Extended Data Figure 10: Pyrin senses B. cenocepacia-induced Asn 41 deamidation of RHOA and induces caspase 1 inflammasome activation in the T6SS-dependent manner.

a–e, Wild-type, Nlrp3^−/−Nlrc4^−/−, Aim2^−/−, Asc^−/− and Mefv^−/− BMDMs were infected with B. cenocepacia J2315 strain (B.c., wild-type or the Δhcp mutant) or S. typhimurium (S.t.) for 3 h at a multiplicity of infection (m.o.i.) of 20:1. f–i, U937 or DC2.4 cells harbouring a vector or expressing human Pyrin (hPyrin) or mouse Pyrin isoform 1 (mPyrin-iso1) were infected with B. cenocepacia (B.c.) (wild type or the Δhcp mutant), S. flexneri (S.f.), EHEC, or stimulated with TcdB as indicated. j, Effects of overexpression of deamidated RHOA on ASC foci formation. The RHOA, B and C triple-knockdown 293T cells obtained in Fig. 4e were transiently transfected with an empty vector or a plasmid overexpressing RHOA wild type or the N41D mutant. Representative caspase 1 immunoblot from at least three repetitions are shown in a, b, e, f, h and i. ELISA assay of IL-1β release in c, d and g and percentages of cells showing the ASC foci in j are mean ± s.d. (n = 3). For j, the P value was determined by Student’s t-test.