Study On The MicroRNA Expression Profiles Of Pseudorabies Virus And Bovine Herpesvirus Type5

Pseudorabies virus (PRV) and Bovine herpesvirus type5(BHV-5) belongs to Alphaherpesvirinae subfamily which causes Aujeszky’s disease in pig and meningoencephalitis in young cattle, respectively, resulting in enormous economic loss worldwide. Viral miRNAs play significant roles in the life cycles and virus-host interaction of herpesviruses, however, research on PRV-encoded or BHV-5-encoded miRNAs are extremely limited. This study aim to identify miRNA expression profiles of the two viruses and to analyse the functions of these viral miRNAs in the viruses’life cycles and virus-host interactions.Total RNAs from PRV-infected PK-15cells and BHV-5-infected MDBK cells at different time points were extracted and then subjected to solexa sequencing. Identified viral miRNAs were further confirmed using northern blot and stem-loop RT-PCR. Bioinformatic approaches were adopted to investigate the miRNAs-targets interaction networks between viral miRNAs and viral targets or between viral miRNAs and host targets. The host miRNA expression profiles in response to virus infection were also analysed.Deep sequencing data revealed that the-4.6kb intron of the PRV-encoded LLT functions as a pri-miRNA that encodes a cluster of11distinct miRNAs, and7of them are novel. Viral miRNAs were further confirmed by stem-loop RT-PCR and northern blot analysis.209known and39novel porcine miRNAs were detected in PRV-infected PK-15cells. All of these viral miRNAs exhibited terminal heterogeneity both at the5’and3’ ends. Seven pre-miRNAs produced mature miRNAs from both arms and two of the pre-miRNAs were partially overlapped. Additionally, a terminal loop-derived small RNA with high abundance and one special moRNA were possiblely processed from a same viral miRNA precursor. Target analysis revealed a complex network formed by viral miRNAs and host targets or viral targets. GO enrichment analysis of host targets suggests that PRV miRNAs are involved in complex cellular pathways including cell death, immune system process, metabolic pathway, etc..Meanwhile,17pre-miRNA loci, containing11sequence-specific pre-miRNAs which were processed to16mature miRNAs and miRNA s were identified in BHV-5genome. Compared with BHV-1-encoded miRNAs, both have several miRNAs located in their UL regions and repeat regions. In BHV-5-infected MDBK,339annotated and327novel bovine miRNAs were identified, while the No. for mock-infected MDBK are285and336respectively. Unexpectedly, only one concerved miRNA was identified between the two viruses. Prominent change of the host miRNA expression profile was observed after the BHV-5infection. In the total137differentially expressed bovine miRNAs,31of them were significantly down-regulated and44were predominantly up-regulated. Target analysis and GO analysis revealed that the differentially expressed host miRNAs are invoved in various cellular processes. Bioinformatic analysis shows that BHV-5-encoded miRNAs can not only target viral genes, e.g., bICPO and bICP22, but also regulate numorous host targets.This is the first study which identified miRNA expression profiles of PRV and BHV-5in epithelial cells. Simultaneously, the host miRNA profiles were analysed as well. Besides, we made comprehensive analysis of the regulation networks of viral miRNAs with either viral targets or host targets. Collectively, this study layed foundation for functional studies on these viral miRNAs, and will provide guidance for subsequent relevant research.