provides insight into the evolution of TLRs and the putative members of the intracellular signal transduction cascade within the genus Anopheles.
{"title":"The molecular toll pathway repertoire in anopheline mosquitoes","authors":"Victoria L. Rhodes , Robert M. Waterhouse , Kristin Michel","doi":"10.1016/j.dci.2024.105287","DOIUrl":"10.1016/j.dci.2024.105287","url":null,"abstract":"<div><div>Innate immunity in mosquitoes has received much attention due to its potential impact on vector competence for vector-borne disease pathogens, including malaria parasites. The nuclear factor (NF)-κB-dependent Toll pathway is a major regulator of innate immunity in insects. In mosquitoes, this pathway controls transcription of the majority of the known canonical humoral immune effectors, mediates anti-bacterial, anti-fungal and anti-viral immune responses, and contributes to malaria parasite killing. However, besides initial gene annotation of putative Toll pathway members and genetic analysis of the contribution of few key components to immunity, the molecular make-up and function of the Toll pathway in mosquitoes is largely unexplored. To facilitate functional analyses of the Toll pathway in mosquitoes, we report here manually annotated and refined gene models of Toll-like receptors and all putative components of the intracellular signal transduction cascade across 19 anopheline genomes, and in two culicine genomes. In addition, based on phylogenetic analyses, we identified differing levels of evolutionary constraint across the intracellular Toll pathway members, and identified a recent radiation of TOLL1/5 within the <em>Anopheles gambiae</em> complex. Together, this study provides insight into the evolution of TLRs and the putative members of the intracellular signal transduction cascade within the genus <em>Anopheles</em>.</div></div>","PeriodicalId":11228,"journal":{"name":"Developmental and comparative immunology","volume":"162 ","pages":"Article 105287"},"PeriodicalIF":2.7,"publicationDate":"2024-11-08","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142616555","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"农林科学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
One of the most highly induced genes in zebrafish (Danio rerio) larvae after infection with the nodavirus red-spotted grouper nervous necrosis virus (RGNNV) was a member of the immunoglobulin superfamily (IgSF), which has remained uncharacterized and erroneously annotated in zebrafish and other fish species as galectin 17 (lgals17). We characterized this gene and named it immunoglobulin (Ig)-like domain-containing protein (igldcp), a new member of the IgSF that does not possess orthologs in mammals. Igldcp expression is induced by viral infection and it belongs to the group of interferon-stimulated genes (ISGs). In vitro overexpression of igldcp decreased RGNNV replication, whereas in vivo knockdown of this gene had the opposite effect, resulting in increased larval mortality. RNA-Seq analyses of larvae overexpressing igldcp in the absence or presence of infection with RGNNV showed that the main processes affected by Igldcp could be directly involved in the regulation of various cellular processes associated with the modulation of the immune system.
{"title":"Characterization of a fish-specific immunoglobulin-like domain-containing protein (Igldcp) in zebrafish (Danio rerio) induced after nodavirus infection","authors":"Nieves Martínez-López, Patricia Pereiro, Amaro Saco, Raquel Lama, Antonio Figueras, Beatriz Novoa","doi":"10.1016/j.dci.2024.105285","DOIUrl":"10.1016/j.dci.2024.105285","url":null,"abstract":"<div><div>One of the most highly induced genes in zebrafish (<em>Danio rerio</em>) larvae after infection with the nodavirus red-spotted grouper nervous necrosis virus (RGNNV) was a member of the immunoglobulin superfamily (IgSF), which has remained uncharacterized and erroneously annotated in zebrafish and other fish species as <em>galectin 17</em> (<em>lgals17</em>). We characterized this gene and named it <em>immunoglobulin (Ig)-like domain-containing protein</em> (<em>igldcp</em>), a new member of the IgSF that does not possess orthologs in mammals. Igldcp expression is induced by viral infection and it belongs to the group of interferon-stimulated genes (ISGs). <em>In vitro</em> overexpression of <em>igldcp</em> decreased RGNNV replication, whereas <em>in vivo</em> knockdown of this gene had the opposite effect, resulting in increased larval mortality. RNA-Seq analyses of larvae overexpressing <em>igldcp</em> in the absence or presence of infection with RGNNV showed that the main processes affected by Igldcp could be directly involved in the regulation of various cellular processes associated with the modulation of the immune system.</div></div>","PeriodicalId":11228,"journal":{"name":"Developmental and comparative immunology","volume":"162 ","pages":"Article 105285"},"PeriodicalIF":2.7,"publicationDate":"2024-11-06","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142603423","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"农林科学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-11-02DOI: 10.1016/j.dci.2024.105284
Xue Kong , Wei Wang , Sunan Xia , Ying Zhi , Yuefeng Cai , Haibin Zhang , Xin Shen
Within cold seep environments, the Vesicomyidae clam emerges as a prevalent species, distinguished by its symbiotic relationship with microorganisms housed within its organ gill. Given the extreme conditions and the symbiotic nature of this association, investigating the host's immune genes, particularly immune recognition receptors, is essential for understanding their role in facilitating host-symbiotic interactions. Three short peptidoglycan recognition proteins (PGRPs) were identified in the clam. AmPGRP-S1, -S2, and -S3 were found to possess conserved amidase binding sites and Zn2+ binding sites. Quantitative Real-time PCR (qRT-PCR) analysis revealed differential expression patterns among the PGRPs. AmPGRP-S1 and AmPGRP-S2 exhibited elevated expression levels in the gill, while AmPGRP-S3 displayed the highest expression in the adductor muscle. Functional experiments demonstrated that recombinant AmPGRP-S1, -S2, and -S3 (rAmPGRPs) exhibited binding capabilities to both L-PGN and D-PGN (peptidoglycan). Notably, rAmPGRP-S1 and -S2 possessed Zn2+-independent amidase activity, while rAmPGRP-S3 lacked this enzymatic function. rAmPGRPs were shown to bind to five different bacterial species. Among these, rAmPGRP-S1 inhibited Escherichia coli and Bacillus subtilis, while rAmPGRP-S2 and -S3 inhibited Bacillus subtilis in the absence of Zn2+. In the presence of Zn2+, rAmPGRP-S1 and -S2 exhibited enhanced inhibitory activity against Staphylococcus aureus or Bacillus subtilis. These findings suggest that AmPGRPs may play a pivotal role in mediating the interaction between the host and endosymbiotic bacteria, functioning as PGN and microbe receptors, antibacterial effectors, and regulators of host-microbe symbiosis. These results contribute to our understanding of the adaptive mechanisms of deep-sea organisms to the challenging cold seep environments.
{"title":"Molecular and functional characterization of short peptidoglycan recognition proteins in Vesicomyidae clam","authors":"Xue Kong , Wei Wang , Sunan Xia , Ying Zhi , Yuefeng Cai , Haibin Zhang , Xin Shen","doi":"10.1016/j.dci.2024.105284","DOIUrl":"10.1016/j.dci.2024.105284","url":null,"abstract":"<div><div>Within cold seep environments, the Vesicomyidae clam emerges as a prevalent species, distinguished by its symbiotic relationship with microorganisms housed within its organ gill. Given the extreme conditions and the symbiotic nature of this association, investigating the host's immune genes, particularly immune recognition receptors, is essential for understanding their role in facilitating host-symbiotic interactions. Three short peptidoglycan recognition proteins (PGRPs) were identified in the clam. AmPGRP-S1, -S2, and -S3 were found to possess conserved amidase binding sites and Zn<sup>2+</sup> binding sites. Quantitative Real-time PCR (qRT-PCR) analysis revealed differential expression patterns among the PGRPs. AmPGRP-S1 and AmPGRP-S2 exhibited elevated expression levels in the gill, while AmPGRP-S3 displayed the highest expression in the adductor muscle. Functional experiments demonstrated that recombinant AmPGRP-S1, -S2, and -S3 (rAmPGRPs) exhibited binding capabilities to both L-PGN and D-PGN (peptidoglycan). Notably, rAmPGRP-S1 and -S2 possessed Zn<sup>2+</sup>-independent amidase activity, while rAmPGRP-S3 lacked this enzymatic function. rAmPGRPs were shown to bind to five different bacterial species. Among these, rAmPGRP-S1 inhibited <em>Escherichia coli</em> and <em>Bacillus subtilis</em>, while rAmPGRP-S2 and -S3 inhibited <em>Bacillus subtilis</em> in the absence of Zn<sup>2<em>+</em></sup>. In the presence of Zn<sup>2+</sup>, rAmPGRP-S1 and -S2 exhibited enhanced inhibitory activity against <em>Staphylococcus aureus</em> or <em>Bacillus subtilis</em>. These findings suggest that AmPGRPs may play a pivotal role in mediating the interaction between the host and endosymbiotic bacteria, functioning as PGN and microbe receptors, antibacterial effectors, and regulators of host-microbe symbiosis. These results contribute to our understanding of the adaptive mechanisms of deep-sea organisms to the challenging cold seep environments.</div></div>","PeriodicalId":11228,"journal":{"name":"Developmental and comparative immunology","volume":"162 ","pages":"Article 105284"},"PeriodicalIF":2.7,"publicationDate":"2024-11-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142567814","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"农林科学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Membrane-associated Ring-CH 5 (MARCH5) is a mitochondrial E3 ubiquitin ligase playing a key role in the regulation of mitochondrial dynamics. In mammals, MARCH5 negatively regulates mitochondrial antiviral signaling (MAVS) protein aggregation during viral infection and hampers downstream type I interferon signaling to prevent excessive immune activation. However, its precise functional role in the teleost immune system remains unclear. This study investigated the molecular characteristics and immune response of the MARCH5 ortholog in Amphiprion clarkii (A. clarkii; AcMARCH5). The predicted AcMARCH5 protein sequence consists of 287 amino acids with a molecular weight of 32.02 kDa and a theoretical isoelectric point of 9.11. It contains four C-terminal transmembrane (TM) domains and an N-terminal RING cysteine-histidine (CH) domain, which directly regulates ubiquitin transfer. Multiple sequence alignment revealed a high level of conservation between AcMARCH5 and its orthologs in other vertebrate species. Under normal physiological conditions, AcMARCH5 showed the highest mRNA expression in the muscle, brain, and kidney tissues of A. clarkii. Upon stimulation with polyinosinic:polycytidylic acid (Poly I:C), lipopolysaccharide (LPS), and Vibrio harveyi, AcMARCH5 expression was drastically modulated. Functional assays showed that overexpression of AcMARCH5 in fathead minnow (FHM) cells downregulated antiviral gene expression, accompanied by enhanced viral hemorrhagic septicemia virus (VHSV) replication. In murine macrophages, AcMARCH5 overexpression markedly reduced the production of pro-inflammatory cytokines in response to poly I:C treatment. Additionally, AcMARCH5 exhibited an anti-apoptotic effect in H2O2-treated FHM cells. Collectively, these results suggest that AcMARCH5 may play a role in maintaining cellular homeostasis under disease and stress conditions in A. clarkii.
膜相关环-CH 5（MARCH5）是一种线粒体 E3 泛素连接酶，在线粒体动力学调控中发挥着关键作用。在哺乳动物中，MARCH5 在病毒感染期间负向调节线粒体抗病毒信号转导（MAVS）蛋白的聚集，并阻碍下游 I 型干扰素信号转导，以防止过度的免疫激活。然而，它在远洋鱼类免疫系统中的确切功能作用仍不清楚。本研究调查了克氏栉水母（Amphiprion clarkii；AcMARCH5）MARCH5直向同源物的分子特征和免疫反应。预测的 AcMARCH5 蛋白序列由 287 个氨基酸组成，分子量为 32.02 kDa，理论等电点为 9.11。它包含四个 C 端跨膜（TM）结构域和一个 N 端 RING 半胱氨酸-组氨酸（CH）结构域，后者直接调节泛素的转移。多重序列比对显示，AcMARCH5与其在其他脊椎动物中的同源物之间存在高度保守性。在正常生理条件下，AcMARCH5 在克氏原鲤的肌肉、脑和肾组织中的 mRNA 表达量最高。在多聚肌苷酸：多聚胞苷酸（Poly I:C）、脂多糖（LPS）和哈维弧菌的刺激下，AcMARCH5的表达受到了极大的影响。功能测试显示，在黑头鲦鱼（FHM）细胞中过表达 AcMARCH5 会降低抗病毒基因的表达，同时增强病毒性出血性败血症病毒（VHSV）的复制。在小鼠巨噬细胞中，AcMARCH5 的过表达明显减少了聚 I:C 处理后促炎细胞因子的产生。此外，AcMARCH5 在 H2O2 处理的 FHM 细胞中表现出抗凋亡作用。总之，这些结果表明，AcMARCH5 可能在 A. clarkii 疾病和应激条件下维持细胞稳态方面发挥作用。
{"title":"Molecular depiction and functional delineation of E3 ubiquitin ligase MARCH5 in yellowtail clownfish (Amphiprion clarkii)","authors":"B.P.M. Vileka Jayamali , H.M.S.M. Wijerathna , D.M.K.P. Sirisena , H.A.C.R. Hanchapola , W.A.D.L.R. Warnakula , U.P.E. Arachchi , D.S. Liyanage , Sumi Jung , Qiang Wan , Jehee Lee","doi":"10.1016/j.dci.2024.105283","DOIUrl":"10.1016/j.dci.2024.105283","url":null,"abstract":"<div><div>Membrane-associated Ring-CH 5 (MARCH5) is a mitochondrial E3 ubiquitin ligase playing a key role in the regulation of mitochondrial dynamics. In mammals, MARCH5 negatively regulates mitochondrial antiviral signaling (MAVS) protein aggregation during viral infection and hampers downstream type I interferon signaling to prevent excessive immune activation. However, its precise functional role in the teleost immune system remains unclear. This study investigated the molecular characteristics and immune response of the MARCH5 ortholog in <em>Amphiprion clarkii</em> (<em>A. clarkii</em>; AcMARCH5). The predicted AcMARCH5 protein sequence consists of 287 amino acids with a molecular weight of 32.02 kDa and a theoretical isoelectric point of 9.11. It contains four C-terminal transmembrane (TM) domains and an N-terminal RING cysteine-histidine (CH) domain, which directly regulates ubiquitin transfer. Multiple sequence alignment revealed a high level of conservation between AcMARCH5 and its orthologs in other vertebrate species. Under normal physiological conditions, <em>AcMARCH5</em> showed the highest mRNA expression in the muscle, brain, and kidney tissues of <em>A. clarkii</em>. Upon stimulation with polyinosinic:polycytidylic acid (Poly I:C), lipopolysaccharide (LPS), and <em>Vibrio harveyi</em>, <em>AcMARCH5</em> expression was drastically modulated. Functional assays showed that overexpression of AcMARCH5 in fathead minnow (FHM) cells downregulated antiviral gene expression, accompanied by enhanced viral hemorrhagic septicemia virus (VHSV) replication. In murine macrophages, AcMARCH5 overexpression markedly reduced the production of pro-inflammatory cytokines in response to poly I:C treatment. Additionally, AcMARCH5 exhibited an anti-apoptotic effect in H<sub>2</sub>O<sub>2</sub>-treated FHM cells. Collectively, these results suggest that AcMARCH5 may play a role in maintaining cellular homeostasis under disease and stress conditions in <em>A. clarkii</em>.</div></div>","PeriodicalId":11228,"journal":{"name":"Developmental and comparative immunology","volume":"162 ","pages":"Article 105283"},"PeriodicalIF":2.7,"publicationDate":"2024-10-29","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142557392","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"农林科学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
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