Functional Analysis of microRNAs:mRNA Interactions Between Pseudorabies Virus (PrV) and Its Natural Pig Host

During a viral infection, complex interactions take place between the virus and its host. It has been shown that microRNAs play a major role in this context. Some host microRNAs are crucial to immune responses, with few of them proposed to directly target viral genes and to inhibit viral replication, while microRNAs encoded by some viruses can modulate the transition from latent to lytic replication and attenuate the host antiviral immune responses. Such interactions occur through molecular interactions with target mRNAs via the RNA-induced silencing complex (RISC).

An interesting model to study these interactions is the pseudorabies virus (alphaherpesvirus, PrV), causing Aujeszky's disease in pigs. We have previously shown that a deletion containing a cluster of nine microRNAs in the Latency-Associated-Transcript Locus of the PrV genome did not hamper the establishment of PrV latency in trigeminal ganglia. However the absence of miRNAs affected the immune pathways of the host response. Prv-miR-LLT1, prv-miR-LLT2 and prv-miR-LLT10 were the most expressed PrV miRNAs by the wild-type virus.

Here we adopted a functional approach to test for the potential effect of the most expressed PrV miRNAs on the host cell. Our strategy is based on microRNA transfection followed by RNA-binding protein immunoprecipitation and microarray analysis of the RISC-bound mRNA (RIP-Chip). We identified PrV microRNA host targets using RIP-Chip enrichment analysis performed by the Bioconductor package anota. A total of 193 PrV-miR targets, that have functional roles in relevant pathways, were identified.