RNA Interference to Evaluate Role of Host Factors in Influenza RNA Expression

RM 207

Influenza virus causes health and economic hardships worldwide. Influenza is controlled with annual vaccine of variable efficacy, and antivirals that become ineffective with use due to selection of resistance. Host proteins are integral for viral replication; host proteins that are essential for viral replication but redundant for the host cell represent a novel class of antiviral targets. As Peer Research Consultant for BIO 592, I led students to employ RNA interference to knockdown host factors and determine the effect on influenza virus RNA expression. Student’s transfected cells with siRNA or no siRNA control for 48 hours prior to infection for 4 hours. I collected cells and students isolated and analyzed RNA by reverse transcription-quantitative PCR. Influenza transcription occurs in the nucleus, so student groups examined host RNA nuclear export factors, including Nxf1, responsible for mRNA nuclear export, Crm1, responsible for rRNA nuclear export, XpoT, responsible for tRNA nuclear export, and Xpo5, responsible for microRNA nuclear export. Each siRNA treated sample demonstrated a decrease in target mRNA, as expected. We found Nxf1 inhibition results in a severe decrease in viral RNA expression, verifying previous results. Interestingly, Xpo5 inhibition showed an increase in viral RNA expression, supporting the idea microRNAs function to counter influenza replication. One group targeted host RNA helicases, UAP56 and URH49, paralogs implicated in influenza replication. Unfortunately, URH49 siRNA cross-reacts with UAP56, limiting our conclusions. Future studies will continue to use RNA technologies with the aim to identify host factors as viable antiviral targets to counter influenza.