Fig. 6: Enzymatic activity of USP18 is dispensable for its function in RLR pathway. | Nature Communications

Fig. 6: Enzymatic activity of USP18 is dispensable for its function in RLR pathway.

From: USP18 positively regulates innate antiviral immunity by promoting K63-linked polyubiquitination of MAVS

Fig. 6

a Top panel: schematic diagram of USP18 and USP18 with C64S mutation (USP18C64S). Bottom panel: DNA sequencing result of plasmids expressing Myc-USP18 or USP18C64S. b Co-IP analysis of the interaction between MAVS and USP18 or USP18C64S in HEK293T cells transfected with the plasmids expressing Flag-MAVS, with Myc vector, Myc-USP18 or Myc-USP18C64S utilizing either anti-Flag magnetic beads (top panel) or anti-Myc magnetic beads (bottom panel). c Immunoprecipitation analysis of ubiquitination of MAVS in HEK293T cells transfected with the plasmids expressing Flag-MAVS, HA-wild type or HA-K63 only Ub, and Myc vector, Myc-USP18, or Myc-USP18C64S. d SDD-AGE (top) and SDS-PAGE (bottom) analysis of MAVS aggregation in HEK293T cells transfected with the plasmids expressing Flag-MAVS, with Myc vector, Myc-USP18, or Myc-USP18C64S. e Representative confocal images of immunofluorescence staining for HeLa cells transfected with the plasmids expressing Flag-tagged MAVS, DsRED2-Mito together with Myc vector, Myc-USP18, or Myc-USP18C64S, scale bar, 10 µm. f Immunoblot analysis of SeV and pIRF3 in lysates of HEK293T cells transfected with Myc-USP18 (top) and Myc-USP18C64S (bottom) in different dosages (1 µg, 2 µg, and 3 µg) for 12 h followed by infection with SeV for 8 h.

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