Alterations In MicroRNA Expression Profile In Street Strain Rabies Virus-infected Mouse Neurons

Rabies virus (RABV), a member of the genus Lyssavirus of the familyRhabdoviridae, is a highly neurotropic virus that spreads along neural pathways andinvades the central nervous system (CNS), where it causes an acute infection. Rabiesoccurs in more than150countries and territories. Annually, more than55,000individuals die from rabies worldwide. The deadly RABV infection produces avariety of nervous system symptoms; however, patients eventually die of circulatoryinsufficiency. Despite the catastrophic clinical outcome of rabies encephalomyelitis,the fact that inflammatory changes and degenerative neuronal changes may beminimal or even absent under natural conditions, indicates that neuronal dysfunction,rather than neuronal death, is likely responsible for the clinical disease and fataloutcome in rabies under natural conditions. Although there are several hypothesesunder active investigation at the present time, the fundamental basis for neuronaldysfunction in rabies has not yet been demonstrated.MicroRNAs (miRNAs), an extensive class of non-coding RNA molecules, thesemolecules are22nucleotides in length and negatively regulates gene expression bybinding completely or partially to target protein-coding mRNAs at theposttranscriptional level for degradation or translational inhibition. Thousands ofmiRNAs have since been identified in various organisms and play essential roles ingene expression regulation of many critical cellular processes such as development,differentiation, proliferation, and hematopoiesis. A large body of evidence indicatesthat the alterations in cellular miRNA expression that take place during the viralinfection may be an important determinant of virulence and pathogenesis. Forexample, human T-cell lymphotropic virus type1, human cytomegalovirus,Epstein-Barr virus, influenza virus.Given the above mentioned information, this study was aimed to identify potential dysregulated miRNAs in street rabies virus-infected neuron. We havegenerated a global miRNA expression profile from primary neuronal cultures infectedwith mock and RABV, and two distinct trends were presented in which one trend wascontinuously upregulated with19miRNAs while the other was continuouslydownregulated with34miRNAs.Then we focuced on the miRNAs which specificlyexpressed in neuron and performed qRT-PCR measurement to verify the expressionlevels of ten differentially expressed miRNAs (miR-466i-5p，miR-29a，miR-21，miR-671，miR-680，miR-107，miR-124，miR-181a，miR-9，miR-99b) in primaryneuronal cultures and hippocampal tissues of mice. TargetScan successfully predicted48target genes of the differentially expressed miRNAs. A total of6172genes werepredicted as target genes for up-regulated miRNAs, while14780genes were predictedfor down-regulated miRNAs. To better understand the miRNA function in thepathogenesis of RABV, we submitted the predicted target genes to DAVID databasefor annotating these genes using GO term which provide a controlled vocabulary todescribe gene/gene product attributes, and for performing GO enrichmentanalysis.The GO enrichment analysis results of particular interest to neuronalfunction-related GO biological processes and cellular component, respectively(Bonferroni adjusted P <0.1). We conducted the pathway enrichment analysis of thepredicted target genes in DAVID database using KEGG pathway. The result ofparticular interest to neuronal function-related pathway (Bonferroni adjusted P <0.05).These target genes were significantly involved in axon guidance, Wnt signalingpathway, regulation of actin cytoskeleton, neurotrophin signaling pathway, mTORsignaling pathway and long-term potentiation. It demonstrated that miRNAs withsignificant change are involved in regulation of target genes related to neuronaldysfunction in response to street RABV infection.According to the existed miRNA expression profile, we pick the STX1A for theexperiment. Finally we used the Luciferase report array system and Western blot tovarify the target genes.In summary, this study contributes to the cognitive and molecular mechanism ofpathogenic RABV infection and the general rule of virus and host gene function relationships, Exploration and raised a host of miRNA-based RABV-host interactionsin the academic academic point of view, from the host system for the discovery ofdrug targets against RABV accumulate academic foundation, which would be veryhelpful in understanding neuronal dysfunction in rabies..