Figure 3.

C. elegans REST orthologs mediate longevity in daf-2 loss-of-function mutants. a, The REST orthologs spr-4 and spr-3 are required for maximal longevity in daf-2 mutant worms. Lifespan analysis was performed on wild-type and daf-2(1370) loss-of-function mutant worms, and the indicated combinations of daf-2 and spr-4/spr-3 mutations. The spr-4/spr-3 mutations significantly reduced the lifespan of daf-2 mutant worms. n=29-59 worms per genotype, replicated at least three times per genotype. P<0.001 for all curves relative to daf-2, by log-rank test. b, Neuronal expression of spr-3 and spr-4 mediate lifespan extension in daf-2 mutant worms. Shown are lifespans of worms with neuronal targeting of RNAi by neuronal expression of a sid-1 transgene in otherwise sid-1 null daf-2(1370) mutants, or untargeted RNAi in sid-1 wild-type daf-2(1370) mutants. Lifespan effect of neuronal targeting of spr-4;3 RNAi versus EV control RNAi is significant by log rank test (P=2.5e-6), n=22-56 worms per curve replicated at least 4 times. c, SPR-3 and SPR-4 repress genes that mediate neural excitation. Shown are significantly enriched GO terms for upregulated genes related to neural excitation in RNA-seq analysis of day 2 spr-4;3;daf-2 triple mutants versus daf-2 single mutant worms. P-values were calculated using Fisher’s exact test (see Methods), n=3 biological replicates per genotype.