Extended Data Fig. 3: CTH binds bacterial lipoproteins.
From: Cytosolic CTH senses bacterial lipoproteins and drives noncanonical inflammasome activation
a, Immunoblots of BMDMs from wild-type mice primed with 1 μg/ml Pam3CSK4 overnight before transfected with DOTAP alone or together with 200 ng/ml LPS or 10 μg/ml Pam3CSK4, or transfected with cell penetrating peptides alone or together with SA-Lpp (100 ng/μl) for 16 h. Sup., supernatant; SA-Lpp, S. aureus lipoprotein. b, Immunoblots of BMDMs from Cth+/+ and Cth−/− mice primed with 1 μg/ml Pam3CSK4 overnight before transfected with 200 ng/ml LPS for 16 h. Otherwise, cells primed with 1 μg/ml LPS for 3 h then stimulated with 2 mM ATP for 30 min or 2 μg/ml poly(dA:dT) for 4 h. Sup., supernatant. c, RT-PCR analysis of indicated gene in BMDMs from Cth+/+ and Cth−/− mice primed with 1 μg/ml Pam3CSK4 overnight. d, Immunoblots of BMDMs from Tlr2flox/flox (Tlr2+/+) and Tlr2flox/flox; Lyz2-Cre (Tlr2−/−) mice primed with 10 μg/ml poly(I:C) overnight before transfected with DOTAP together with 10 μg/ml Pam3CSK4 for 16 h. Sup., supernatant. e, Quantification of pyroptosis in BMDMs from Tlr2+/+ and Tlr2−/− mice treated as in d. ELISA assays of IL-1β in supernatants, LDH release assays of cell death in supernatants and ATP assays of cell viability in cell pellets. In c and e, data are pooled from three independent experiments (n = 3) and shown as means ± SD. Unpaired two-tailed Student’s t-test was used. Exact P values are indicated in the figures. Data are representative of three independent experiments with similar results in a-e.
