Inhibiting The Expression Of HCMV UL54 By Using EGS And Its Effect On Viral Replication

Aim: A few HCMV UL54 gene specific EGS were proved to be able to cleave target gene in our previous study. Amongst them, two DNA-based EGS, dEGS-T6 and dEGS-T7 were selected for the investigation on their efficiency in inhibiting the gene expression of HCMV UL54 and their effect on the viral replication In this study.Methods: The HFF cells were transfected with dEGS-T6 or dEGS-T7 at different concentration (100nM,80nM,60nM) followed by infection with AD169 (moi=1). The total cellular RNAs was collected 8hrs post virus-infection and EGS-transfection. The inhibition of target gene expression was monitored by quantitate the UL54 mRNA using real time fluorescent quantitative PCR. The supernatants of the infected HFFs cells were collected at 1-day intervals through 7 days postinfection. The number of copies of virus were caculated and used to draw the reduplication curve for analyzing the influence of the EGS on viral replication.Results: Although non-specific inhibition was observed with EGS application at high concentration (at both 100nM and 80nM), specific inhibition on UL54 expression was observed with dEGS-T6 at 60nM while no inhibition was observed with dEGS-T6 and dEGS-TK. Further studies was carried out on the potential influence that dEGS-T7 might have on viral replication. A reduction of about 67.27±0.10%～76.47±1.23% in HCMV growth was observed in the HFFs that were treated with dEGS-T7.Conclusion: The results in this study showed that dEGS-T7 had significant inhibition on both target gene expression and viral replication. It provided a potential application of dEGS-T7 as a new anti-HCMV drug.