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. 2022 Jun 11;14(6):1273. doi: 10.3390/v14061273

Figure 6.

Figure 6

ORF6 blocks mRNA nuclear export along with transcriptional inhibition. (a) The effects at transcriptional level of ORF6 on IFN-β and IL-6 expression were evaluated in A549 cells. Total RNA was purified from empty vector (Ctr-), green-fluorescent protein (GFP), ORF6, or its M58R mutant transfected cells collected at both 24 h and 48 h post-transfection. Specific IFN-β and IL6 mRNA content was detected by quantitative reverse-transcription polymerase chain reaction (RT-qPCR). RNaseP gene expression was used for relative quantification based on 2−ΔΔCt method. (b) Total RNA was purified from A549 nuclear and cytoplasmic cellular fractions from negative control (Ctr-), green-fluorescent protein (GFP), ORF6 or its mutant M58R expressing cells, collected at both 24 h and 48 h post-transfection. Relative quantification of IFN-β and IL6 mRNAs was performed by 2−ΔΔCt method and the ratio between nuclear and cytoplasmic specific mRNAs content was used as a comparative method. (a,b) Three (n = 3) independent experiments were performed and representative data are presented as mean values ± standard deviations. Significance was evaluated as p < 0.05 (*** p < 0.0005, ** p < 0.005, * p < 0.05, n.s. = not significant).