Extended Data Figure 8: Binding and ubiquitination of different GBP-family members by IpaH9.8.
From: Ubiquitination and degradation of GBPs by a Shigella effector to suppress host defence
a, b, 293T cells were co-transfected with Flag–IpaH9.8-C337A and a haemagglutinin-tagged human (a) or mouse (b) GBP-family member. c, In vitro ubiquitination of Flag-tagged representative GBP-family members by IpaH9.8. d, The haemagglutinin-tagged GTPase domain (N) or helical domain (C) of the indicated GBP proteins was co-transfected with Flag–IpaH9.8-C337A or Flag–RFP control into 293T cells. e, Haemagglutinin-tagged mGBP1, mGBP2, or the indicated chimaera construct, were co-transfected with 6×Myc–IpaH9.8 (wild-type or C337A mutant (C/A)) or a vector control (−) into 293T cells. 1N, GTPase domain of mGBP1; 2N, GTPase domain of mGBP2; 1C, helical domain of mGBP1; 2C, helical domain of mGBP2. Cell lysates (a, b, d) or reaction mixtures (c) were immunoprecipitated with Flag antibody (Flag IP) and immunoblotted with Flag, haemagglutinin, tubulin or myc antibodies. Coomassie blue-stained SDS–PAGE gels (c) show the recombinant proteins added into the reaction. All data shown are representative of two independent experiments.
