Cloning and functional characterization of IRAK1 from rainbow trout (Oncorhynchus mykiss)

As a key molecule in innate immune signalling pathway, interleukin (IL)-1 receptor-associated kinase 1 (IRAK1) mediates downstream signalling cascades in immune response. In the present study, an IRAK1 orthologue was characterized from rainbow trout (Oncorhynchus mykiss), with a 2115 bp open reading frame (ORF), encoding a protein of 704 amino acids (aa). Multiple alignments showed that IRAK1 contains highly conserved features among different species, with a conservative N-terminal death domain (DD) and a C-terminal conserved serine/threonine protein kinase (STKc) domain. Expression analysis indicated that IRAK1 was widely expressed in examined organs/tissues, with the highest level observed in muscle and lowest in stomach. In RTG-2 cell line, the induced expression of IRAK1 was observed following the stimulation by the fish bacterial pathogen Flayobacterium columnare. Luciferase activity assays revealed that IRAK1 induced significantly the activity of NF-kappa B in Human embryonic kidney 293T (HEK293T) cell line; but after co-transfected with rainbow trout IL-1 receptorassociated kinase 4 (IRAK4), the induction was significantly down-regulated when compared with the expression of IRAK1 alone. Co-immunoprecipitation (Co-IP) assays indicated that IRAK1 was associated with rainbow trout myeloid differentiation factor 88 (MyD88), IRAK4 and TNF receptor associated factor 6 (TRAF6) in transfected HEK293T cells, and may form a complex with MyD88, IRAK4 and TRAF6 during the signalling pathway

Molecular characterization of a cyprinid fish (Ancherythroculter nigrocauda) TBK1 and its kinase activity in IFN regulation

TANK-binding kinase 1 (TBK1) plays a vital role in activating interferon (IFN) production and positively regulating antiviral response in mammals. Research on more species of fish is necessary to clarify whether the function of fish TBK1 is conserved compared to that in mammals. Here, a cyprinid fish (Ancherythroculter nigrocauda) TBK1 (AnTBK1) was functionally identified and characterized. The full-length open reading frame (ORF) of AnTBK1 consists of 2184 nucleotides encoding 727 amino acids and contains a conserved Serine/Threonine protein kinase catalytic domain (S_TKc) in the N-terminal, similar to TBK1 in other species. The transcripts of AnTBK1 were found in all the tissues evaluated and the cellular distribution indicated that AnTBK1 was localized in the cytoplasm. In terms of functional identification, AnTBK1 induced a variety of IFN promoter activities as well as the expression of downstream IFN-stimulated genes (ISGs). In addition, AnTBK1 interacted with and significantly phosphorylated IFN regulatory factor 3 (IRF3), exhibiting the canonical kinase activity of TBK1. Finally, AnTBK1 presented strong antiviral activity against spring viremia of carp virus (SVCV) infection. Taken together, our research on the features and functions of AnTBK1 demonstrated that AnTBK1 plays a central role in IFN induction against SVCV infection

Species-specific responses of submergedmacrophytes to the presence of a small omnivorous bitterling Acheilognathus macropterus

Recovery of submerged macrophytes has been considered a key factor in the restoration of shallow eutrophic lakes. However, in some subtropical restored lakes, small omnivorous fish dominate the fish assemblages and feed in part on submerged macrophytes. Knowledge of the effects of small omnivores on the growth of submerged macrophytes is scarce and their responses are potentially species-specific, i.e. the growth of some species may be hampered by fish grazing while growth of others may be promoted by the nutrients becoming available by fish excretion. We conducted mesocosm experiments to examine the effects of the small omnivorous bitterling Acheilognathus macropterus, a common species in restored subtropical lakes in China, on nutrient concentrations and the growth of four species of submerged macrophytes (Hydrilla verticillata, Vallisneria denseserrulata, Ceratophyllum demersum and Myriophyllum spicatum). We found that the bitterling significantly increased nutrient concentrations via excretion and thereby enhanced the net growth of the less grazed nuisance macrophyte M. spicatum. In contrast, the net growth of C. demersum was reduced by the bitterling, most likely due to grazing as indicated by gut content analyses. Dominance by bitterling may, therefore, pose a threat to the long-term success of lake restoration by provoking a shift in the submerged macrophyte community towards nuisance species through selective grazing. Nutrient excretion may potentially also stimulate the growth of phytoplankton and periphyton, hampering the growth of submerged macrophyte. (C) 2020 Elsevier B.V. All rights reserved

The expanding and function of NLRC3 or NLRC3-like in teleost fish: Recent advances and novel insights

The nucleotide-binding domain and leucine-rich repeat-containing family (NLR) proteins are innate immune sensors which recognize highly conserved pathogen-associated molecular patterns (PAMPs). Mammals have small numbers of NLR proteins, whereas in some species such as in invertebrates and jawless vertebrates, NLRs have expanded into very large families. Nearly 400 NLR proteins are identified in the zebrafish genome. Members of the NLR family can be divided into two functional sub-groups based on their ability to either positively or negatively regulate host immune response or inflammatory signaling cascades. Mammalian NLRC3 has been identified as an inhibitory NLR, and serves as a negative regulator in the NE-KB-mediated inflammatory response, STING-mediated DNA sensing and PI3K-mTOR pathways. Different from mammalian NLRC3, the analysis from genomes or transcriptomes revealed that the expansions of NLRC3 existed in different species of fish. Furthermore, piscine NLRC3-like genes were confirmed to have a negative or positive regulatory function in response to different kinds of pathogen infections and in the production of pminflammatory cytokines. In this review, we summarize recent advances in our understanding of the expanding and function of NLRC3 or NLRC3-like genes in teleost fish, and give our view of important directions for future studies. The knowledge of piscine NLRC3 or expansive NLRC3-like genes-mediated biological functions in homeostasis and diseases will shed new light on the prevention and control of inflammatory and/or infectious diseases

The expanding and function of NLRC3 or NLRC3-like in teleost fish: Recent advances and novel insights

The nucleotide-binding domain and leucine-rich repeat-containing family (NLR) proteins are innate immune sensors which recognize highly conserved pathogen-associated molecular patterns (PAMPs). Mammals have small numbers of NLR proteins, whereas in some species such as in invertebrates and jawless vertebrates, NLRs have expanded into very large families. Nearly 400 NLR proteins are identified in the zebrafish genome. Members of the NLR family can be divided into two functional sub-groups based on their ability to either positively or negatively regulate host immune response or inflammatory signaling cascades. Mammalian NLRC3 has been identified as an inhibitory NLR, and serves as a negative regulator in the NE-KB-mediated inflammatory response, STING-mediated DNA sensing and PI3K-mTOR pathways. Different from mammalian NLRC3, the analysis from genomes or transcriptomes revealed that the expansions of NLRC3 existed in different species of fish. Furthermore, piscine NLRC3-like genes were confirmed to have a negative or positive regulatory function in response to different kinds of pathogen infections and in the production of pminflammatory cytokines. In this review, we summarize recent advances in our understanding of the expanding and function of NLRC3 or NLRC3-like genes in teleost fish, and give our view of important directions for future studies. The knowledge of piscine NLRC3 or expansive NLRC3-like genes-mediated biological functions in homeostasis and diseases will shed new light on the prevention and control of inflammatory and/or infectious diseases

Production, functional stability, and effect of rhamnolipid biosurfactant from Klebsiella sp. on phenanthrene degradation in various medium systems

The present study investigated the stability and efficacy of a biosurfactant produced by Klebsiella sp. KOD36 under extreme conditions and its potential for enhancing the solubilization and degradation of phenanthrene in various environmental matrices. Klebsiella sp. KOD36 produced a mono-rhamnolipids biosurfactant with a low critical micelle concentration (CMC) value. The biosurfactant was stable under extreme conditions (60 degrees C, pH 10 and 10% salinity) and could lower surface tension by 30% and maintained an emulsification index of > 40%. The emulsion index was also higher (17-43%) in the presence of petroleum hydrocarbons compared to synthetic surfactant Triton X-100. Investigation on phenanthrene degradation in three different environmental matrices (aqueous, soil-slurry and soil) confirmed that the biosurfactant enhanced the solubilization and biodegradation of phenanthrene in all matrices. The high functional stability and performance of the biosurfactant under extreme conditions on phenanthrene degradation show the great potential of the biosurfactant for remediation applications under harsh environmental conditions

Production, functional stability, and effect of rhamnolipid biosurfactant from Klebsiella sp. on phenanthrene degradation in various medium systems

The present study investigated the stability and efficacy of a biosurfactant produced by Klebsiella sp. KOD36 under extreme conditions and its potential for enhancing the solubilization and degradation of phenanthrene in various environmental matrices. Klebsiella sp. KOD36 produced a mono-rhamnolipids biosurfactant with a low critical micelle concentration (CMC) value. The biosurfactant was stable under extreme conditions (60 degrees C, pH 10 and 10% salinity) and could lower surface tension by 30% and maintained an emulsification index of > 40%. The emulsion index was also higher (17-43%) in the presence of petroleum hydrocarbons compared to synthetic surfactant Triton X-100. Investigation on phenanthrene degradation in three different environmental matrices (aqueous, soil-slurry and soil) confirmed that the biosurfactant enhanced the solubilization and biodegradation of phenanthrene in all matrices. The high functional stability and performance of the biosurfactant under extreme conditions on phenanthrene degradation show the great potential of the biosurfactant for remediation applications under harsh environmental conditions

An identified PfHMGB1 promotes microcystin-LR-induced liver injury of yellow catfish (Pelteobagrus fulvidraco)

Microcystin-LR (MC-LR) is a potent hepatotoxin that can cause liver inflammation and injury. However, the mode of action of related inflammatory factors is not fully understood. PfHMGB1 is an inflammatory factor induced at the mRNA level in the liver of juvenile yellow catfish (Pelteobagrus fulvidraco) that were intraperi-toneally injected with 50 mu g/kg MC-LR. The PfHMGB1 mRNA level was highest in the liver and muscle among 11 tissues examined. The full-length cDNA sequence of PfHMGB1 was cloned and overexpressed in E. coli, and the purified protein rPfHMGB1 demonstrated DNA binding affinity. Endotoxin-free rPfHMGB1 (6-150 mu g/mL) also showed dose-dependent hepatotoxicity and induced inflammatory gene expression of primary hepatocytes. PfHMGB1 antibody (anti-PfHMGB1) in vitro reduced MC-LR (30 and 50 mu mol/L)-induced hepatotoxicity, suggesting PfHMGB1 is important in the toxic effects of MC-LR. In vivo study showed that MC-LR upregulated PfHMGB1 protein in the liver. The anti-PfHMGB1 blocked its counterpart and reduced ALT/AST activities after MC-LR exposure. Anti-PfHMGB1 partly neutralized MC-LR-induced hepatocyte disorganization, nucleus shrinkage, mitochondria, and rough endoplasmic reticula destruction. These findings suggest that PfHMGB1 promotes MC-LR-induced liver damage in the yellow catfish. HMGB1 may help protect catfish against widespread microcystin pollution

Cloning and functional characterization of IRAK1 from rainbow trout (Oncorhynchus mykiss)

As a key molecule in innate immune signalling pathway, interleukin (IL)-1 receptor-associated kinase 1 (IRAK1) mediates downstream signalling cascades in immune response. In the present study, an IRAK1 orthologue was characterized from rainbow trout (Oncorhynchus mykiss), with a 2115 bp open reading frame (ORF), encoding a protein of 704 amino acids (aa). Multiple alignments showed that IRAK1 contains highly conserved features among different species, with a conservative N-terminal death domain (DD) and a C-terminal conserved serine/threonine protein kinase (STKc) domain. Expression analysis indicated that IRAK1 was widely expressed in examined organs/tissues, with the highest level observed in muscle and lowest in stomach. In RTG-2 cell line, the induced expression of IRAK1 was observed following the stimulation by the fish bacterial pathogen Flayobacterium columnare. Luciferase activity assays revealed that IRAK1 induced significantly the activity of NF-kappa B in Human embryonic kidney 293T (HEK293T) cell line; but after co-transfected with rainbow trout IL-1 receptorassociated kinase 4 (IRAK4), the induction was significantly down-regulated when compared with the expression of IRAK1 alone. Co-immunoprecipitation (Co-IP) assays indicated that IRAK1 was associated with rainbow trout myeloid differentiation factor 88 (MyD88), IRAK4 and TNF receptor associated factor 6 (TRAF6) in transfected HEK293T cells, and may form a complex with MyD88, IRAK4 and TRAF6 during the signalling pathway

IRF11 regulates positively type I IFN transcription and antiviral response in mandarin fish, Siniperca chuatsi

In vertebrates, a total of eleven interferon (IFN) regulatory factors (IRFs), IRF1 to IRF11 are reported, with the conserved presence of IRF1 to IRF9 in all classes of vertebrates. However, IRF10 has been reported only in fish and birds, and IRF11 seems to be a fish specific IRF member. In this study, IRF11 in mandarin fish Siniperca chuatsi was found upregulated following virus infection, and IRF11 was localized constitutively in nucleus as revealed through immunofluorescence test. The overexpression and/or luciferase reporter assays showed that IRF11 can induce transcriptionally the ISRE activity, and the expression of type I IFNs, IFNc and IFNh, as well as the IFN-stimulated gene, Mx, thus inhibiting the Siniperca chuatsi rhabdovirus (SCRV) replication as indicated in the reduced expression of virus protein genes. It is thus suggested that IRF11 in mandarin fish and probably in other teleost fish can exert its antiviral effect through the upregulation of type I IFNs and ISGs