A small-molecule Psora-4 acts as a caloric restriction mimetic to promote longevity in C. elegans

Abstract

In populations around the world, the fraction of humans aged 65 and above is increasing at an unprecedented rate. Aging is the main risk factor for the most important degenerative diseases and this demographic shift poses significant social, economic, and medical challenges. Pharmacological interventions directly targeting mechanisms of aging are an emerging strategy to delay or prevent age-dependent diseases. Successful application of this approach has the potential to yield dramatic health, social, and economic benefits. Psora-4 is an inhibitor of the voltage-gated potassium channel, Kv1.3, that has previously been shown to increase longevity and health span in the nematode Caenorhabditis elegans (C. elegans). Our recent discovery that Psora-4 lifespan benefits in C. elegans are synergistic with those of several other lifespan-extending drugs has motivated us to investigate further the mechanism by which Psora-4 extends lifespan. Here, we report that Psora-4 increases the production of free radicals and modulates genes related to stress response and that its effect intersects closely with the target set of caloric restriction (CR) genes, suggesting that it, in part, acts as CR mimetic. This effect may be related to the role of potassium channels in energy metabolism. Our discovery of a potassium channel blocker as a CR mimetic suggests a novel avenue for mimicking CR and extending a healthy lifespan.

Keywords: Aging; Caloric restriction; Drug; Lifespan; Potassium channel; Psora-4.

Fig. 1

Psora-4 increases healthy lifespan of wild-type N2 nematodes. a Mean lifespan in days of several repeats of different doses of Psora-4 (previous and new data combined). 100 μM showed the best lifespan extension (log-rank). b Health span category of worms. Worms under Psora-4 treatment experienced a significantly larger percentage (51%) of their already extended lifespan at optimal health compared to untreated controls (39.2%) (P-value < 0.01). 100 worms per condition. Log-rank test was used to determine statistically significant differences in health span by taking category “A” animals as “healthy” and combining category “B” and “C” animals as “compromised.” c Health span category of worms using the scoring scheme of Herndon et al. Worms under Psora-4 treatment showed better health span across all stages of lifespan. d Multiple sequence alignment of human Kv1.3 potassium channel with C. elegans homolog, using EMBL-EBI Clustal Omega [26]. (Table S4 for percent identity score). Shadowed boxes indicate SF and S6 amino acid sequence regions respectively. “*”means the same letter/amino acid in each; “:”means very similar sequence; “.”means similar sequence; red means strongly aligned; blue means not strongly aligned. One-way ANOVA, ***P < 0.001, *P < 0.05

Fig. 2

Psora-4 increases ROS production and improves stress resistance. a Psora-4 resulted in a significant increase in mitochondrial ROS production (mean ± SD). NAC completely blocks the increase in ROS in worms treated with Psora-4 but does not affect endogenous ROS levels in untreated animals. 150 worms per well, 8 replicates. b Psora-4-treated worms exhibit improvement in stress resistance as measured by survival time on solid NGM plates containing 20 mM paraquat. 50 worms per condition. Log-rank test, P < 0.05. c The exogenous antioxidant/redox modulator NAC alone does not extend or decrease the lifespan of nematodes. Lifespan extension by Psora-4 is not affected by co-treatment with NAC. Lifespan extension in the presence and absence of NAC is unchanged (statistically insignificant, log-rank test). d Mean lifespan of nematode under psora-4 and NAC treatment. 100 worms per condition. One-way ANOVA, ***P < 0.0001

Fig. 3

Transcriptome profile of Psora-4 confirmed that nematodes treated with psora-4 have enhanced stress response. a GO term analysis, top GO terms targeted by Psora-4 were defensive responses, transmembrane transport, and oxidation-reduction process, based on DEGs with |LFC| > 1 and P-value < 0.05. b As in a only for downregulated genes. Top hits are metabolic processes. c As in a for upregulated genes. Top hits are oxidation-reduction processes. d Top signaling pathways targeted by Psora-4 differentially expressed genes based on panther pathway analysis. e Panther GO-slim molecular function showed the top classes of differentially expressed genes by Psora-4 are antioxidant activities. f GO terms enriched by DEGs of Psora-4 were analyzed by DAVID [32] and summarized by REVIGO [33]. The top GO terms are metabolic processes. X- and Y-axes represent GO terms’ semantic similarity measures in multidirectional scaling; the closeness of the groups on the plot reflects their relative similarity based on this metric

Fig. 3

Transcriptome profile of Psora-4 confirmed that nematodes treated with psora-4 have enhanced stress response. a GO term analysis, top GO terms targeted by Psora-4 were defensive responses, transmembrane transport, and oxidation-reduction process, based on DEGs with |LFC| > 1 and P-value < 0.05. b As in a only for downregulated genes. Top hits are metabolic processes. c As in a for upregulated genes. Top hits are oxidation-reduction processes. d Top signaling pathways targeted by Psora-4 differentially expressed genes based on panther pathway analysis. e Panther GO-slim molecular function showed the top classes of differentially expressed genes by Psora-4 are antioxidant activities. f GO terms enriched by DEGs of Psora-4 were analyzed by DAVID [32] and summarized by REVIGO [33]. The top GO terms are metabolic processes. X- and Y-axes represent GO terms’ semantic similarity measures in multidirectional scaling; the closeness of the groups on the plot reflects their relative similarity based on this metric

Fig. 4.

Comparison of Psora-4 with eat-2 and effect on the lifespan of worms fed on different concentrations of bacteria. a A Venn diagram of differentially expressed genes. 193 genes were affected by both Psora-4 and in eat-2 mutants. Among these genes impacted by both Psora-4 and eat-2, 26 were known aging genes (as determined by having entries in the GenAge database). Overall, Psora-4 impacted 43 GenAge genes; therefore, 59% of GenAge genes (26 out of 43) affected by Psora-4 were also affected in eat-2 mutants. b Circos plot (generated by Metascape) shows gene expression overlaps for Psora-4 and eat-2. The outside arc represents genes that are up and downregulated by eat-2 or Psora-4. On the inside arc, dark orange represents genes affected by both eat-2 and Psora-4 and light orange represents genes unique to either conditions. Purple lines link genes commonly affected by eat-2 and Psora-4. c Lifespan of untreated worms fed on different concentrations of OP50-1. d Psora-4 extends the lifespan of worms fed on a normal diet (10¹⁰ CFU/ml). e Psora-4 results in only a small lifespan extension effect in worms fed on 10⁹ CFU/ml OP50-1 (P-value < 0.05). f Psora-4 was unable to further extend the lifespan of worms fed on 10⁸ CFU/ml (CR). g Direct comparison of Psora-4 lifespan in worms fed on different concentrations of OP50-1; lifespan of Psora-4-treated animals is similar on all food concentrations and indistinguishable from lifespan at highest level of CR (lowest food concentration)

Fig. 4.

Comparison of Psora-4 with eat-2 and effect on the lifespan of worms fed on different concentrations of bacteria. a A Venn diagram of differentially expressed genes. 193 genes were affected by both Psora-4 and in eat-2 mutants. Among these genes impacted by both Psora-4 and eat-2, 26 were known aging genes (as determined by having entries in the GenAge database). Overall, Psora-4 impacted 43 GenAge genes; therefore, 59% of GenAge genes (26 out of 43) affected by Psora-4 were also affected in eat-2 mutants. b Circos plot (generated by Metascape) shows gene expression overlaps for Psora-4 and eat-2. The outside arc represents genes that are up and downregulated by eat-2 or Psora-4. On the inside arc, dark orange represents genes affected by both eat-2 and Psora-4 and light orange represents genes unique to either conditions. Purple lines link genes commonly affected by eat-2 and Psora-4. c Lifespan of untreated worms fed on different concentrations of OP50-1. d Psora-4 extends the lifespan of worms fed on a normal diet (10¹⁰ CFU/ml). e Psora-4 results in only a small lifespan extension effect in worms fed on 10⁹ CFU/ml OP50-1 (P-value < 0.05). f Psora-4 was unable to further extend the lifespan of worms fed on 10⁸ CFU/ml (CR). g Direct comparison of Psora-4 lifespan in worms fed on different concentrations of OP50-1; lifespan of Psora-4-treated animals is similar on all food concentrations and indistinguishable from lifespan at highest level of CR (lowest food concentration)

Fig. 5

Psora-4 decreases accumulation of long-chain fatty acyl–containing TAGs. a Heatmap of TAG species. Based on TAG abundance Psora-4-treated N2 (N2 + Psora-4) and untreated eat-2 (eat-2) were separated from untreated N2 (N2) and clustered together. b Two days of Psora-4-treated N2 worms and eat-2 mutants has lower total TAG accumulation than wild-type N2 nematodes. c Both Psora-4 treatment and eat-2 did not affect the abundance of medium-chain fatty acyl–containing TAGs. d The decrease in total TAGs is due to a decrease in long-chained fatty acyl–containing TAGs. e DEGs of Psora-4 treatment and eat-2 mutant that are downstream of daf-16 but not daf-2. Highlighted in purple are genes downregulated by Psora-4 and eat-2 in a similar direction. *** P-value < 0.001, ** P-value < 0.01