Extended Data Fig. 1: m-3M3FBS activates the inflammasome independent of NLRP3.
From: Mitochondrial damage activates the NLRP10 inflammasome
(a) IL-1β secretion from WT iMacs, LPS-primed (200 ng/ml) and stimulated with nigericin (10 μM), poly-(dA:dT) (2 μg/ml) or m-3M3FBS (80 μM) (n = 3). (b) ASC specks per nuclei in iMacs expressing NLRP3 and ASCmCerulean, stimulated as in (a) (n = 3). (c) IL-1β secretion from WT iMacs, LPS-primed (200 ng/ml) and stimulated with nigericin (10 μM), poly-(dA:dT) (2 μg/ml) or m-3M3FBS (85 μM), in the presence of increasing doses of KCl (0, 5, 10, 25, 50, 75, 100, 125 mM) (n = 3). (d) ASC specks per nuclei in iMacs expressing NLRP3 and ASCmCerulean, stimulated as in (c), in the presence of increasing doses of KCl (0, 5, 10, 25, 50, 75, 100, 125 mM) (n = 3). (e) ASC specks per nuclei in WT iMacs expressing ASCmCerulean, with or without LPS priming (200 ng/ml) (41, 42), stimulated as in (c). While nigericin-induced ASC speck formation depends on priming, m-3M3FBS and poly-(dA:dT) trigger ASC speck formation in both LPS-primed and -unprimed cells (n = 3). (f) IL-1β secretion from WT iMacs, LPS-primed (200 ng/ml) and stimulated as in (c), in the presence or absence of the NLRP3 inhibitor CRID3 (5 μM) (43-45) (n = 3). (g) ASC specks per nuclei in iMacs expressing NLRP3 and ASCmCerulean, stimulated as in (c), in the presence or absence of the NLRP3 inhibitor CRID3 (5 μM) (n = 3). P values were calculated by 2-way ANOVA with Šídák’s multiple comparisons test. **P = 0.0068, ****P < 0.0001, ns P = 0.5730-0.9999. (h) ASC speck formation in HEK cells stably expressing ASCTagBFP and transfected with an empty vector (EV) or human NLRC4, Pyrin or AIM2, stimulated with increasing doses of m-3M3FBS (40, 55, 70, or 85 μM); normalized to the unstimulated control (n = 3). (i) ASC specks per nuclei in HEK cells expressing human NLRP10 or mCitrine and ASCTagBFP, stimulated with nigericin (10 μM), poly-(dA:dT) (2 μg/ml) or m-3M3FBS (85 μM) (n = 4). Individual data points are means of technical triplicates (a, b, e, h, i) or duplicates (c, d, f, g); error bars represent SD.
