Cell nonautonomous activation of flavin-containing monooxygenase promotes longevity and health span

Abstract

Stabilization of the hypoxia-inducible factor 1 (HIF-1) increases life span and health span in nematodes through an unknown mechanism. We report that neuronal stabilization of HIF-1 mediates these effects in Caenorhabditis elegans through a cell nonautonomous signal to the intestine, which results in activation of the xenobiotic detoxification enzyme flavin-containing monooxygenase-2 (FMO-2). This prolongevity signal requires the serotonin biosynthetic enzyme TPH-1 in neurons and the serotonin receptor SER-7 in the intestine. Intestinal FMO-2 is also activated by dietary restriction (DR) and is necessary for DR-mediated life-span extension, which suggests that this enzyme represents a point of convergence for two distinct longevity pathways. FMOs are conserved in eukaryotes and induced by multiple life span-extending interventions in mice, which suggests that these enzymes may play a critical role in promoting health and longevity across phyla.

Fig. 1. A screen for age-associated autofluorescence identifies FMO-2 as a modulator of longevity and healthspan in the hypoxic response pathway

(A) Lifespans of vhl-1(ok161) animals on empty vector (EV), fmo-2 RNAi, or hif-1 RNAi. (B) Lifespans of Wild-Type worms and worms overexpressing FMO-2 ubiquitously (eft-3 promoter, FMO-2 OE). (C, D) Thrashing, pumping and autofluorescence measurements of Wild-Type, FMO-2 overexpression worms, and vhl-1(ok161) mutant worms during adulthood (day 10, 13 and 5, respectively). * indicates statistical difference (p<0.05) from Wild-Type by individual t-test for each strain. Error bars represent SEM, N ≥ 3 for all experiments.

Fig. 2. FMO-2 modulates proteostasis and longevity downstream of HIF-1 and DR

(A-C) Control, fmo-2(ok2147), vhl-1(ok161), and FMO-2 overexpression (FMO-2 OE) resistance to tunicamycin (growth from egg), dithiothreitol (survival at L4), and heat (survival at L4). (D) Wild-Type and fmo-2(ok2147) lifespans on dietary restriction (sDR). (E) fmo-2p::GFP reporter worms on fed and fasted conditions (F) Quantitative measurements of fmo-2 fluorescence shown in (E). * indicates statistical difference (p<0.05) from Wild-Type (** from hif-1) by individual t-test for each strain. Error bars represent SEM, N ≥ 3 for all experiments.

Fig. 3. Neuronal HIF-1 activates intestinal fmo-2 to increase longevity

(A) Lifespans of worms overexpressing FMO-2 under an intestinal (vha-6p) promoter. (B) Lifespans of control worms and worms expressing non-degradable HIF-1 (HIF-1^S) under neuronal (unc-14p) and ubiquitous (hif-1p) promoters. (C) Growth in hypoxia (0.5% oxygen) of wild-type, hif-1(ia04), and hif-1(ia04)::neuro-HIF-1^S worms after 6 days from egg. (D) Lifespans of control, hif-1(ia04), and hif-1(ia04) worms with stabilized neuronal HIF-1. (E) QPCR measurement of fmo-2 transcript in multiple strains. (F) Lifespans of hif-1(ia04) and hif-1(ia04) worms with stabilized neuro-HIF-1^S in control (EV) and fmo-2 RNAi. * indicates statistical difference (p<0.05) from Wild-Type (** from hif-1) by individual t-test for each strain. Error bars represent SEM, N ≥ 3 for all experiments.

Fig. 4. The signaling pathway from neuronal HIF-1 to intestinal FMO-2 involves serotonin and HLH-30

(A) Fluorescence images and quantification of fmo-2p::GFP reporter worms in normoxia (~21% O₂) and hypoxia (0.1% O₂) on control, tph-1, and ser-7 RNAi. (B, C) Lifespans of wild-type, vhl-1(ok161) mutant, and hif-1(ia04) with stabilized neuro-HIF-1^S worms on control (EV, solid lines), ser-7 RNAi (B, dashed lines) and tph-1 RNAi (C, dashed lines). (D) Lifespans of worms expressing HIF-1^S under the panneuronal (unc-54p) and serotonergic (tph-1p) promoters. (E) Expression of fmo-2p::GFP reporter in fed, fasted and hypoxic conditions under control and hlh-30 RNAi. (F) Lifespans of wild-type, vhl-1(ok161) mutant, and hif-1(ia04) with stabilized neuro-HIF-1^S worms on control (EV, solid lines) and hlh-30 RNAi (dashed lines) (G) Model of hypoxic response and dietary restriction converging on intestinal FMO-2. * indicates statistical difference (p<0.05) from Wild-Type by individual t-test for each strain. Error bars represent SEM, N ≥ 3 for all experiments.