Cell-nonautonomous effects of dFOXO/DAF-16 in aging

Abstract

Drosophila melanogaster and Caenorhabditis elegans each carry a single representative of the Forkhead box O (FoxO) family of transcription factors, dFOXO and DAF-16, respectively. Both are required for lifespan extension by reduced insulin/Igf signaling, and their activation in key tissues can extend lifespan. Aging of these tissues may limit lifespan. Alternatively, FoxOs may promote longevity cell nonautonomously by signaling to themselves (FoxO to FoxO) or other factors (FoxO to other) in distal tissues. Here, we show that activation of dFOXO and DAF-16 in the gut/fat body does not require dfoxo/daf-16 elsewhere to extend lifespan. Rather, in Drosophila, activation of dFOXO in the gut/fat body or in neuroendocrine cells acts on other organs to promote healthy aging by signaling to other, as-yet-unidentified factors. Whereas FoxO-to-FoxO signaling appears to be required for metabolic homeostasis, our results pinpoint FoxO-to-other signaling as an important mechanism through which localized FoxO activity ameliorates aging.

Graphical abstract

Figure 1

dfoxo-to-dfoxo Signaling Is Not Required for the Antiaging Effects of Increased dFOXO Activity in the Gut/Fat Body or mNSC (A) Western blots of dFOXO and the tubulin loading control on whole-fly protein extracts from S₁106>dfoxo or dfoxoΔ/Δ S₁106>dfoxo female flies in the absence of the inducer. (B) dfoxo transcript levels (relative to Act and with S₁106>dfoxo -RU486 set to 1) in S₁106>dfoxo or dfoxoΔ/Δ S₁106>dfoxo female flies fed or not RU486. (C) Survival of S₁106>dfoxo or dfoxoΔ/Δ S₁106>dfoxo female flies in presence or absence of RU486 determined in two experimental trials (top and bottom panel). (D) Survival of dilp2GAL4>dfoxo female flies, or the two genetic controls (dilp2-GAL4 or UAS-dfoxo alone), in wild-type (WT) or dfoxoΔ/Δ backgrounds. (E) The proportion of high climbers (top panel) or combined medium and high climbers (bottom panel) in three cohorts (combined) of S₁106>dfoxo or dfoxoΔ/Δ S₁106>dfoxo female flies in the presence or absence of RU486. Note the same color code is used in (B), (C), and (E) and is given in (C). See Table 1 for statistical analysis of data in (B)–(E). Where used, box plots indicate median, first and third quartile, data range, and outliers.See also Figure S1.

Figure 2

Transcriptional Regulation by Gut/Fat Body dfoxo and the Physiological Relevance of dfoxo-to-dfoxo Signaling (A) Log₂ fold change in transcript levels upon RU486 administration in S₁106>dfoxo (x axis) or dfoxoΔ/Δ S₁106>dfoxo (y axis) female flies for the genes significantly changed in S₁106>dfoxo. Red indicates the genes whose response to RU486 is significantly altered by genotype (significant genotype:RU486 interaction in the linear model). The red line is the regression line for these genes (slope = 0.49) and the black line is for the others (slope = 0.77). The significant difference in slope (p = 1 × 10⁻⁷) indicates the genes marked in red are overall less responsive to RU486 in dfoxoΔ/Δ S₁106>dfoxo than in S₁106>dfoxo female flies. Gene lists are given in Table S1. (B) Nplp4 transcript levels (relative to Act and with body -RU486 set to 1) in bodies or heads of S₁106>dfoxo female flies fed or not RU486. (C) dfoxo transcript levels (relative to Act and with body -RU486 set to 1) in bodies or heads of S₁106>dfoxo female flies fed or not RU486. (D) Protein content of individual S₁106>dfoxo or dfoxoΔ/Δ S₁106>dfoxo female flies after 5-day feeding with RU486 or not. (E) Individual fly weight for the same conditions. Note the same color code is used in (B)–(E) and is given in (B). In (D) and (E), asterisk indicates significant difference at p < 0.05 by post hoc, pair-wise t test between − and + RU486 conditions. See Table 1 for statistical analysis of data in (B)–(E). See Figure S2 for further data.

Figure 3

daf-16-to-daf-16 Signaling Is Not Essential for the Lifespan Benefit of Increased DAF-16 Activity in the Intestine (A) Survival of wild-type (WT) and daf-16(mu86) worms in combination with daf-16 overexpression from a gut-specific promoter (muEx211[ges-1p::GFP::daf-16]) determined in two experimental trials (top and bottom panel) in worms fed HT115 bacteria. (B) Worm protein content under the four conditions described in (A). Color codes are the same in (A) and (B). See Figure S3A for further lifespan trials and Table 1 for statistical analysis. See also Figures S3B and S3D for the effect of an independent transgene. See Figure S3C for the lifespan effects on OP50 bacteria.