Fig. 1

TRIM56 interacts with cGAS. a HEK293T cells were transiently transfected with cGAS-FLAG and protein complexes were purified from cell lysates with anti-FLAG M2 beads. The immunoprecipitates were eluted with FLAG peptides and fractionated using SDS-PAGE followed by Coommassie blue staining. The bands for TRIM56 and cGAS identified by mass spectrometry are indicated on the right. b Co-immunoprecipitation analysis of the interaction between cGAS-FLAG and TRIM56-V5. HEK293T cells expressing cGAS-FLAG were co-transfected with TRIM56-V5 or TRIM25-V5. cGAS-FLAG immunoprecipitates were probed with FLAG and V5 antibody and whole-cell lysates (WCLs) were probed with FLAG and V5 antibody, as indicated. c cGAS interacts with endogenously expressed TRIM56. HEK293T cells were transfected with cGAS-FLAG. Rabbit IgG or TRIM56 immunoprecipitates were probed with FLAG antibody, and WCLs were probed with TRIM56 and FLAG, as indicated. d cGAS-FLAG was transiently coexpressed with TRIM56-V5 in HEK293T cells. TRIM56-V5 was pulled down for SiMPull analysis. Top panel: schematic depiction of cGAS SiMPull; middle panel: representative GFP fluorescence images; bottom panel: average number of molecules per imaging area (N_f). e The N-terminal regulatory domain of cGAS interacts with the C-terminal NHL homologous region of TRIM56. Left panel: the regulatory domain of cGAS binds to TRIM56. WCLs of HEK293T cells transfected with TRIM56-V5 and cGAS truncation mutants were used for immunoprecipitation and immunoblotting with the indicated antibodies. Right panel: the C-terminus domain (308–531) of TRIM56 interacts with cGAS. WCLs of HEK293T cells transfected with cGAS-FLAG and TRIM56 truncation mutants were used for immunoprecipitation and immunoblotting with the indicated antibodies. Data are representative of two independent experiments a–e. Error bars indicate mean ± s.d. of n = 3. *P < 0.05 versus control using Student’s t-test (d). Full blots are shown in Supplementary Fig. 10