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. 2022 Apr 28;18(4):e1010163. doi: 10.1371/journal.ppat.1010163

Fig 2. Effects of Sec61A1 and Loquacious depletion on DENV2 RNA and protein abundances, and viral titers.

Fig 2

(A) Experimental outline. Mosquito Aag2 cells were transfected with double stranded RNAs (dsRNA) directed against GFP, Sec61A1, Loqs (targeting both PA and PB isoforms) or Loqs-PB mRNAs. 24 hrs post transfection, cells were infected with DENV2-NGC at an MOI of 0.1 and harvested 96 hrs post infection for analyses. (B) RT-qPCR measurement of DENV RNA abundances in dsRNA-treated cells plotted as fold change over treatment with dsGFP. Data was normalized to internal control RPL32 mRNA levels (n = 3, ****p<0.0001). (C) Effects of dsRNA treatment on DENV RNA and DENV subgenomic RNA (DENV-sfRNA) abundances, measured by Northern blot analysis. Methylene blue (MB) staining of rRNA was used as a loading control. Representative image from three independent experiments is shown. (D) Effects of dsRNA treatment on DENV NS3 and actin protein levels examined by western blot analysis. Numbers below the lanes represent the relative abundance of NS3 normalized against actin in each sample, as quantified by densitometric analysis using Image Lab 6.0. Representative image from three independent experiments is shown. (E) Effects of dsRNA treatment on infectious viral titers determined by plaque forming assays (PFU/ml, n = 4, *p<0.05).