Figure 5. SGs prevent dsRNA-triggered cell death by suppressing RLR, PKR and OAS pathways.

A. Cell death in U2OS cells as measured by Sytox uptake (left) and bright field microscopy (right) at 24 hr post-dsRNA.

B. Apoptotic caspase cleavage in Δ U2OS cells at 14 or 24 hr post-dsRNA.

C. Levels of IFNβ and TNFα mRNAs in U2OS cells at 6 hr post-dsRNA. Data were normalized to WT at 6 hr post-dsRNA.

D. Heat map of z-scores for differentially expressed genes in apoptosis pathway (KEGG pathway hsa04210) in U2OS cells at 6 hr post-dsRNA stimulation.

E. Level of secreted TNFα in U2OS cells 6 hr post-dsRNA.

F. Effect of anti-TNFα antibody on dsRNA-triggered cell death in U2OSΔG3BPs. Cells were pre-treated with anti-TNFα antibody (0.01, 0.1 and 1 μg/ml) 30 min prior to transfection with dsRNA. Cell death was measured by Sytox uptake at 24 hr post-dsRNA.

G. Cell death in ΔG3BPs and ΔG3BPsΔPKR at 24 hr post-dsRNA.

H. Cell death in ΔG3BPs and ΔG3BPsΔRNase L at 24 hr post-dsRNA.

I. Schematic for dsRNA-induced cell death in ΔG3BPs cells. The lack of SGs make ΔG3BPs cells hypersensitive to dsRNA, resulting in more potent activation of RLR, PKR and OASes. The TNFα signaling branch (but not the IRF3-IFN branch) downstream of RLR-MAVS makes the primary contribution to cell death in U2OS cells. PKR and OASes-RNase L also contribute, likely by suppressing global protein synthesis.

Data are presented in means ± SD. p values were calculated using two-tailed unpaired Student’s t test (ns, p>0.05). All data are representative of three independent experiments.