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Methyltransferase (METTL3), the most important N6-methyladenosine (m6A) writer, plays a vital role in regulating immune-related signaling pathways. However, the underlying mechanism of METTL3 action remains largely unknown, especially in lower vertebrates. The results of this study show that METTL3 inhibits innate immune response and promotes the infection of miiuy croaker, Miichthys miiuy, by Siniperca chuatsi rhabdovirus and Vibrio anguillarum. Significantly, the function of METTL3 in inhibiting immunity depends on its methylase activity. Mechanistically, METTL3 increases the methylation level of trif and myd88 mRNA, rendering them sensitive to degradation by the YTHDF2/3 reader proteins. By contrast, we found that the YTHDF1 reader protein promotes the translation of myd88 mRNA. In summary, these results indicate that METTL3-mediated m6A modification of trif and myd88 mRNAs suppresses innate immunity by inhibiting the TLR pathway, unveiling a molecular mechanism by which RNA methylation controls innate immunity to pathogens in the teleost fish.

Interferon-mediated innate immune response is the first line of defense against foreign pathogen infection. Overexpression of MITA can activate the expression of interferon and promote the innate immune response of the body to the virus. These innate immune responses are tightly controlled to prevent the host from over-immunizing itself. In this study, we reported that structurally highly conserved PCNA negatively regulates MITA. PCNA overexpression can promote MITA degradation and block the expression of interferon, while the autophagy inhibitor 3-MA significantly inhibits MITA degradation, indicating that PCNA can degrade MITA through the autophagy pathway. PCNA inhibits interferon production by targeting MITA and avoids excessive immune response. In summary, our results indicate that PCNA is involved in the immune response by degrading MITA through the autophagy pathway, which will provide new ideas for further studies on the regulatory mechanism of immune signaling pathways in lower vertebrates.

Accumulating evidence has demonstrated that microRNAs (miRNAs) regulate various physiological and pathological processes at the transcriptional level, thus called novel regulators in immune response. In this study, we used bioinformatics and functional experiments to determine the role of miR-103 and miR-190 in the regulation of IL-1R1 gene involved in the immune and inflammatory responses in miiuy croakers. First, we predicted the target genes of miR-103 and miR-190 through bioinformatics and found that IL-1R1 is a direct target gene of miR-103 and miR-190. This was further confirmed by the dual-luciferase reporter assay that the over-expression of miR-103, miR-190 mimics and the pre-miR-103, pre-miR-190 plasmids inhibit the luciferase levels of the wild-type of IL-1R1 3'UTR. miR-103 and miR-190 inhibitors increase the luciferase levels of IL-1R1-3'UTR. Additionally, we found that miR-103 and miR-190 could negatively regulate the mRNA expression of IL-1R1. Importantly, we demonstrated that miR-103 and miR-190 significantly inhibit the NF-κB signaling pathway by targeting IL-1R1 upon LPS stimulation. Collectively, these results provide strong evidence for an important regulatory mechanism of miR-103 and miR-190 targeting the IL-1R1 gene, thereby preventing excessive inflammatory immune responses from causing autoimmunity.

The inhibitory protein IκBα plays a key role in the inflammatory process and immune response by regulating the activity of the transcription factor NF-κB. microRNA (miR) is a small non-coding RNA that can regulate many biochemical processes, such as cell growth, proliferation, and immune response. In this study, it was first predicted that IκBα is the target of miR-122 through bioinformatics, and it was confirmed by dual fluorescence experiments. Then we found that miR-122 can inhibit the expression of IκBα at the mRNA and protein levels, thereby promoting the p65-activated NF-κB pathway. It is speculated that miR-122 plays an important role in the innate immunity of teleost fish. This study will help to further understand miRNAs regulatory mechanism in teleost fish.

Sciaenops ocellatus, also known as red drum, is a valuable commercial fish that is widely cultured in China. In recent years, while the genetic research and molecular breeding on S. ocellatus has been widely performed, whole-genome resources are lacking. In our study, we assembled the S. ocellatus genome by high-quality genome sequencing, followed by complete annotation. The size of final genome was 686.62 Mb and N50 length of contig and scaffold is 99.71 Kb and 25.62 Mb, respectively. Using the Hi-C technology, we obtained 24 pseudochromosomes containing 87.82% of the total scaffolds. We annotated 20,053 protein-coding genes in S. ocellatus genome, of which 99.93% were homologous to proteins in different database. Totally, we detected 133.54 Mb repeat sequences, which accounts for 19.45% of the assembly. According to the phylogenetic analysis, S. ocellatus is most closed to L. croea with a divergence time about 30 million years ago. We assembled a high-quality genome that will be a valuable resource to further study the biology of S. ocellatus and will inform how to breed the red drum to enhance its economic traits.

This paper analyzes tourists’ perceived preferences for cultural and creative product elements using human-computer interaction technology and constructs the innovation and transformation path of cultural and creative products from four dimensions: concept, elements, content, and structure. The Great Wall tourism cultural and creative products are used as an example. The findings demonstrate that: (1) From a behavioral data viewpoint, cultural and creative items’ overall inventiveness, formal design, manufacturing method, area, cultural collection value, and function have varying degrees of influence on visitors’ perceived preferences; (2) The richness and attraction of character expression, action, and form components from the hotspot map and matrix map can boost the visual engagement impact of visitors. Scenic area architecture may enhance visitors’ immersion experiences of local culture since it serves as the design prototype for cultural and creative businesses. (3) The number of fixation points, total fixation time, and saccade frequency of cultural and creative products with various design elements differ significantly when viewed from the perspective of the eye movement index, and these differences are further presented as individualized tourist behavior characteristics. (4) From a design standpoint, it is essential that the circumstances of the product satisfy the needs of visitors in order to produce high-quality cultural and creative products. Innovative ideas should be used to steer the innovation and transformation of cultural and creative products, enhancing the universal design of products with element innovation, enhancing the cultural legacies of products with content innovation, and lengthening the market cycle of products with structural innovation. The use of modern technology broadens the research methodologies for the tourism field and creates new research environments for tourism experimentation.

NF-κB is a typical transcription factor that regulates expression of various genes involved in inflammatory and immune responses. Therefore, it is essential that NF-κB signaling tightly regulated to maintain immune balance. Compared with those of mammals, the regulatory mechanisms of NF-κB signaling is rarely reported in teleost fish. IκBα is a prominent negative feedback regulator in the NF-κB signaling system. In this study, we determined that IRF11 enhances the inhibitory effect of IκBα on NF-κB activation in teleost fish. Overexpression of IRF11 can inhibit IκBα degradation, whereas its knockdown has the opposite effect of IκBα. Our study further indicates that IκBα was regulated via ubiquitin-proteasome degradation pathway, IRF11 inhibits IκBα in ubiquitin-proteasome degradation. This study provides a novel evidence on the regulation of innate immune signaling pathways in teleost fish and thus provides new insights into the regulatory mechanisms in mammals.

The innate immune response is the first line of defense against pathogen infection. Eradication of pathogen infection requires appropriate immune and inflammatory responses, but excessive inflammation may cause inflammatory and autoimmune diseases. MicroRNAs (miRNAs) are a group of small noncoding RNAs, and accumulating evidence has shown that in mammals, they can act as negative regulators that participate in the regulation of inflammation and immune responses. However, the miRNA-mediated immune regulation networks in the inflammatory responses of lower vertebrates are largely unknown. In this study, we report an miRNA, miR-132, identified from miiuy croaker, that acts as a negative regulator in the host's bacterium-induced inflammatory response. We found that miR-132 expression was dramatically increased upon infection by the Gram-negative bacterium Vibrio harveyi and lipopolysaccharide (LPS). Inducible miR-132 inhibits the production of inflammatory cytokines by targeting tumor necrosis factor receptor-associated factor 6 (TRAF6), transforming growth factor-activated protein kinase 1 (TAK1), and TAK1 binding protein 1 (TAB1), thus avoiding an excessive inflammatory response. Furthermore, we demonstrate that miR-132 modulates the inflammatory response through a TRAF6-, TAK1-, and TAB1-mediated NF-κB signaling pathway. These results collectively reveal that miR-132 plays a negative regulatory role in the host antibacterial immune response, which will help to gain insight into the intricate network of host resistance to pathogen infection in lower vertebrates.

Transforming growth factor-beta activated kinase 1 (TAK1) is an adaptor molecular in TLR-mediated NF-κB signaling pathway and plays indispensable roles in innate immunity. As the most typical innate immune pathway, the strict regulation of NF-κB signaling pathway is particularly important. Rho-GDP-dissociation inhibitor-γ (Rho-GDIγ) is a member of the Rho protein family that regulates many important physiological processes. In this study, we demonstrated the mechanism of suppressing TAK1 expression in the teleost and found that Rho-GDIγ negatively regulated the NF-κB signaling pathway mediated by TAK1. We determined that TAK1 could directly interact with Rho-GDIγ. It is interesting that Rho-GDIγ promotes TAK1 degradation through the ubiquitin proteasome pathway. This study brings a new experimental basis to the teleost fish innate immune signaling pathway. Moreover, this discovery may provide new insights into innate immune regulation mechanism in mammals.

The diversity of endophytic bacteria in the progeny is related to the parental lines. In this study, the traditional separation method was used to study the dominant endophytic bacteria of the shared paternal line and its pollen, different maternal lines and their F1 progeny. And the results showed that the dominant endophytic bacteria in maize seeds and the pollen were Bacillus and Pantoea. The Bacillus diversity of the progeny JMC121 and JN728 were the same as both the paternal line and the maternal line, including Bacillus subtilis, Bacillus velezensis, Bacillus mojavensis, and Bacillus licheniformis. The Bacillus subtilis and Bacillus velezensis in JN828 were the same as both the paternal line and the maternal line, while Bacillus licheniformis was only the same as the paternal line. Through the RAPD molecular typing, there was the same strain of Bacillus mojavensis existed in the paternal line J2416, the pollen and the progeny JN728; this meant that the paternal line passed its dominant endophytic bacteria to the progeny through pollen in vertical transmission. This study showed that the dominant endophytic bacteria in maize seeds and the pollen were Bacillus, and the diversity of F1 progeny was related to both the paternal line and the maternal line.

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.

The suppressor of cytokine signaling (SOCS) was first described as inhibitors of cytokine signaling. The SOCS1, as a number of SOCS family, is an important negative regulator in the IFN signaling pathways in mammals. While data on functional characterization of SOCS1 in lower vertebrates are limited. In this study, we identified and characterized the full length SOCS1b gene of miiuy croaker (Miichthys miiuy). The sequence alignment analysis results showed that miiuy croaker SOCS1b (mmSOCS1b) have only a conserved SH2 domain that is similar to other vertebrates. To further study the functions of mmSOCS1b, we identified and determined its potential ability to perceive poly (I:C) stimulation. Stimulation experiments with poly (I:C) showed the significantly upregulated expression of mmSOCS1b in crucial immune-related tissues of spleen and kidney, indicating that mmSOCS1b might participate in the immune responses. Furthermore, the immunofluorescence assay indicated that mmSOCS1b present in the cytoplasmic of HeLa cells. In addition, mmSOCS1b could inhibit IFNα or IFNγ-induced ISRE reporter gene. In a word, we systematically and comprehensively analyzed the characterizations and functions of mmSOCS1b, which not only enriches the current knowledge of SOCS in IFN signaling regulation but also offer the basis for future research of fish SOCS family.

Upon recognition of the pathogen components by PRR (pattern recognition receptors), then the cells could be activated to produce inflammatory cytokines and type I interferons. The inflammation is tightly modulated by the host to prevent inappropriate inflammatory responses. MicroRNAs (miRNAs) are non-coding and small RNAs that can inhibit gene expression and participate in various biological functions, including maintaining a balanced immune response in the host. To maintain the balance of the immune response, these pathways are closely regulated by the host to prevent inappropriate reactions of the cells. However, in low vertebrates, the miRNA-mediated inflammatory response regulatory networks remain largely unknown. Here, we report that two miRNAs, miR-20-1 and miR-101a are identified as negative regulators in teleost inflammatory responses. Initially, we find that both miR-20-1 and miR-101a dramatically increased after lipopolysaccharide (LPS) stimulation and Vibrio harveyi infection. Upregulated miR-20-1 and miR-101a inhibit LPS-induced inflammatory cytokines production by targeting TNF receptor-associated factor 6 (TRAF6), thus avoiding excessive inflammation. Moreover, miR-20-1 and miR-101a regulate the inflammatory responses through the TRAF6-mediated nuclear factor kappa (NF-κB) signaling pathways. Collectively, these data indicate that miR-20-1 and miR-101a act as negative regulators through regulating the TRAF6-mediated NF-κB signaling pathway, and participate in the host antibacterial immune responses, which will provide new insight into the intricate networks of the host-pathogen interaction in the lower vertebrates.

The diversity of endophytic bacteria in the progeny is related to the parental lines. In this study, the traditional separation method was used to study the dominant endophytic bacteria of the shared paternal line and its pollen, different maternal lines and their F1 progeny. And the results showed that the dominant endophytic bacteria in maize seeds and the pollen were Bacillus and Pantoea. The Bacillus diversity of the progeny JMC121 and JN728 were the same as both the paternal line and the maternal line, including Bacillus subtilis, Bacillus velezensis, Bacillus mojavensis, and Bacillus licheniformis. The Bacillus subtilis and Bacillus velezensis in JN828 were the same as both the paternal line and the maternal line, while Bacillus licheniformis was only the same as the paternal line. Through the RAPD molecular typing, there was the same strain of Bacillus mojavensis existed in the paternal line J2416, the pollen and the progeny JN728, this meant that the paternal line passed its dominant endophytic bacteria to the progeny might through pollen in vertical transmission. This study showed that the dominant endophytic bacteria in maize seeds and the pollen were Bacillus, and the diversity of F1 progeny was related to both the paternal line and the maternal line.

Long noncoding RNAs (lncRNAs) function as microregulatory factors that influence gene expression after a variety of pathogenic infections, and they have been extensively studied in the past few years. Although less attention has been paid to lncRNAs in lower vertebrates than in mammals, current studies reveals that lncRNAs play a vital role in fish stimulated by pathogens. Here, we discovered a new lncRNA, termed MIR2187HG, which can function as a precursor of a small RNA, miR-2187-3p, with regulatory functions in the miiuy croaker (Miichthys miiuy). Upon Siniperca chuatsi rhabdovirus (SCRV) virus infection, the expression levels of MIR2187HG were remarkably enhanced. Elevated MIR2187HG expression can act as a pivotally negative regulator that participates in the innate immune response of teleost fish to inhibit the intracellular TANK-binding kinase 1 (TBK1)-mediated antiviral signaling pathways, which can effectively avoid excessive immunity. In addition, we found that SCRV could also utilize MIR2187HG to enhance its own numbers. Our results not only provide evidence regarding the involvement of the lncRNAs in response to viruses in fish but also broaden our understanding of the function of lncRNAs as precursor microRNAs (miRNAs) in teleost fish for the first time. IMPORTANCE SCRV infection upregulates MIR2187HG levels, which in turn suppresses SCRV-triggered type I interferon production, thus promoting viral replication in miiuy croaker. Notably, MIR2187HG regulates the release of miR-2187-3p, and TBK1 is a target of miR-2187-3p. MIR2187HG could acquire from miR-2187-3p the function of inhibiting TBK1 expression and subsequently modulate TBK1-mediated NF-κB and IRF3 signaling. The collective results suggest that the novel regulation mechanism of TBK1-mediated antiviral response during RNA viral infection was regulated by MIR2187HG. Therefore, a new regulation mechanism for lncRNAs to regulate antiviral immune responses in fish is proposed.

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.

MicroRNAs (miRNA) are non-coding RNAs that regulate many biochemical processes, such as cell growth, proliferation and immune responses. In this study, we investigated miR-144 as a regulator of IκBα that promotes the activation of NF-κB signaling pathway. And IκBα interact with p65 blocks nuclear translocation of NF-κB and anchors NF-κB in cytoplasmic quiescent cells in an inactive form. The seed region of miR-144 can regulate gene expression by binding to the 3' UTR of IκBα and repress IκBα expression at the post-transcriptional level. More importantly, miR-144 can promote the activation of p65 by inhibiting IκBα, thus affecting the NF-κB signaling pathway. Thus, preventing excessive inflammatory responses from causing autoimmune diseases will help to further understand the immunoregulatory mechanisms of miRNAs in fish after invasion by pathogens.

As a non-coding RNA with regulatory functions, microRNAs(miRNAs) can regulate gene expression and participate in a variety of physiological and pathological processes. In recent years, although there have been many studies on miRNA, the regulation mechanisms of miRNA in teleost fish have not been fully elucidated. In this study, it was first predicted that MITA is the target of miR-27c through bioinformatics, and it was confirmed by dual fluorescence experiments. Then we found that miR-27c can inhibit the expression of MITA at the mRNA and protein levels, thereby promoting the NF-κB or IRF3 pathway. It is speculated that miR-27c plays an important role in the innate immunity of teleost fish. This study will help to further understand miRNAs regulatory mechanism in teleost fish.

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.

MDA5 is a member of retinoic acid-inducible gene I (RIG-I)-like receptors (RLR receptors), which may play a crucial role in the immune regulation process. Recently, microRNAs (miRNAs) have been shown to act as an important regulator in the RLRs signaling pathway. Additionally, the MDA5 gene, as a significant cytosolic pathogen recognition receptor (PRR), its characteristics and functions have been extensively investigated, while less research has been done on the mechanisms of MDA5-miRNA mediated gene regulation. In this study, the evolution and functional characterization of MDA5 from miiuy croaker (mmiMDA5) were characterized. Comparative genomic analysis demonstrated that the ascidiacea and superclass do not have the MDA5 gene in the process of evolution. MDA5 contains four structural domains: CARD, ResIII, Helicase C, and RIG-I C-RD. The MDA5 was ubiquitously expressed in all tested miiuy croaker tissues. Moreover, the expressions were significantly up-regulated after stimulation with poly (I: C), which indicated that MDA5 might be involved in the antiviral immune response. The bioinformatics predicted programs have indicated that miR-203 has a direct negative regulatory effect on MDA5 in miiuy croaker. Furthermore, the dual-luciferase reporter assay have showed that miR-203 was the direct negative regulator of MDA5 in miiuy croaker. This study is the first to demonstrate that miRNA can suppress cytokines by regulating the RLR signaling pathway in teleost fish, providing some new ideas for studying miRNA-mediated regulation of immune responses in mammals.

Miiuy croaker, Miichthys miiuy is an ecologically important teleost species which is widely distributed in southeast coast of China. In this study, we present a chromosomal-scale genome assembly of the miiuy croaker which is an important Sciaenidae fish and economical species. We adopted Oxford Nanopore and Hi-C sequencing techniques to achieve an assembly with high accuracy and completeness. The investigation of genome characteristic and functional features may provide insights into the study of phylogenetic diversity of miiuy croaker. This study can also be beneficial to improve molecular assisted breeding techniques. Moreover, it can be a great resource to better conduct further study of other sciaenids.

MyD88 is a critical adaptor in the TLRs signaling pathway, which can activate NF-κB signaling pathway and promote proinflammatory cytokines production. However, the molecular mechanisms that modulate MyD88 expression, especially in teleost, remain largely unknown. In this study, we showed that NOP56 serve as a negative regulator of the MyD88-mediated NF-κB signaling pathway. NOP56 overexpression inhibited the protein expression of MyD88. Whereas, siRNA knockdown of NOP56 had opposite effect. Furthermore, we found that the NOSIC domain is responsible for the suppressive effect of NOP56 in MyD88 expression at protein level. Therefore, we identified NOP56 as a negative regulator of MyD88-mediated NF-κB signaling by inhibiting MyD88 expression and provided new insight into the regulation mechanism in teleost fish.

MyD88 is a typical street protein of the TLRs signaling pathway and is a central player in innate immune signaling, which can regulate the NF-κB signaling pathway and promote downstream inflammatory factors. However, studies on the molecular mechanisms of the MyD88-mediated NF-κB signaling pathway in teleosts have been poorly reported. In this study, we report that Zw10 targets MyD88 to inhibit NF-κB activation. Zw10 inhibits cell proliferation and MyD88-mediated innate immunity in fish. Zw10 interacts with MyD88, and its Δ2 domain is very critical for MyD88 degradation. In addition, we found that Zw10 degrade MyD88 by autophagy, thereby negatively regulating the MyD88-mediated NF-κB signaling pathway. This study not only enriches the research on the innate immunity of teleost fish, but also provides insights for the regulating mechanism for mammals.

Circular RNAs (circRNAs) are a subgroup of endogenous noncoding RNA that is covalently closed rings and widely expressed. In recent years, there is accumulating evidence indicating that circRNAs are a class of important regulators, which play an important role in various biological processes. However, the biological functions and regulation mechanism of circRNAs in lower vertebrates are little known. In this study, we discovered a circRNA Samd4a (circSamd4a) that is related to the antiviral immune response of teleost fish. It can act as a key regulator of the host’s antiviral response and play a key role in inhibiting Sininiperca chuatsi rhabdovirus replication. Further studies have shown that circSamd4a may act as a competing endogenous RNA, which can enhance the STING-mediated NF-κB/IRF3 signaling pathway by adsorbing miR-29a-3p, thereby enhancing the antiviral immune response. Therefore, circSamd4a plays an active regulatory role in the antiviral immune response of bony fish. Our research results provide a strong foundation for circular RNA to play a regulatory role in the antiviral immune response of 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.

The innate immune system is the first line of defense against the invasion of pathogens. It can make a rapid immune response to the invading pathogenic microorganisms, thereby eliminating the invading pathogens and protecting the body from harm. microRNAs are a family of small non-coding ribonucleic acid molecules, which are important and multifunctional regulator of immune response. In this study, we studied the role of miR-132 as a key regulatory factor of IL-1β-mediated inflammation. The seed region of miR-132 can regulate gene expression by binding to the 3'UTR of IL-1β, and inhibit the expression of IL-1β at the post-transcriptional level. More importantly, miR-132 inhibits LPS-induced NF-κB signaling pathway by targeting IL-1β, thereby preventing excessive inflammatory response from causing autoimmune diseases. These results will help to better understand the complex regulatory mechanisms of teleost fishes.

The inflammatory response is a protective process to clear detrimental stimuli, constitutes the defense against infectious pathogens. Clearing pathogen infection requires appropriate immune and inflammatory response, but excessive inflammatory response can lead to uncontrolled inflammation, autoimmune disease, or pathogen transmission. Accumulating evidences show that miRNAs are important and multifunctional regulators of innate immunity and inflammation. However, in the inflammatory response of lower vertebrates, the miRNAs regulatory networks are largely unknown. In this study, a combination of bioinformatics and experimental techniques were used to investigate the functions of miR-148. IL-1β is a hypothetical target gene of miR-148 predicted by bioinformatics. In addition, dual-luciferase reporter gene experiment was used to verify the targeting effect of miR-148 on IL-1β-3'UTR. miR-148 inhibits IL-1β expression in a dose-dependent manner at protein and mRNA levels. It is important that miR-148 participates in regulation of LPS-induced the NF-κB signaling pathway by inhibiting IL-1β. These results will improve our understanding of the regulation of miRNAs in fish on the immune response.

Circular RNAs (circRNAs) represent a class of widespread, diverse, and covalently closed circRNAs that function as microRNA (miRNA) sponges and crucial regulators of gene expression in mammals. However, the regulation and function of circRNAs in lower vertebrates are still unknown. Here, we first discover a highly conserved circRNA termed circRasGEF1B, which displays a high conservation from mammals to fish and serves as key regulator in eliciting antiviral immunity in teleost fish. Results indicate that circRasGEF1B was highly expressed in Siniperca chuatsi rhabdovirus-infected tissues and cells. Functionally, miR-21-3p could inhibit cellular antiviral responses significantly, whereas circRasGEF1B counteracts the effects of miR-21-3p. In mechanism, the results demonstrate that circRasGEF1B acts as a competing endogenous RNA (ceRNA) of miR-21-3p to relieve the repressive effect of miR-21-3p on its target MITA, then enhance the innate antiviral responses. Our results not only provide a novel insight into the functions of circRNAs in lower vertebrates but also broaden our understanding of circRNAs in viral infection. IMPORTANCE Siniperca chuatsi rhabdovirus (SCRV) is a typical fish RNA rhabdovirus, which is one of the most significant viral pathogens in teleost fish and can cause severe hemorrhagic septicemia in freshwater and marine fishes. Here, we discovered a highly conserved circRNA called circRasGEF1B, which acts as a key regulator for innate antiviral responses upon SCRV infection. circRasGEF1B acts as an endogenous sponge of miR-21-3p that downregulates miR-21-3p expression levels. circRasGEF1B is able to bind to miR-21-3p directly and regulates MITA expression. To our knowledge, this report is the first to characterize circRNA-miRNA regulatory networks that exist in lower vertebrates.

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., Author summary Increasing evidence indicates that circRNAs participate in innate and adaptive immunity. However, the relationship between circRNAs and host antiviral responses remains unknown, particularly in lower vertebrates. Our results provided direct evidence that a circRNA, namely, circDtx1, is related to the antiviral responses in lower vertebrates. In addition, our study also found a microRNA, namely, miR-15a-5p, which could inhibit an antiviral immune response and promote viral replication by targeting TRIF. Moreover, circRNAs can serve as competing endogenous RNAs (ceRNAs) and cross-talk with mRNAs by competing shared miRNAs. Such ceRNAs regulate the distribution of miRNA molecules on their targets and apply an additional level of post-transcriptional regulation. In our study, circDtx1 functions as a ceRNA for miR-15a-5p to control protein abundance of fish TRIF, thereby inhibiting viral replication and promoting antiviral responses. Our study demonstrates the ceRNA regulatory networks existing in lower vertebrates, which can provide new insights into understanding the effects of circRNAs on host-virus interactions.

The innate immune response is first line of host defense against pathogen invasion. However, excessive activation of immune responses may cause autoimmune diseases and excessive inflammation. Retinoic acid-inducible gene I (RIG-I) is an important cytoplasmic pathogen recognition receptor that is activated on virus infection. TNF-receptor-associated factor 6 (TRAF6) plays an essential role in the RIG-I-mediated signaling pathway. MicroRNAs (miRNAs) are noncoding RNAs that are emerging as important regulators of immune responses. In this study, we found that the overexpression of miR-489 mimics and pre-miR-489 significantly suppressed the luciferase activity of the wild-type TRAF6 3'UTR, whereas mutant-type led to a complete abrogation of the negative effect. In addition, we also observed that miR-489 can negatively regulate TRAF6 at the level of translation. More importantly, we demonstrated that miR-489 is a negative regulator of TRAF6 involved in the immune response to poly(I:C) stimulation. These common findings indicated that miR-489 plays a regulatory role in host-virus interactions by targeting TRAF6. Overall, all of the present results provide direct evidence that miR-489 is involved in the regulation of TRAF6 expression in miiuy croaker, which will help to better understand the complex regulatory networks of teleost fish.

Optimal activation of NF-κB signaling is crucial for the initiation of inflammatory responses and eliminating invading bacteria. Bacteria have likewise evolved the ability to evade immunity; however, mechanisms by which bacteria dysregulate host NF-κB signaling are unclear. In this study, we identify eukaryotic translation initiation factor eIF3k, a nonessential member of the eIF3 translation initiation complex, as a suppressor of the NF-κB pathway. Mechanistically, we show that eIF3k expression induced by Vibrio harveyi enhances E3 ligase Nrdp1-mediated K27-linked ubiquitination of MyD88, an upstream regulator of NF-κB pathway activation. Furthermore, we show that eIF3k acts as a bridge linking ubiquitin-tagged MyD88 and ATG5, an important mediator of autophagy. We demonstrate that the MyD88-eIF3k-ATG5 complex is transported to the autophagosome for degradation, and that innate immune signaling is subsequently terminated and does not attack invading V. harveyi. Therefore, our study identifies eIF3k as a specific inhibitor of the MyD88-dependent NF-κB pathway and suggests that eIF3k may act as a selective autophagic receptor that synergizes with ATG5 to promote the autophagic degradation of MyD88, which helps V. harveyi to evade innate immunity. We conclude that V. harveyi can manipulate a host's autophagy process to evade immunity in fish and also provide a new perspective on mammalian resistance to bacterial invasion.

Increasing evidence shows that the long noncoding RNA (lncRNA) is a major regulator and participates in the regulation of various physiological and pathological processes, such as cell proliferation, differentiation, metastasis, and apoptosis. Unlike mammals, however, the study of lncRNA in lower invertebrates is just beginning and the extent of lncRNA-mediate regulation remains unclear. Here, we for the first time identify an lncRNA, termed nucleotide oligomerization domain 1 (NOD1) antibacterial and antiviral-related lncRNA (NARL), as a key regulator for innate immunity in teleost fish. We found that NOD1 plays an important role in the antibacterial and antiviral process in fish and that the microRNA miR-217-5p inhibits NOD1 expression and thus weakens the NF-κB and the IRF3-driven signaling pathway. Furthermore, our results indicated that NARL functions as a competing endogenous RNA (ceRNA) for miR-217-5p to regulate protein abundance of NOD1; thus, invading microorganisms are eliminated and immune responses are promoted. Our study also demonstrates the regulation mechanism that lncRNA NARL can competitive adsorption miR-217-5p to regulate the miR-217-5p/NOD1 axis is widespread in teleost fish. Taken together, our results reveal that NARL in fish is a critical positive regulator of innate immune responses to viral and bacterial infection by suppressing a feedback to NOD1-NF-κB/IRF3-mediated signaling.

The inflammatory response is a self-defense process that fights the pathogen invasion by eliminating harmful stimuli. However, excessive inflammation may disrupt immune homeostasis, even causing chronic inflammation or autoimmune diseases. MicroRNAs (miRNAs) are a crucial regulator that can negatively regulate gene expression and participate in multiple biological processes of growth, development, and immune response in organisms. However, the miRNA-mediated modulation networks of inflammatory responses remain largely unclear in lower vertebrates. In this study, miR-128 was identified as a negative regulator to participate in the NF-κB signaling pathway by targeting TAB2 in miiuy croaker. First, we predicted target genes of miR-128 through the bioinformatics software programs and found that TAB2 is a direct target of miR-128. We also found that miR-128 can inhibit TAB2 expression at the mRNA and protein levels. Besides, upon LPS stimulation, miR-128 inhibits the expression of inflammatory cytokines by targeting TAB2 to avoid excessive inflammation. Particularly, we found that miR-128 can regulate TAB2-mediated NF-κB signaling pathways. In summary, our results indicate that miR-128 plays a critical role in suppressing inflammatory responses by regulating the TAB2-mediated NF-κB signaling pathway in miiuy croaker.

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.

Pathogen infection can cause the production of inflammatory cytokines, which are key mediators that cause the host’s innate immune response. Therefore, proper regulation of immune genes associated with inflammation is essential for immune response. Among them, microRNAs (miRNAs) as gene regulator have been widely reported to be involved in the innate immune response of mammals. However, the regulatory network in which miRNAs are involved in the development of inflammation is largely unknown in lower vertebrates. Here, we identified two miRNAs from miiuy croaker (Miichthys miiuy), miR-210 and miR-3570, which play a negative regulatory role in host antibacterial immunity. We found that the expressions of miR-210 and miR-3570 were significantly upregulated under the stimulation of Gram-negative bacterium vibrio harveyi and LPS (lipopolysaccharide). Induced miR-210 and miR-3570 inhibit inflammatory cytokine production by targeting RIPK2, thereby avoiding excessive inflammation. In particular, we found that miR-210 and miR-3570 negatively regulate antimicrobial immunity by regulating the RIPK2-mediated NF-κB signaling pathway. The collective results indicated that both miRNAs are used as negative feedback regulators to regulate RIPK2-mediated NF-κB signaling pathway and thus play a regulatory role in bacteria-induced inflammatory response.

Inflammation is a self-protection mechanism that can be triggered when innate immune cells detect infection. Eradication of pathogen infection requires appropriate immune and inflammatory responses, but excessive inflammatory responses can cause uncontrolled inflammation, autoimmune diseases, or pathogen dissemination. Mounting evidence has shown that microRNAs (miRNAs) in mammals act as important and versatile regulators of innate immunity and inflammation. However, miRNA-mediated regulation networks are largely unknown in inflammatory responses in lower vertebrates. Here miR-144 and miR-217 are identified as negative regulators in teleost inflammatory responses. We find that Vibrio harveyi and lipopolysaccharide (LPS) treatment significantly upregulate the expression of fish miR-144 and miR-217. Upregulated miR-144 and miR-217 suppress LPS-induced inflammatory cytokine expression by targeting nucleotide-binding oligomerization domain-containing protein 1 (NOD1), thereby avoiding excessive inflammatory responses. In addition, miR-144 and miR-217 regulate inflammatory responses through NOD1-induced nuclear factor kappa (NF-kB) signaling pathways. These findings demonstrate that miR-144 and miR-217 play regulatory roles in inflammatory responses by modulating the NOD1-induced NF-κB signaling pathway.

Upon recognition of bacterial or viral components by pattern recognition receptors, cells could be activated to produce inflammatory cytokines, type I IFN, and IFN-stimulated genes. These antibacterial and antiviral immunities are tightly regulated by the host to prevent inappropriate immune responses. MicroRNAs (miRNAs) have emerged as an essential regulatory network with profound effects on mammalian inflammation and immune responses, but the regulatory networks of miRNA-mediated immune response in lower vertebrates remain largely unknown. In this study, we report a miRNA, miR-217, identified from miiuy croaker, which plays a negative role in host antiviral and antibacterial immunity. We found that miR-217 could be abundantly expressed upon Gram-negative bacteria, as well as rhabdovirus infection. Inducible miR-217 suppresses the production of inflammatory cytokines and type I IFN by targeting TAK1, thereby avoiding excessive inflammation. Particularly, we revealed that miR-217 modulates the antibacterial and antiviral immunity through TAK1-mediated NF-κB and IRF3 signaling pathways. The collective results indicate that miR-217 acts as a negative feedback regulator involved in host antibacterial and antiviral immune responses, which will provide insights into the intricate networks of host–virus interaction in lower vertebrates.

Objectives. To study the quantification of the components in rat plasma after oral administration of Shenyanyihao oral solution. Methods. Shenyanyihao oral solution has been traditionally used for the treatments of chronic nephritis in clinics. Stachydrine, Danshensu, chlorogenic acid, protocatechuic acid, plantamajoside, aesculetin, isoquercitrin, ferulic acid, baicalin, and baicalein are regarded as the main compounds in Shenyanyihao oral solution. A sensitive, efficient, and precise UPLC-MS/MS method was established and validated for the quantification of the components in rat plasma after oral administration of Shenyanyihao oral solution. Results. The main pharmacokinetic parameters of the components were acquired based on the analysis of the plasma sample by a noncompartmental method using the WinNonlin7.0 pharmacokinetic program. Danshensu, protocatechuic acid, isoquercitrin, and ferulic acid from Shenyanyihao oral solution were quickly absorbed, and their peak concentration occurred at less than 0.5 h. The pharmacokinetic parameter of the average t1/2 from Danshensu was 3.91 h in rats, and it was the most rapid distribution and elimination among the components. In addition, the Cmax of stachydrine and baicalin were revealed as the higher plasma concentrations in rats. Conclusions. This pharmacokinetic study seems to be useful for a further clinical study of Shenyanyihao oral solution in the treatments of chronic nephritis.

This research aimed to investigate the composition and diversity of endophytic bacterial community in seeds of four hybrid maize and their parental lines, which was used to reveal the potential relationship and association of endophytic bacteria between maize genotypes and their genetic relevance. High-throughput sequencing (HTS) technology showed that a total of 1419 OTUs (46.6%) were parental lines unique and 1052 OTUs (34.5%) were hybrid varieties unique, with only 575 core OTUs revealed in all the samples. Most OTUs belonged to Proteobacteria. Enterobacter (23.2%), Shigella (21.2%), Pseudomonas (15.8%) and Achromobacter (10.1%) were the major genera; the bacterial community composition and diversity of endophytic bacteria were inconsistent among different seed genotypes. Based on principal component analysis (PCA), the results referred that the endophytic composition of hybrid sample showed obvious correlation with their female parental lines, and in ‘Jingke968’ and ‘MC738’ with the same female line the endophytic community was more similar than other hybrid samples. This was the first ever use of HTS technology for investigating the endophytic bacterial diversity and community structures in seeds of genetically related maize genotypes, it was shown that, there were core microbes shared among all genotypes of seed samples, and the female parental line was more significant to impact on the composition of their hybrid seeds than male parental line. This study would provide scientific clues for the future research on the vertical transmission of endophytes among maize generations. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (10.1007/s13205-019-2034-8) contains supplementary material, which is available to authorized users.