Extended Data Fig. 9: Age-related hyperactivation of RAC by increased EPS-8 levels increases JNK phosphorylation.

a, Intestinal-specific knockdown of eps-8 after development does not affect lifespan (P = 0.1486). b, Epidermal-specific knockdown of eps-8 during adulthood does not affect lifespan (P = 0.3149). c, Single knockdown of RAC orthologues mig-2 (P < 0.0001) and rac-2 (P = 0.0008) during adulthood extends lifespan in wild-type worms. Knockdown of RAC orthologue ced-10 does not affect lifespan (P = 0.3227). d, Intestinal-specific knockdown of either mig-2 (P = 0.7340) or rac-2 (P = 0.6021) during adulthood does not affect lifespan. e, Muscle-specific knockdown of either mig-2 (P < 0.0001) or rac-2 (P = 0.0003) during adulthood extends lifespan. f, Neuronal-specific knockdown of either mig-2 (P < 0.0001) or rac-2 (P < 0.0001) during adulthood extends lifespan. g, Western blot analysis with antibodies to phosphorylated JNK (P-JNK), total JNK and α-tubulin of wild-type worms at different days of adulthood. Representative of three independent experiments. h, Western blot analysis with antibodies to P-JNK, total JNK and α-tubulin of wild-type worms at day 10 of adulthood. eps-8 RNAi was initiated during adulthood. Representative of two independent experiments. i, Knockdown of kgb-1 after development extends longevity (EPS-8 (WT) Vector RNAi versus EPS-8 (WT) kgb-1 RNAi, P < 0.0001) and rescues the short lifespan induced by ubiquitin-less EPS-8 mutant variant (EPS-8 (WT) Vector RNAi versus EPS-8 (Ub-less) Vector RNAi (P < 0.0001); EPS-8 (WT) kgb-1 RNAi versus EPS-8 (Ub-less) kgb-1 RNAi (P = 0.8061)). In each lifespan experiment, RNAi was initiated at day 1 of adulthood. P values were determined by two-sided log-rank test, n = 96 worms per condition. Supplementary Table 11 contains statistics and replicate data of independent lifespan experiments. For gel source data, see Supplementary Fig. 1.