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. 2021 Mar 15;17(3):e1009401. doi: 10.1371/journal.ppat.1009401

Fig 5. Molecular characterization of cGAS neddylation.

Fig 5

(A) HEK293T cells were transfected with indicated plasmids. 48h after transfection, cells were denatured with 1% SDS, then cells were lysed and immunoprecipitation with anti-Flag antibody and then analyzed by immunoblotting with indicated antibodies. (B) HEK293T cells were transfected with indicated plasmids. 45h after transfection, one group of cells was treated with 5mM MLN4924 for 3h, then cells were subjected to denatured immunoprecipitation with anti-Flag antibody and then analyzed by immunoblotting with indicated antibodies. (C) HEK293T cells were transfected with HA-Nedd8, Flag-cGAS, myc-Rnf111 (WT), myc-Rnf111(C937A). 48h after transfection, cells were subjected to denatured immunoprecipitation with anti-Flag antibody and then analyzed by immunoblotting with indicated antibodies. (D) Recombinant cGAS was incubated with NEDD8, E1, UBE2M and RNF111 in the presence of ATP, after the reaction, proteins were subjected to SDS-PAGE, and immunoblotting with indicated antibodies. (E) MEFs were infected with HSV-1 (MOI = 0.3) for indicated times, then cell lysis was subjected to denatured immunoprecipitation with anti-NEDD8 antibody and then analyzed by immunoblotting with indicated antibodies.