Fig. 5
From: Modulation of innate immune response to viruses including SARS-CoV-2 by progesterone
SRC mediates phosphorylation of IRF3 at Y107 and its activation. a Effects of progesterone on endogenous association of IRF3 with SRC. T-47D cells were treated with P4 (1 μM) for 1 h, and then left uninfected or infected with SeV for the indicated times before coimmunoprecipitation and immunoblotting analysis with the indicated antibodies. b Effects of SRC on tyrosine phosphorylation of IRF3. HEK293 cells were transfected with the indicated plasmids for 24 h before coimmunoprecipitation and immunoblotting analysis with the indicated antibodies. c SRC does not mediate IRF3Y107F phosphorylation. HEK293 cells were transfected with the indicated plasmids for 24 h before coimmunoprecipitation and immunoblotting analysis with the indicated antibodies. d SRC mediates phosphorylation of IRF3Y107. HEK293 cells were transfected with the indicated plasmids for 24 h. The cell lysates were denatured by 1% SDS, which were then diluted with regular lysis buffer till the final concentration of SDS reached to 0.1%. The lysates were then immunoprecipitated with a Flag antibody, and then the immunoprecipitation were analyzed by immunoblots with the indicated antibodies. e Effects of progesterone on virus-induced phosphorylation of IRF3Y107 in SRC-knockdown cells. The control and SRC-knockdown T-47D cells were treated with P4 (1 μM) for 1 h, and then left uninfected or infected with SeV for the indicated times before immunoblotting analysis with the indicated antibodies. f Effects of IRF3 and IRF3Y107F on SeV-induced activation of the IFN-β promoter. HEK293 cells were transfected with the indicated plasmids for 24 h, and then infected with SeV for 12 h before luciferase assays. Data shown are mean ± SD (n = 3). *P < 0.05; **P < 0.01; ***P < 0.001. g Effects of IRF3Y107 mutant on progesterone-potentiated activation of the IFN-β promoter. The PGR-overexpressed HEK293 cells were transfected with the indicated plasmids for 24 h, and then left treated or untreated with P4 (1 μM) for 1 h followed by SeV infection for 12 h before luciferase assays. Data shown are mean ± SD (n = 3). *P < 0.05; **P < 0.01; ***P < 0.001. h. Effects of IRF3 Y107 mutation on its association with VISA and TBK1 or self-association. HEK293 were transfected with the indicated plasmids for 24 h before coimmunoprecipitation and immunoblotting analysis with the indicated antibodies. i Effects of IRF3 Y107 mutation on its dimerization. HEK293 were transfected with the indicated plasmids for 24 h, and then left treated or untreated with P4 (1 μM) for 1 h followed by SeV infection for 6 h. The cells were lysed for native PAGE and immunoblotting analysis with the indicated antibodies
