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. 2025 Mar 31;99(5):e00004-25. doi: 10.1128/jvi.00004-25

Fig 3.

Blots depict TRIM22 interactions with MAVS, IRF3, and TBK1. Graphs depict MAVS mRNA expression. Immunoprecipitation confirms TRIM22-MAVS binding. Immunofluorescence depicts TRIM22 and MAVS colocalization. Diagrams depict TRIM22 and MAVS domain structures.

The PRY/SPRY domain of TRIM22 interacts with the proline-rich domain of MAVS. (A) Western blot analysis using anti-Flag or anti-His antibodies for proteins that co-immunoprecipitated with Flag-tagged TRIM22 from lysates of HEK293T cells that were also co-transfected with plasmids encoding His-tagged IRF3, TBK1, or MAVS. Input represents western blot analysis of whole-cell lysates without IP. (B) RT-qPCR analysis of MAVS mRNA in TRIM22 knockdown or sh-NC A549 cells. (C) RT-qPCR analysis of MAVS mRNA in TRIM22 overexpression or vector A549 cells. (D) Co-immunoprecipitation analysis to assess the interaction of endogenous TRIM22 with MAVS in A549 cells. (E and F) Co-immunoprecipitation analysis to assess the interaction of endogenous TRIM22 with MAVS in A549 cells infected with IAV-H1N1 or IAV-H3N2 (MOI = 0.1) for 24 h. Input represents western blot analysis of whole-cell lysates without IP. (G) RT-qPCR analysis of MAVS mRNA in A549 cells infected with IAV (MOI = 0.1) for 24 h. (H) Fluorescence microscopy of A549 cells transfected for 48 h with plasmids expressing Flag-TRIM22 and His-MAVS, followed by labeling of TRIM22 and MAVS with a Flag- or His-specific primary antibody and an Alexa Fluor 568-conjugated goat anti-mouse IgG secondary antibody (red) and an Alexa Fluor 488-conjugated goat anti-rabbit IgG secondary antibody (green). Scale bars: 20 µm. (I) Binding of Flag-TRIM22 or its truncation mutants with His-MAVS in co-transfected HEK293T cells, as determined by co-immunoprecipitation and western blot analysis. Flag-TRIM22 domains and truncation mutants are shown on top. (J) Binding of His-MAVS or its truncation mutants with Flag-TRIM22 in co-transfected HEK293T cells, as determined by co-immunoprecipitation and western blot analysis. His-MAVS domains and truncation mutants are shown on top. A one-way ANOVA test was used for panels B, C, and F. All data are representative of three independent experiments with three biological replicates and are presented as mean ± S.D. (ns, nonsignificant; *P < 0.05; ***P < 0.001; ****P < 0.0001).