| Innate immunity,also known as natural immunity,is the first line to defense pathogens.Different organisms live in different environments and therefore have their own set of immune systems.This system functions through various pattern recognition receptors(PRRs)to rapidly recognize pathogen-associated molecular patterns(PAMPs).The immune system quickly transmits signals to key downstream adaptor proteins to activate various hosts.immune response.In this process,the host cells of the body can respond quickly,produce corresponding immune factors,and quickly eliminate or block pathogenic microorganisms to deal with the invasion of foreign pathogens.Long-term studies have shown that non-coding RNAs(ncRNAs)exist in a class of RNAs that can act as immune regulators in organisms,known as long non-coding RNAs(lncRNAs),which can be involved in gene imprinting,dose compensation,transcriptional enhancement and antisense regulation.lncRNAs can modulate targets through different mechanistic models.For example,some lncRNAs can directly interact with heteronuclear ribonucleoproteins or chromatin modification complexes,while some lncRNAs function as competing endogenous RNAs(ce RNAs)acting as competition elements and interact with m RNAs by competing with shared mi RNAs effect.Mediator of IRF3 activation,MITA,also known as stimulator of interferon genes(STING),has been identified as a key mediator of the innate antiviral response.In mammals,various studies have identified MITA as an important transcriptional inducible signaling molecule in type I interferon responses to DNA viruses and certain RNA viruses.Furthermore,the mechanism by which MITA mediates signaling pathways has been extensively studied in innate antiviral responses.In teleost fish,it has also been confirmed that MITA can target and regulate some gene-mediated signaling pathways to regulate innate immunity.Since most of the above studies were performed in mammals,there are few studies on lncRNAs in lower vertebrates.Miiuy croaker(Miichthys miiuy),as one of the important marine economic fish,can bring huge economic benefits to my country’s fishery production every year.Its whole body has high economic and medicinal value.In recent years,with the progress of industrialization,marine pollution has intensified,and pathogens have become more and more serious to the destruction of marine organisms.Therefore,there is an urgent need to reduce the economic losses caused by pathogenic microorganisms to the aquaculture industry by studying the relevant immune regulation mechanisms of minnows.In the present study,a novel lncRNA,LTCONS7822,was reported,which could positively regulate the antiviral immunity of miiuy croaker.Through targeted immunological experiments,the conclusions of this study are as follows:1.Through the gene characterization analysis of lncRNA LTCONS7822,we compared the lncRNA sequences of closely related species with miiuy croaker,and found that it has a high similarity in teleost fish,and we believe that it has certain evolutionary conservation in vivo.Linear gene analysis of miiuy croaker showed that the lncRNA LTCONS7822 was located between the genes ccnd2 b and llph.The lncRNA LTCONS7822 was predicted to have shallow coding potential by the Potential Coding Calculator(CPC),and the non-coding nature of lncRNA LTCONS7822 was verified by Western blotting experiments.2.In the q RT-PCR experiment,the expression level of lncRNA LTCONS7822 in miiuy croaker liver cells(MLC)after poly(I:C)stimulation or Siniperca chuatsi rhabdovirus(SCRV)infection stimulation was detected in this study.The experimental results showed that the expression of lncRNA LTCONS7822 changed significantly under poly(I:C)stimulation or SCRV infection.In addition,lncRNA LTCONS7822 and viral factors were transfected into miiuy croaker liver cells,and then the transfected mink fish liver cells were treated with different stimuli respectively.The results of real-time fluorescence quantitative assay showed that lncRNA LTCONS7822 could promote the expression of antiviral genes.3.This study found that the lncRNA LTCONS7822 could up-regulate the expression of MITA at the m RNA and protein levels.By using q RT-PCR experiments,the experimental results proved that the overexpression plasmid of lncRNA LTCONS7822 can up-regulate the expression of MITA,and further demonstrated by western blotting experiments that at the protein level,lncRNA LTCONS7822 can promote the protein expression level of MITA.4.Based on MITA identified as a key signaling molecule in innate immunity triggered by cytosolic virus or microbial nucleic acid,and has been shown to regulate NF-κB and IRF3 signaling pathways in mammals and teleost fish,the lncRNA LTCONS7822 was further explored.By targeting the MITA-involved regulation of the signaling pathway,and by detecting the effect of the over-expressed MITA expression plasmid in Endothelial Progenitor Cells(EPC cells)on the activity of different reporter genes under stimulation,the experimental results show that the over-expression of the MITA-expressing plasmid can significantly activate the reporter gene.And further verified that lncRNA LTCONS7822 upregulates NF-κB and IRF3 signaling pathways by targeting MITA.In conclusion,lncRNA LTCONS7822,as a regulator,plays an important regulatory role in MITA-mediated signaling pathways.In summary,the experimental results indicated that lncRNA LTCONS7822,as a regulator,is involved in regulating the immune response induced by viral infection.This study provides a theoretical basis for further exploring the role of lncRNAs in the regulatory network of immune response in fish pathogen infection.This tightly regulated mechanism of positive regulation is critical for mitigating viral responses and avoiding viral evasion.These data not only provide information on lncRNAs as feedback regulators involved in antiviral immune responses in fish,but also enrich the network of hostpathogen interactions mediated by lncRNAs. |
