Response In Teleost Fish Research Articles

MiR-144 affects the immune response and activation of inflammatory responses in Cynoglossus semilaevis by regulating the expression of CsMAPK6

MicroRNAs are increasingly recognized for their pivotal role in the immune system, yet the specific regulatory functions of fish-derived microRNAs remain largely unexplored. In this research, we discovered a novel miRNA, Cse-miR-144, in the Chinese tongue sole (Cynoglossus semilaevis), characterized by a 73-base pair precursor and a 21-nucleotide mature sequence. Our findings revealed that the expression of Cse-miR-144 was notably inhibited by various Vibrio species. Utilizing bioinformatics and dual-luciferase assay techniques, we established that the pro-inflammatory cytokine gene CsMAPK6 is a direct target of Cse-miR-144. Subsequent in vitro and in vivo western blotting analyses confirmed that Cse-miR-144 can effectively reduce the protein levels of CsMAPK6 post-transcriptionally. Moreover, CsMAPK6 is known to be involved in the activation of the Nuclear Factor kappa-light-chain-enhancer of activated B cells (NF-kB). Additional investigations using qPCR and ELISA demonstrated that suppression of Cse-miR-144 leads to an upsurge in the liver mRNA levels of various immune genes (including MYD88, TRAF6, NF-κB, TRAF2, TRAF3, and TNF), alongside a marked increase in the production and secretion of pro-inflammatory cytokines (IL-1β, IL-6, and IL-8) in the bloodstream of C. semilaevis. These findings collectively underscore the potential of Cse-miR-144 as a key inhibitor of CsMAPK and its crucial role in modulating the immune and inflammatory responses in teleost fish. Compared to the siRNA, miRNA is a better tool in controlling the expression of target gene with a lower cost.

Galectin 9 restricts viral replication in teleost via autophagy-antiviral pathway and polarizes M2 macrophages for anti-inflammatory response: New insights into functional properties of fish Galectin-9 from Planiliza haematocheilus

Galectin 9 (Gal9) is a tandem repeat type ß-galactoside-binding galectin that mediates various cellular biochemical and immunological functions. Many studies have investigated the functional properties of Gal9 in mammals; however, knowledge of fish Gal9 is limited to antibacterial studies. In this context, our aim was to clone Gal9 from Planiliza haematocheilus (PhGal9) and investigate its structural and functional characteristics. We discovered the PhGal9 open reading frame, which was 969 base pairs long and encoded a 322 amino acid protein. PhGal9 had a projected molecular weight of 35.385 kDa but no signal peptide sequence. PhGal9 mRNA was ubiquitously produced in all investigated tissues but was predominant in the intestine, spleen, and brain. Its mRNA expression was increased in response to stimulation by Poly(I:C), LPS, and L. garvieae. The rPhGal9 exhibited a dose-dependent agglutination potential toward gram-positive and gram-negative bacteria at a minimum concentration of 50 μg/mL. Overexpression of PhGal9 promoted M2-like phenotype changes in mouse macrophages, and RT-qPCR analysis of M1 and M2 marker genes confirmed M2 polarization with upregulation of M2 marker genes. In the antiviral assay, the expression levels of Viral Hemorrhagic Septicemia Virus (VHSV) glycoproteins, phosphoproteins, nucleoproteins, non-virion proteins, matrix proteins, and RNA polymerase were significantly reduced in PhGal9-overexpressed cells. Furthermore, the mRNA expression of autophagic genes (sqstm1, tax1bp1b, rnf13, lc3, and atg5) and antiviral genes (viperin) were upregulated in PhGal9 overexpressed cells. For the first time in teleosts, our study demonstrated that PhGal9 promotes M2 macrophage polarization by upregulating M2-associated genes (egr2 and cmyc) and suppressing M1-associated genes (iNOS and IL-6). Furthermore, our results show that exogenous and endogenous PhGal9 prevented VHSV attachment and replication by neutralizing virion and autophagy, respectively. Gal9 may be a potent modulator of the antimicrobial immune response in teleost fish.

March2 negatively regulates antiviral immune response by targeting tbk1 in grass carp (Ctenopharyngodon idella).

Grass carp is one of the most economically important fish species. Hemorrhagic diseases caused by grass carp reovirus (GCRV) can seriously damage the economic yield of grass carp. Therefore, antiviral research on grass carp is urgently needed. Membrane-associated RING-CH2 (MARCH2) negatively regulates the innate immune response in mice. However, little is known about the role of march2 in the antiviral innate immune response in teleost fish. Our present study showed that march2 has high homology in grass carp, its orthologs, and mammals, and has the same amino acid sequence in grass carp and crucian carp. Overexpression of Cimarch2 (Ctenopharyngodon idella march2) significantly inhibited interferon (IFN) activation induced by Polyinosinic-polycytidylic acid (poly I: C), spring viremia of carp virus (SVCV), and GCRV. However, knocking down Cimarch2 enhanced the activation of IFN induced by poly I: C, SVCV, and GCRV. Overexpression of Cimarch2 can promotes viral replication. Mechanistically, Cimarch2 tightly bound to TANK-binding kinase 1 (tbk1) and downregulated tbk1 through the proteasome pathway. Our results demonstrated the potential role of Cimarch2 in the antiviral breeding of grass carp.

HnRNPM suppressed IRF7-mediated IFN signaling in the antiviral innate immunity in triploid hybrid fish

Mammalian heterogeneous nuclear ribonucleoproteins M (hnRNPM) is a critical splicing regulatory protein that has been reported to negatively regulate the RLR signaling pathway by impairing the binding of RIG-I and MDA5 to viral RNA. To explore the role of hnRNPM in the antiviral innate immune response in teleost fish, the hnRNPM homologue of triploid fish (3nhnRNPM) has been cloned and identified in this paper. The CDS of 3nhnRNPM gene is composed of 2016 nucleotides and encodes 671 amino acids. 3nhnRNPM migrated around 71 kDa in immunoblotting assay and was mainly detected in the nucleus in nucleo-cytoplasmic separation assay and immunofluorescent staining test. When 3nhnRNPM and 3nIRF7 were co-expressed in EPC cells, 3nhnRNPM significantly reduced the 3nIRF7-induced interferon (IFN) promoter transcription. Correspondingly, the mRNA levels of the SVCV-M, -N, -P, and -G genes were noteworthily enhanced, but the transcription levels of epcIFNφ1, epcMx1, epcPKR, and epcISG15 were dramatically decreased. Additionally, the knockdown of 3nhnRNPM resulted in restricted SVCV replication and enhanced host cell antiviral activity. Furthermore, the association between 3nhnRNPM and 3nIRF7 has been identified by the co-immunoprecipitation assay. In addition, we found that 3nIRF7 was detained in the nucleus when co-expressed with 3nhnRNPM. To sum up, our data supported the conclusion that 3nhnRNPM suppressed 3nIRF7-mediated IFN signaling in the antiviral innate immunity.

Circular RNA circPlce1 regulates innate immune response in miiuy croaker, Miichthys miiuy

In recent years, more and more researchers have devoted to the study of circular RNAs (circRNAs) in noncoding RNAs. As an important regulator in a variety of biological processes, circRNAs are relatively abundant in the study of mammals, while research in lower vertebrates is still lacking. In this study, we found a circRNA, circPlce1, related to innate immune response in Miichthys miiuy (miiuy croaker). The experimental results confirmed that circPlce1 could promote the production of antiviral genes and inflammatory response under the stimulation of poly (I: C) and LPS. We also confirmed that circPlce1 can promote NF-κB and IRF3 pathways through luciferase reporter assay experiment. In addition, we also found that circPlce1 can promote cell proliferation and improve cell viability. In conclusion, our results showed that circPlce1 plays an active role in regulating inflammatory response, cell proliferation and cell viability, providing a foundation for the study of the biological function of circRNAs in the innate immune response in teleost fish.

TGF-β1 suppresses the T-cell response in teleost fish by initiating Smad3- and Foxp3-mediated transcriptional networks

Transforming growth factor-β1 (TGF-β1) can suppress the activation, proliferation, and function of many T-cell subsets, protecting organisms from inflammatory and autoimmune disease caused by an overexuberant immune response. However, whether and how TGF-β1 regulates T-cell immunity in early vertebrates remain unknown. Here, using a Nile tilapia (Oreochromis niloticus) model, we investigated suppression of the T-cell response by TGF-β1 in teleost species. Tilapia encodes an evolutionarily conserved TGF-β1, the expression of which in lymphocytes is significantly induced during the immune response following Edwardsiella piscicida infection. Once activated, tilapia T cells increase TGF-β1 production, which in turn suppresses proinflammatory cytokine expression and inhibits T-cell activation. Notably, we found administration of TGF-β1 cripples the proliferation of tilapia T cells, reduces the potential capacity of Th1/2 differentiation, and impairs the cytotoxic function, rendering the fish more vulnerable to bacterial infection. Mechanistically, TGF-β1 initiates the TGF-βR/Smad signaling pathway and triggers the phosphorylation and nuclear translocation of Smad2/3. Smad3 subsequently interacts with several transcriptional partners to repress transcription of cytokines IL-2 and IFN-γ but promote transcription of immune checkpoint regulator CTLA4 and transcription factor Foxp3. Furthermore, TGF-β1/Smad signaling further utilizes Foxp3 to achieve the cascade regulation of these T-cell genes. Taken together, our findings reveal a detailed mechanism by which TGF-β1 suppresses the T cell-based immunity in Nile tilapia and support the notion that TGF-β1 had already been employed to inhibit the T-cell response early in vertebrate evolution, thus providing novel insights into the evolution of the adaptive immune system.

Development of vaccine adjuvant that effectively induces cell-mediated immune response in teleost fish

Development of vaccine adjuvant that effectively induces cell-mediated immune response in teleost fish

Extensive involvement of CD40 and CD154 costimulators in multiple T cell-mediated responses in a perciform fish Larimichthys crocea

CD40 and CD154 are well-characterized costimulatory molecules involved in adaptive humoral immunity in humans and other mammals. These two costimulatory molecules were found to be originated from teleost fish during vertebrate evolution. However, the functionality of fish CD40 and CD154 remains to be explored. In this study, we identified the CD40 and CD154 homologs (LcCD40 and LcCD154) from large yellow croaker (Larimichthys crocea), a marine species of the perciform fish family. The LcCD40 and LcCD154 share conserved structural features to their mammalian counterparts, and are widely expressed in immune-relevant tissues and leukocytes at different transcriptional levels. Immunofluorescence staining and FCM analysis showed that LcCD40 and LcCD154 proteins are distributed on MHC-II+ APCs and CD4-2+ T cells, and are significantly upregulated in response to antigen stimulation. Co-IP assay exhibited strong association between LcCD40 and LcCD154 proteins. Blockade of LcCD154 with anti-LcCD154 antibody (Ab) or recombinant soluble LcCD40-Ig fusion protein remarkably decreased the MHC-II+ APC-initiated CD4+ T cell response upon Aeromonas hydrophila stimulation, and alloreactive T cell activation as examined by mixed lymphocyte reaction (MLR). These findings highlight the costimulatory role of LcCD40 and LcCD154 in T cell activities in Larimichthys crocea. Thus, the CD40 and CD154 costimulators may extensively participate in the regulation of multiple T cell-mediated immune responses in teleost fish. It is anticipated that this study would provide a cross-species understanding of the evolutionary history of CD40 and CD154 costimulatory signals from fish to mammals.

Exon-Intron Circular RNA circRNF217 Promotes Innate Immunity and Antibacterial Activity in Teleost Fish by Reducing miR-130-3p Function.

Circular RNA (circRNA) is produced by splicing head to tail and is widely distributed in multicellular organisms, and circRNA reportedly can participate in various cell biological processes. In this study, we discovered a novel exon-intron circRNA derived from probable E3 ubiquitin-protein ligase RNF217 (RNF217) gene, namely, circRNF217, which was related to the antibacterial responses in teleost fish. Results indicated that circRNF217 played essential roles in host antibacterial immunity and inhibited the Vibrio anguillarum invasion into cells. Our study also found a microRNA miR-130-3p, which could inhibit antibacterial immune response and promote V. anguillarum invasion into cells by targeting NOD1. Moreover, we also found that the antibacterial effect inhibited by miR-130-3p could be reversed with circRNF217. In mechanism, our data revealed that circRNF217 was a competing endogenous RNA of NOD1 by sponging miR-130-3p, leading to activation of the NF-κB pathway and then enhancing the innate antibacterial responses. In addition, we also found that circRNF217 can promote the antiviral response caused by Siniperca chuatsi rhabdovirus through targeting NOD1. Our study provides new insights for understanding the impact of circRNA on host-pathogen interactions and formulating fish disease prevention to resist the severely harmful V. anguillarum infection.

The long noncoding RNA MIR122HG is a precursor for miR-122-5p and negatively regulates the TAK1-induced innate immune response in teleost fish

Long noncoding RNAs (lncRNAs) are a diverse subset of RNA species of noncoding transcripts that are usually longer than 200 nt. However, the biological role and function of many lncRNAs have not been fully identified. It has been shown that one potential function of lncRNAs is to act as a precursor miRNA and promote the production of multiple miRNAs. However, the function of the miiuy croaker lncRNA MIR122HG has not been explored. In the present study, we show that this differentially expressed teleost fish lncRNA can act as the host gene of miR-122-5p, regulate its expression, and indirectly regulate the expression of potential inflammatory target protein transforming growth factor-β–activated kinase 1. We show that MIR122HG can negatively regulate the transforming growth factor-β–activated kinase 1–triggered NF-κB and interferon regulatory factor 3 signaling pathways and subsequently attenuate the innate immune response. In addition, MIR122HG can promote the replication of Siniperca chuatsi rhabdovirus and exacerbate the pathological effects caused by viral infection. We conclude that the study of lncRNA–miRNA–mRNA interaction through bioinformatics analysis or experimental-supported analysis can provide information for further elucidation of the functions of fish lncRNAs in innate immunity.

Characterization of the glucocorticoid receptor of large yellow croaker (Larimichthys crocea) and its expression in response to salinity and immune stressors

Glucocorticoids are steroidal hormones critical to stress responses in vertebrates. To gain further insight into the role of the glucocorticoid receptor (GR) in acute stress responses in teleost fish, the relevant cDNA of large yellow croaker (Larimichthys crocea; LcGR) was cloned using the rapid amplification of cDNA ends (RACE) technique. Multiple alignment of the amino acids (aa) of LcGR and the GR of other teleosts indicated LcGR contained four commonly conserved domains and lacked the 9-aa insert seen in GR1. Phylogenetic analysis of the amino acid sequence revealed that LcGR grouped most closely with the GR2 of other teleosts and can therefore be considered a GR2 subtype. In healthy L. crocea, Lcgr mRNA was found to be expressed at high levels in the gill, brain, and muscle tissue, expressed at intermediate levels in heart and stomach tissue, and expressed at low levels in the kidney, intestine, head kidney, liver, and spleen tissue. The response of L. crocea to acute low-salinity stress was tested, with a significant increase in plasma cortisol concentration after 3 h, peaking after 6 h, and gradually returning to base levels. Regarding changes of Lcgr expression in different body tissues under the stress, there was up-regulation of the Lcgr transcript in the brain, liver, and gill tissues, but not in muscle tissue. Responses to pathogen mimics were also tested. Injection with lipopolysaccharide resulted in Lcgr expression, with an increase-decrease-increase trend in the head kidney. In contrast, a down-regulation of Lcgr expression in the head kidney was observed throughout the experimental period upon injection of polyinosinic:polycytidylic acid, revealing different roles of Lcgr for different types of pathogens. The results offer novel insights about the effects of different stressors on GR gene expression in L. crocea, and can facilitate further investigations into stress responses in other mariculture fish species.

The circular RNA circBCL2L1 regulates innate immune responses via microRNA-mediated downregulation of TRAF6 in teleost fish

Growing numbers of studies have shown that circular RNAs (circRNAs) can function as regulatory factors to regulate the innate immune response, cell proliferation, cell migration, and other important processes in mammals. However, the function and regulatory mechanism of circRNAs in lower vertebrates are still unclear. Here, we discovered a novel circRNA derived from the gene encoding Bcl-2-like protein 1 (BCL2L1) gene, named circBCL2L1, which was related to the innate immune responses in teleost fish. Results indicated that circBCL2L1 played essential roles in host antiviral immunity and antibacterial immunity. Our study also identified a microRNA, miR-30c-3-3p, which could inhibit the innate immune response by targeting inflammatory mediator TRAF6. And TRAF6 is a key signal transduction factor in innate immune response mediated by TLRs. Moreover, we also found that the antiviral and antibacterial effects inhibited by miR-30c-3-3p could be reversed with the expression of circBCL2L1. Our data revealed that circBCL2L1 functioned as a competing endogenous RNA (ceRNA) of TRAF6 by competing for binding with miR-30c-3-3p, leading to activation of the NF-κB/IRF3 inflammatory pathway and then enhancing the innate immune responses. Our results suggest that circRNAs can play an important role in the innate immune response of teleost fish.

Identification and characterization of CD152 in Nile tilapia (Oreochromis niloticus) playing roles in host immune response against bacterial pathogens

The CD152, as named CTLA-4 (Cytotoxic T lymphocyte-associated antigen-4), is an important costimulatory molecule on the surface of T cells and plays an essential role in the cell-mediated immune response. In this work, Nile tilapia (Oreochromis niloticus) CD152 (OnCD152) was successfully cloned, and its characterization in response to bacterial pathogens and interaction with its ligand OnCD80/86 was also investigated. The open reading frame (ORF) of OnCD152 was 705 bp, encoding a polypeptide with 234 amino acids. The OnCD152 protein shared some homology with other known CD152 molecule from teleost species, containing conservative key sites, namely YPPPY, GNGT and four cysteine residues (Cys 45, Cys 97, Cys 123, Cys 151). The mRNA level of OnCD152 was extensively expressed in the collected healthy tissues. Following the stimulation of Poly (I:C), Streptococcus agalactiae and Aeromonas hydrophila, CD152 was significantly upregulated in the head kidney and spleen. The transcriptional upregulations of OnCD152 were also detected in Nile tilapia leukocytes after in vitro stimulations with Poly (I:C), S. agalactiae and A. hydrophila. In addition, the recombinant protein (r)OnCD152 was purified, and its binding ability to (r)CD80/86 strengthen with the increase of concentration. Taken together, this study demonstrated that OnCD152 was likely involved in host immune response against bacterial pathogens. Moreover, this research first cover about the capacity of OnCD152 binding with ligand CD80/86 in Nile tilapia (Oreochromis niloticus), which enrich the understanding for exploring the CD152 function in immune response in teleost fish in the future.

Functional characterization of TNF-α in pufferfish (Takifugu obscurus) in immune response and apoptosis against Aeromonas hydrophila.

Tumour necrosis factor-α (TNF-α) is a multifunctional cytokine involved in immune system homeostasis, antimicrobial defence, regulation of apoptosis, cell proliferation and differentiation. Although the pro-inflammatory property of TNF-α has been made new progress, detailed research on host defence against bacterial infection and inducing apoptosis remains to be revealed in early vertebrates. Here, we reported the TNF-α homologue (ToTNF-α) from pufferfish (Takifugu obscurus). The open reading frame (ORF) of ToTNF-α was 753bp, encoding a protein of 250 aa contained the TNF family signature and conserved cysteine residues. The mRNA expression of ToTNF-α had a wide range of tested tissues, with the highest expression in the skin. After Aeromonas hydrophila infection, the mRNA expression of ToTNF-α was significantly up-regulated both in vivo and in vitro experiments. After stimulation by recombinant protein of ToTNF-α ((r)ToTNF-α), the relative expressions of endogenous TNF-α, caspase 8, caspase 3, p53, and Bax inhibitor-1 in head kidney leucocytes were all notably up-regulated. These results showed that ToTNF-α might induce apoptosis depend on pro- and anti-apoptotic proteins at mRNA level. Moreover, flow cytometry analysis indicated that the (r)ToTNF-α can induce apoptosis of head kidney leucocytes. Taken together, these characteristics suggest that ToTNF-α can participate in immune response against A. hydrophila and induce apoptosis at mRNA and cellular level, which will help to understand the mechanism of apoptosis and immune response in teleost fish.

Circular RNA circPIKfyve acts as a sponge of miR-21-3p to enhance antiviral immunity through regulating MAVS in teleost fish.

Circular RNAs (circRNAs) are a class of widespread and diverse covalently closed circular endogenous RNAs that exert crucial functions in regulating gene expression in mammals. However, the function and regulation mechanism of circRNAs in lower vertebrates are still unknown. Here, we discovered a novel circRNA derived from PIKfyve, named circPIKfyve, that is related to the antiviral responses in teleost fish. The results showed that circPIKfyve plays essential roles in host antiviral immunity and inhibition of SCRV replication. Moreover, we also found that the antiviral effect inhibited by miR-21-3p could be reversed with the addition of circPIKfyve. In mechanism, our data revealed that circPIKfyve is a competitive endogenous RNA (ceRNA) of MAVS by sponging miR-21-3p, leading to activation of NF-κB/IRF3 pathway, which then enhance the innate antiviral responses. In addition, we firstly found that RNA binding protein QKI is involved in the formation and regulation of circPIKfyve. Our results provided a strong basis that circRNAs to play a regulatory role in antiviral immune responses in teleost fish.Importance: Here, we identified a novel circRNA, namely, circPIKfyve, that can act as a key regulator of the innate immune response in teleost fish. circPIKfyve acts as a molecular sponge by competitive adsorbing of miR-21-3p, thereby increasing the abundance of MAVS and activating the downstream NF-κB/IRF3 pathway to enhance the antiviral response. In addition, this study was the first to find that QKI protein is involved in regulating the formation of circPIKfyve in fish. The overall results of this study suggest that circPIKfyve plays an active regulatory role in the antiviral immune response of teleost fish.

Circular RNA circDtx1 regulates IRF3-mediated antiviral immune responses through suppression of miR-15a-5p-dependent TRIF downregulation in teleost fish.

Circular RNAs (circRNAs) represent a class of widespread and diverse covalently closed circular endogenous RNAs that exert crucial functions in regulating gene expression in mammals. However, the function and regulation mechanism of circRNAs in lower vertebrates are still unknown. Here, we discovered a novel circRNA derived from Deltex E3 ubiquitin ligase 1 (Dtx1) gene, namely, circDtx1, which was related to the antiviral responses in teleost fish. Results indicated that circDtx1 played essential roles in host antiviral immunity and inhibition of SCRV replication. Our study also found a microRNA miR-15a-5p, which could inhibit antiviral immune response and promote viral replication by targeting TRIF. Moreover, we also found that the antiviral effect inhibited by miR-15a-5p could be reversed with the circDtx1. In mechanism, our data revealed that circDtx1 was a competing endogenous RNA (ceRNA) of TRIF by sponging miR-15a-5p, leading to activation of the NF-κB/IRF3 pathway, and then enhancing the innate antiviral responses. Our results indicated that circRNAs played a regulatory role in immune responses in teleost fish.

Ocean acidification compromises energy management in Sparus aurata (Pisces: Teleostei)

The effects of ocean acidification mediated by an increase in water pCO2 levels on marine organisms are currently under debate. Elevated CO2 concentrations in the seawater induce several physiological responses in teleost fish, including acid-base imbalances and osmoregulatory changes. However, the consequences of CO2 levels enhancement on energy metabolism are mostly unknown. Here we show that 5 weeks of exposure to hypercapnia (950 and 1800 μatm CO2) altered intermediary metabolism of gilthead seabream (Sparus aurata) compared to fish acclimated to current ocean values (440 μatm CO2). We found that seabream compromises its physiological acid-base balance with increasing water CO2 levels and the subsequent acidification. Intestinal regions (anterior, mid, and rectum) engaged in maintaining this balance are thus altered, as seen for Na+/K+-ATPase and the vacuolar-type H+-ATPase activities. Moreover, liver and muscle counteracted these effects by increasing catabolic routes e.g., glycogenolysis, glycolysis, amino acid turnover, and lipid catabolism, and plasma energy metabolites were altered. Our results demonstrate how a relatively short period of 5 weeks of water hypercapnia is likely to disrupt the acid-base balance, osmoregulatory capacity and intermediary metabolism in S. aurata. However, long-term studies are necessary to fully understand the consequences of ocean acidification on growth and other energy-demanding activities, such as reproduction.

The Sp1-Responsive microRNA-15b Negatively Regulates Rhabdovirus-Triggered Innate Immune Responses in Lower Vertebrates by Targeting TBK1.

As is known to all, the production of type I interferon (IFN) plays pivotal roles in host innate antiviral immunity, and its moderate production play a positive role in promoting the activation of host innate antiviral immune response. However, the virus will establish a persistent infection model by interfering with the production of IFN, thereby evading the organism inherent antiviral immune response. Therefore, it is of great necessity to research the underlying regulatory mechanisms of type I IFN appropriate production under viral invasion. In this study, we report that a Sp1–responsive miR-15b plays a negative role in siniperca chuatsi rhabdovirus (SCRV)-triggered antiviral response in teleost fish. We found that SCRV could dramatically upregulate miiuy croaker miR-15b expression. Enhanced miR-15b could negatively regulate SCRV-triggered antiviral genes and inflammatory cytokines production by targeting TANK-binding kinase 1 (TBK1), thereby accelerating viral replication. Importantly, we found that miR-15b feedback regulates antiviral innate immune response through NF-κB and IRF3 signaling pathways. These findings highlight that miR-15b plays a crucial role in regulating virus–host interactions, which outlines a new regulation mechanism of fish’s innate immune responses.

Full-length transcriptome sequencing combined with RNA-seq analysis revealed the immune response of fat greenling (Hexagrammos otakii) to Vibrio harveyi in early infection

Fat greenling (Hexagrammos otakii) is an important commercial marine fish species cultured in northeast Asia, but its available gene sequences are limited. Vibrio harveyi is a causative agent of vibriosis in fat greenling and also causes severe losses to the aquaculture industry in China. In order to obtain more high-quality transcript information and investigate the early immune response of fat greenling against V. harveyi, the fish were artificially infected with V. harveyi, and five sampling points were set within 48 h. Iso-Seq combined with RNA-Seq were applied in the comprehensive transcriptome analysis of V. harveyi-infected fat greenling. Total 42,225 consensus isoforms were successfully extracted from the result of Iso-Seq, and more than 19,000 ORFs were predicted. In addition, total three modules were identified by WGCNA which significantly positive correlated to the infection time, and the KEGG analysis showed that the immune-related genes in these modules mainly enriched in TLR signaling pathway, NF-κB signaling pathway and Endocytosis. The activation of inflammation and endocytosis was the most significant characteristics of fat greenling immune response during the early infection. Based on the WGCNA, a series of high–degree nodes in the networks were identified as hub genes. The protein structures of cold-inducible RNA-binding protein (CIRBP), poly [ADP-ribose] polymerase 1 (PARP1) and protein arginine N-methyl transferase 1 (PRMT1) were subsequently found to be highly conserved in vertebrate, and the gene expression pattern of CIRBP, PARP1, PRMT1 and a part of TLR/NF-κB pathway-related genes indicated that these proteins might have similar biological functions in regulation of inflammatory response in teleost fish. The results of this study provided the first systematical full-length transcriptome profile of fat greenling and characterized its immune responses in early infection of V. harvey, which will serve as the foundation for further exploring the molecular mechanism of immune defense against bacterial infection in fat greenling.

Long noncoding RNA AANCR modulates innate antiviral responses by blocking miR-210-dependent MITA downregulation in teleost fish, Miichthys miiuy.

Viral infection induces the initiation of antiviral effectors and cytokines which are critical mediators of innate antiviral responses. The critical molecular determinants are responsible for triggering an appropriate immune response. Long noncoding RNAs (lncRNAs) have emerged as new gene modulators involved in various biological processes, while how lncRNAs operate in lower vertebrates are still unknown. Here, we discover a long noncoding RNA, termed antiviral-associated long noncoding RNA (AANCR), as a novel regulator for innate antiviral responses in teleost fish. The results indicate that fish MITA plays an essential role in host antiviral responses and inhibition of Siniperca chuatsi rhabdovirus (SCRV) production. miR-210 reduces MITA expression and suppress MITA-mediated antiviral responses, which may help viruses evade host antiviral responses. Further, AANCR functions as a competing endogenous RNA (ceRNA) for miR-210 to control protein abundance of MITA, thereby inhibiting SCRV replication and promoting antiviral responses. Our data not only shed new light on understanding the function role of lncRNA in biological processes in teleost fish, but confirmed the hypothesis that ceRNA networks exist widely in vertebrates.