Fig. 1.

Identification of SARS-CoV-2 M as an inhibitor of the viral RNA-triggered innate immune response. a Screening for SARS-CoV-2 proteins that inhibit the SeV-triggered activation of the IFN-β promoter. HEK293T cells were transfected with an IFN-β promoter luciferase plasmid and the indicated SARS-CoV-2 protein expression plasmids for 20 h and then infected with SeV (MOI = 1) or left untreated for 12 h before luciferase assays were performed. b The M protein inhibits the SeV-triggered activation of the IFNβ promoter, ISRE and NF-κB in a dose-dependent manner. HEK293T cells were transfected with the indicated reporter plasmids and increasing amounts of the Flag-M plasmid for 20 h and then infected with SeV (MOI = 1) or left untreated for 12 h before luciferase assays were performed. c The M protein inhibits the SeV-triggered transcription of antiviral genes in HEK293 cells. HEK293 cells stably expressing the M protein were left uninfected or infected with SeV (MOI = 1) for the indicated times before qPCR analysis was performed. d The M protein inhibits the poly (I:C)-triggered transcription of antiviral genes in HEK293 cells. HEK293 cells stably expressing the M protein were mock-transfected or transfected with poly (I:C) for 6 h before qPCR analysis was performed. e The M protein inhibits the SARS-CoV-2-triggered transcription of antiviral genes in HEK293 cells. HEK293-ACE2 cells were transfected with the Flag-M plasmid for 20 h and then infected with SARS-CoV-2 (MOI = 1) or left uninfected for the indicated times before qPCR analysis was performed. f Effects of M on viral genome replication during SARS-CoV-2 infection in HEK293-ACE2 cells. HEK293-ACE2 cells were transfected with the Flag-M plasmid for 20 h and then infected with SARS-CoV-2 (MOI = 1) or left uninfected for the indicated times before qPCR analysis was performed. g The M protein inhibits the SeV- and poly (I:C)-induced secretion of IFN-β and TNF-α in HEK293 cells. HEK293 cells stably expressing the M protein were infected with SeV (MOI = 1) or transfected with poly (I:C) for 8 h before measurement of IFN-β and TNF-α by ELISA. h The M protein impairs the SeV- and poly (I:C)-induced phosphorylation of downstream components. HEK293 cells stably expressing the M protein were infected with SeV (MOI = 1) or transfected with poly (I:C) for the indicated times before immunoblot analysis was performed. i M impairs the SeV-induced nuclear translocation of IRF3. HeLa cells were transfected with the Flag-M plasmid for 20 h and then infected with SeV (MOI = 1) or left uninfected for 10 h before confocal microscopy was performed. j Effects of the M protein on the IFN-β-induced transcription of the ISG56 and CXCL10 genes. HEK293 cells stably expressing the M protein were untreated or treated with IFN-β for 4 h before qPCR analysis was performed. k Effects of the M protein on cell viability. HEK293 cells stably expressing the M protein and vector were measured by MTT assays. Graphs show the mean ± SD; n = 3; ns not significant; *p < 0.05, **p < 0.01 (Student’s unpaired t test)