Analysis Of Expression Profiles Of LncRNA And MRNA In Rabies Virus Infected Cells And Mice

Rabies is an infectious disease caused by rabies virus(Rabies virus,RABV)infection in the mammalian central nervous system(Central nervous system,CNS).It is a zoonotic disease with a high mortality.Once clinical symptoms occur,the mortality rate is as high as 100%.According to statistics from the World Health Organization,more than 50000 peoples die from rabies each year worldwide.The vast majority of cases occur in under-developed countries in Asia and Africa.RABV can infects nearly all warm-blooded animals.Wild animals are the main storage hosts of RABV,and humans and various mammals serve as the hosts of transmission.Viroal particles in the saliva of the RABV-infected animal invade the body by scratching or biting the host's skin,causing infection of CNS.Long-chain non-coding RNA(Long-chain noncoding RNA,lncRNA)is a type of RNA molec?Le with a length of more than 200 nt,which lacks a reading frame and does not encode a protein.It is distributed in the cytoplasm and nucleus of cells.Although lncRNAs do not encode proteins,they are involved in important processes such as DNA methylation,regulation of transcriptional activation and interference,intranuclear transportation and translation.LncRNA is involved in regulating of gene expression through functions of epigenetic modifications,transcriptional regulation and post-transcriptional regulation.The host's anti-RABV infection mainly relies on innate immunity,especially type I interferon response.After viral infection,cell surface receptors and intracellular receptors in vivo recognize the pathogen-associated molecular patterns(PAMPS),activating signaling pathways and inducing the secretion of type I interferons.More and more evidences showed that lncRNA plays an important role in the host antiviral response and innate immunity.However,the expression profile of lncRNA in RABV-induced host immune response is not yet known.Therefore,it is of great significance to elucidate the change of lncRNA expression profile induced by RABV infection in order to deeply explore the pathogenic mechanism of RABV and its prevention and control strategies.In this experiment,lncRNA and mRNA expression profiles in RABV infections were investigated by high-throughput small RNA sequencing analysis at the cellular and mouse brain tissue levels.Firstly,SRV9 and CVS11 were used to infect BHK21 cells respectively.After 12 h,24 h and 48 h infection,total RNA was extracted,library was constructed,and lncRNA and mRNA expression profiles were sequenced.The results showed that 940 lncRNAs were differentially expressed in the infected CVS11 group compared with the uninfected group,whereas 946 lncRNAs were differentially expressed in the infected SRV9 group.In the mRNA expression profile of infected BHK21 cells,5984 mRNAs were differentially expressed in the uninfected group compared to the SRV9 infected group,and 5674 mRNAs were differentially expressed in the infected CVS11 virus group(P < 0.05).GO enrichment analysis of lncRNA target gene showed that after 48 h of infection in the CVS11 group responses to cellular responses,activation of myeloid dendritic cells,modulation of macrophage production,and positive regulation of chemokine receptor activity were significantly enriched.After 48 h of infection in SRV9 group,GO items regulation of signal transduction,protein kinase and embryo morphogenesis were significantly enriched.The results of KEGG enrichment analysis showed that 16 KEGG pathways of the lncRNA target gene were significantly enriched in the CVS11 group after 48 h of infection,including pathways involved in the immune response,viral carcinogenesis,FoxO signaling pathway and MAPK signaling pathway,etc.and similar results were observed in SRV9 infection group.These findings indicate that lncRNAs modulate the immune and anti-viral response during RABV infection modulating its neighboring protein-coding genes.In this study,differentially expressed lncRNA and mRNA were selected for RT-qPCR validation.Secondly,rabies strain CVS11 was used to infect mouse brain tissue.Eight days after infection,tissues of mice was taken to extract total RNA,cDNA library was constructed and lncRNA and mRNA expression profiles were analyzed by sequencing.The results showed that a total of 140 lncRNAs were differentially expressed in the CVS11 infection group compared to the uninfected group.Chromosome 7,12 and 16 had the largest variation in lncRNA;chromosomes 18,19 and chromosome X had the smallest lncRNA differences.At the same time,we detected changes in mRNA expression profiles of CVS11-infected mouse brain tissue.The results showed that 3807 lncRNAs were differentially expressed in the CVS11 infection group compared with uninfected mice(FC ? 2,P < 0.05).The differential expression of chromosome 2,7 and 11 mRNA was the most significant,and the difference between chromosome 16 and 18 mRNA was significant.GO analysis showed that the 100 kb upstream or downstream of the lncRNA was highly enriched for the protein encoded by the isoform.The differentially expressed lncRNA co-localized genes are highly enriched in biological processes such as detoxification of nitrogen compounds,cell detoxification of nitrogen compounds,and nitrobenzene metabolism processes.KEGG analysis results showed that differentially expressed lncRNA targets involved in Toll-like receptors,NF-?B,MAPK,chemokines,herpes simplex and influenza A infection and other important signaling pathways.These findings indicate that lncRNA is involved in modulating the immune response during RABV infection.Subsequent real-time quantitative PCR was used to verify differential expression and the results were consistent with the differential expression of small RNA sequencing analysis of RABV infection.In this study,RABC-infected cells and mice were used to analyze the changes of lncRNA and mRNA expression profiles in vitro and in vivo,and differentially expressed lncRNA and mRNA were selected for RT-qPCR validation.Based on the analysis of biological information,the path of RABV infection-induced immune response in the host is preliminarily identified at the in vitro and in vivo levels.At present,there has been no study on the expression pattern of host lncRNA in RABV infection.This study provides data to support in elucidating the mechanism of immune escape and pathogenesis of RABV.