Fig. 4

UPR-induced antiviral signaling following TBEV infection requires the IRE1 pathway. a, b PERK depletion did not affect TBEV yields. U2OS cells stably transduced with shPERK under puromycin selection were infected with TBEV and harvested for immunoblotting at 16 and 24 h p.i., Actin is the protein loading control (a). At the indicated time points, supernatants from infected cells were used to infect Vero cells to measure virus yields (b). c, d ATF6 depletion did not affect TBEV yields. U2OS cells were transfected with siATF6/siCTRL and after 48 h infected with TBEV. Cells were harvested for immunoblotting at 16 and 24 h.p.i. Actin is the protein loading control (c). At the indicated time points, supernatants from infected cells were used to infect Vero cells to measure virus yields (d). e, f IRE1 depletion increased TBEV yields. U2OS cells were treated as above (c, d) with siIRE1. g, h IRE1 depletion increased TBEV replication but did not affect IFNβ mRNA. U2OS cells were depleted for IRE1 as above (e). Total RNA was extracted at the indicated time points and used as template for real-time qPCR using primers specific for TBEV (g) or IFNβ (h). TBEV RNA amplicons were normalized to β-actin RNA and plotted as fold change from time 0. i, j IRE1 depletion partially rescued TBEV replication in TM-treated cells. U2OS cells were either infected with TBEV (blue bars) or treated with TM immediately after infection (black bars) in conditions of IRE1 depletion. Immunoblot for IRE1 and TBEV RNA quantification was performed at 24 h.p.i. k, l IRF3 depletion partially rescued TBEV replication in TM-treated cells. U2OS cells were treated as above (i, j) in conditions of IRF3 depletion. Immunoblot for IRF3 and TBEV RNA quantification was performed at 24 h.p.i. m, n RIG-I depletion partially rescued TBEV replication in TM-treated cells. U2OS cells were treated as above (i, j) in conditions of RIG-I depletion. Immunoblot for RIG-I and TBEV RNA quantification was performed at 24 h.p.i. Statistics as already described in the legend to Fig. 1. Source data are provided as a Source Data file