Effects of atg RNAi on longevity are highly variable and condition dependent.

(A) The autophagy pathway, and genes tested in this study. (B-D) Effects at 20°C. (B) Effects on N2 (wild type). (C) Effects on daf-2(e1368). (D) Effects on daf-2(e1370). (E, F) Effects at 25°C. (E) Effects on N2. (F) Effects on daf-2(e1368). (G) Effects of bec-1 RNAi on daf-2(e1370) at 15°C. (B-G summed data, N = 2; for individual trial data and statistical comparisons, see Table S2, S5.

Degree of suppression of daf-2 longevity by atg gene RNAi differs greatly between daf-2 alleles and atg RNAi treatments (20°C), cf. Figure 1B-D (same data).

Summed data, N = 2; for individual trials and statistical comparisons, see Table S2. All trials were performed in parallel, i.e. all lifespans (summed or in each trial) are directly comparable.

FUDR but not infection alters outcome on atg gene RNAi (20°C).

(A-C) Effects of 0 μM, 15 μM and 800 μM FUDR. We note that in (A) the lifespan of the gfp RNAi control is somewhat lower than in other experiments (mean 14.96 days, Table S6); see Discussion for consideration of possible reasons for inter-trial variability. (D) No alteration by kanamycin of atg-13 RNAi effect on N2 lifespan. Summed data, N ≥ 2; for individual trials and statistical comparisons, see Table S6, S7.

Degree of suppression of glp-1(e2141) longevity by atg gene RNAi differs greatly between genes.

(A, B) 20°C. (A) Effects on N2. (B) Effects on glp-1. (C, D) 25°C. (C) Effects on N2. (D) Effects on glp-1. Summed data, N = 2-5; for individual trials and statistical comparisons, see Table S9. That bec-1 RNAi increases N2 lifespan in (A) (+16.7%, p < 0.0001) but not (B) could imply an interaction with temperature during development, or merely variability of atg RNAi effects (see Discussion). (E) Overview of effects of suppression of Age by RNAi of genes specifying autophagy. Dark blue, reduction of daf-2 or glp-1 Age that is significantly greater than in N2. Light blue, reduction of daf-2 or glp-1 Age that is not significantly greater than in N2. R, robust suppression, i.e. knockdown reduces the extended lifespan of daf-2 or glp-1 to within <30% of the mean lifespan of N2 under the same RNAi. This designation (“robust”) indicates a high degree of suppression of the mutant longevity phenotype (see Figure 2, Figure 5 and Fig. S2). Note that the bec-1 RNAi effect on glp-1 at 20°C is not classified as robust here even though it reduces lifespan to within <30% of the mean lifespan of N2 under bec-1 RNAi, since the fact that it does so partly reflects an increase in N2 lifespan, rather than a robust life-shortening effect on glp-1.

Degree of suppression of glp-1(e2141) longevity by atg gene RNAi differs greatly between genes (20°C), cf. Figure 4A,B (same data).

Summed data, N = 2-5; for individual trials and statistical comparisons, see Table S9.

Absence of effect of atg RNAi on vitellogenin accumulation.

Fold change of yolk proteins YP170, YP115 and YP88 in N2 and fog-2 normalized to gfp day 1 (N = 3). In no case is vitellogenin level significantly different to that in the gfp RNAi control at any time point (two-way ANOVA, Table S10). For raw data for vitellogenin levels see Supplementary Dataset 3.