Figure 5
MiR-281 enhances virus replication in C6/36 cells. (A) miR-281 mimic treatment up-regulates DENV-2 gRNA. mock: normal cells + DENV-2; control mimic: control mimic + DENV-2; miR-281 mimic: miR-281 mimic + DENV-2. (B) antagomiR-281 treatment down-regulates DENV-2 gRNA. mock: normal cells + DENV-2; control antagomiR: control antagomiR + DENV-2; antagomiR-281: miR-281 antagomiR + DENV-2. The results of (A) (B) were analyzed using the 2-Δct method. DENV-2 gRNA (NS1 region) was quantified by RT-qPCR and normalized to rpS7 gene. Data shown represent three independent experiments with three triplicates each. P values were calculated using ANOVA. (C) Western blot analyses of C6/36 cells treated as in (A) and (B) for the detection of DENV-2 E protein. The E protein level is up-regulated by the miR-281 mimic and down-regulated by the miR-281 antagomiR. Mosquito beta-actin was used as control for protein loading. Control: normal cells; Mock: normal cells + DENV-2; Con mimic: control mimic + DENV-2; mimic: miR-281 mimic + DENV-2; Con antagomiR: control antagomiR + DENV-2; antagomiR-281: miR-281 antagomiR + DENV-2. (D) Semiquantitative analysis of E protein levels. Data shown represent three independent experiments. The western blot band intensities were measured with ImageJ software. E protein levels were normalized to beta-actin levels. P values were calculated using ANOVA. (E) DENV-2 titers of cells treated as (B) were measured by TCID50 in C6/36 cells and were calculated by log10TCID50. Data shown represent three independent experiments with three triplicates each. P values were calculated using ANOVA.