Journal: Nature Communications

Article Title: A conserved long-range RNA interaction in SARS-CoV-2 recruits ADAR1 to enhance virus proliferation

doi: 10.1038/s41467-025-63297-2

Figure Lengend Snippet: a Structure model of a 158-base-long mini-construct that contains the long-range interaction LR1 base-pairing (mLR1). Mutations intended to disrupt the base-pairing from one side (mLR1A, green) or another side (mLR1B, orange) are indicated next to the original bases. Compensatory mutant mLR1AB contains both A and B mutations restoring the pairing of mLR1. b Top, PAGE showing proteins that were immunoprecipitated using biotinylated mLR1-WT RNA and negative control RNAs mLR1A (lane A), mLR1B (lane B) and Tetrahymena ribozyme (Teth). Input proteins from the whole cell lysate of Vero-E6 are also shown. Bottom, Western blot analysis of proteins from the pull-down assay using anti-ADAR1 antibody. c EMSA experiment showing the binding of different concentrations of ADAR1 with indicated RNAs. d Quantification of the bound RNA fraction shown in c . Data are presented as mean ± SD ( n = 3 independent experiments). e A-to-I RNA editing level along SARS-CoV-2 WT, LR1A, LR1B and LR1AB mutant genomes in infected cells. N = 3 independent experiments (total 6 replicates). f Box-plot showing the distribution of editing level of each edited site. The total number of edited sites across three biological replicates is indicated. g Box-plot showing the distribution of the editing index ( n = 6) in samples as described for panel ( e ). h A-to-I RNA editing level along SARS-CoV-2 WT genome in cells transfected using a random ASO (ASO_CTRL), an ASO that targets a different region (ASO_1, against 1563-1582), or an ASO designed against the LR1 region (ASO_LR1). N = 2 independent experiments (total 4 replicates). i Box-plot showing the distribution of the editing level of each edited site. Total number of edited sites across two biological replicates is indicated. j Box-plot showing the editing index in samples treated as described for panel h ( n = 4). For box-plots, the box represents the 25–75th percentiles, median is indicated as the line. The whiskers show the minimum and maximum values. P -values were calculated using two-sided Wilcoxon Rank Sum Test. ns, not significant. Source data are provided as a Source Data file.

Article Snippet: Western blot analysis was performed according to standard procedures using a 1:1000 dilution of anti-ADAR1 (E6X9R) XP(R) rabbit monoclonal antibody (Cell Signalling Technology) and a 1:5000 dilution of secondary anti-rabbit antibody.

Techniques: Construct, Mutagenesis, Immunoprecipitation, Negative Control, Western Blot, Pull Down Assay, Binding Assay, Infection, Transfection