Fig. 2: Age-related deubiquitination impairs targeted degradation of longevity regulators.
From: Rewiring of the ubiquitinated proteome determines ageing in C. elegans
a, Ten Ub-proteins increase after rpn-6 RNAi treatment in young wild-type adults (rpn-6 RNAi/Vector RNAi) and become less ubiquitinated but more abundant with age (day 15/day 5). b, Volcano plot of proteins containing Lys48-linked polyUb in day-5 adult wild-type worms (n = 3, FDR < 0.05). The −log10(P value) of a two-sided t-test is plotted against the log2-transformed fold change values from immunoprecipitation (IP) with an antibody against Lys48-linked polyUb compared with a control anti-Flag antibody. Red dots indicate age-dysregulated proteasome targets. c, d, Knockdown of either ifb-2 (c) or eps-8 (d) after development extends lifespan (P < 0.0001). e, Western blot analysis with an antibody against IFB-2 of wild-type worms at different days of adulthood. α-tubulin is the loading control. Representative of four independent experiments. f, Western blot analysis with an antibody against EPS-8 of wild-type worms. Representative of three independent experiments. g, Western blot analysis with an antibody against IFB-2 of wild-type and IFB-2(K255R/K341R) (Ub-less) mutant worms at day 2 of adulthood. Representative of three independent experiments. h, Ubiquitin-less IFB-2 mutant worms have a shorter lifespan than wild-type worms (P < 0.0001). i, Western blot analysis with an antibody against EPS-8 of worms expressing endogenous wild-type EPS-8::HA or EPS-8(K524R/K583R/K621R::HA) (Ub-less) at day 1 of adulthood. Representative of three independent experiments. j, EPS-8 (Ub-less) mutants are short-lived (P < 0.0001). In lifespan experiments, P values were determined by two-sided log-rank test; n = 96 worms per condition. Lifespan data are representative of at least two independent experiments. Supplementary Table 11 contains replicate data of independent lifespan experiments. For gel source data, see Supplementary Fig. 1.
