FIGURE 1. ADAR1 suppresses viral RNA-stimulated type I IFN expression in primary mouse cells.

Mouse primary fibroblasts were prepared from embryos carrying floxed ADAR1 gene and inducible Cre-ER gene (iKO MEF) or littermates without Cre gene (cMEF). (A) The floxed ADAR1 gene allele is illustrated before and after tamoxifen (TM) induction in iKO MEF and cMEF. (B) ADAR1 protein was completely depleted in iKO MEFs between 24–48 hours after adding 10nm 4-OH TM to culture medium, but did not affect ADAR1 protein in cMEFs. (C) iKO MEFs were split into two groups in parallel for TM treatment and non-TM treatment. IFN production was compared in the exact same cells, except TM induction. cMEFs were included to exclude the potential effect of TM. (D, E) Sendai virus (SeV) was used to stimulate the cells for IFN production. SeV infection induced significant IFN-α and IFN-β expression in these primary cells, as shown by the mRNA levels measured by real-time PCR. TM had no effect on cMEFs, but it caused dramatic increases in IFN-α and IFN-β expression in iKO-MEFs. (F, G) Transfection of poly I:C into iKO MEFs caused the same responses with or without TM induction, as observed in SeV infection. IFN mRNA was measured after six hours of SeV infection or poly I:C transfection. Expression level of each sample was normalized by its beta-actin expression and the fold change was relative to the non-stimulated INF expression level. Data shown in this figure is mean ± S.D. (n=3. Experiments repeated five times with MEFs from different embryos). * indicates p<0.05; ** p<0.01; *** p<0.001.