Published online Nov 26, 2014. doi: 10.4331/wjbc.v5.i4.437
Revised: September 9, 2014
Accepted: October 1, 2014
Published online: November 26, 2014
AIM: To test whether the replication of human rhinovirus (HRV) is regulated by microRNAs in human bronchial epithelial cells.
METHODS: For the present study, the human cell line BEAS-2B (derived from normal human bronchial epithelial cells) was adopted. DICER knock-down, by siRNA transfection in BEAS-2B cells, was performed in order to inhibit microRNA maturation globally. Alternatively, antisense oligonucleotides (anti-miRs) were transfected to inhibit the activity of specific microRNAs. Cells were infected with HRV-1B. Viral replication was assessed by measuring the genomic viral RNA by reverse transcription quantitative polymerase chain reaction (RT-qPCR). Association between microRNA-induced-silencing-complex and viral RNA was detected by Ago2 co-immunoprecipitation followed by RT-qPCR. Targetscan v.6 was used to predict microRNA target sites on several HRV strains.
RESULTS: Here, we show that microRNAs affect replication of HRV-1B. DICER knock-down significantly reduced the expression of mature microRNAs in a bronchial epithelial cell line (BEAS-2B) and in turn, increased the synthesis of HRV-1B RNA. Additionally, HRV-1B RNA co-immunoprecipitated with argonaute 2 protein, an important effector for microRNA activity suggesting that microRNAs bind to viral RNA during infection. In order to identify specific microRNAs involved in this interaction, we employed bioinformatics analysis, and selected a group of microRNAs that have been reported to be under-expressed in asthmatic bronchial epithelial cells and were predicted to target different strains of rhinoviruses (HRV-1B, -16, -14, -27). Our results suggest that, out of this group of microRNAs, miR-128 and miR-155 contribute to the innate defense against HRV-1B: transfection of specific anti-miRs increased viral replication, as anticipated in-silico.
CONCLUSION: Taken together, our results suggest that pathological changes in microRNA expression, as already reported for asthma or chronic obstructive pulmonary disease have the potential to affect Rhinovirus replication and therefore may play a role in virus-induced exacerbations.
Core tip: Our results show for the first time that: (1) DICER knock-down increases HRV-1B replication in human bronchial epithelial cells; (2) the genomic RNA of human rhinovirus (HRV)-1B interacts directly with the miRISC during infection; and (3) inhibition of two microRNAs predicted to target HRV-1B, i.e., miR-128 and miR-155, favors viral replication. This supports a role for cellular microRNAs in the antiviral response to HRV-1B mounted by bronchial epithelial cells, and suggests that pathological microRNA dysregulation may contribute to the poor antiviral immunity in diseases such as asthma and chronic obstructive pulmonary disease.