Figure 4. HIF-1 mediates lifespan extension of maa-1–deficient animals. (A) Downregulation of hif-1 reduces the longevity of maa-1(ok2033) mutants. Lifespans of wild-type and maa-1(ok2033) mutants subjected to hif-1 or control RNAi (P=0.1091 for control RNAi vs hif-1 RNAi of maa-1(ok2033) mutants). (B) Deletion of hif-1 reverses the lifespan extension conferred by the maa-1 mutation. (P=0.2661 for wild-type vs maa-1(ok2033); hif-1(ia4) mutants; P<0.0001 for maa-1(ok2033) vs maa-1(ok2033); hif-1(ia4) mutants). (C) qPCR analysis of the HIF-1 targets nhr-57 and F22B5.4 in maa-1(ok2033) and maa-1(ok2033); hif-1(ia4) mutants. Results are relative to levels in wildtype animals. Error bars represent SEM (t-test: *P<0.05, **P<0.001 for maa-1(ok2033) vs wildtype animals). (D) HIF-1 stability is not affected by downregulation of maa-1. Western blot of protein extracts from wildtype, transgenic iaIs34 animals carrying HIF-1 (P621G)::myc, and transgenic iaIs28 animals carrying HIF-1::myc, subjected to control or maa-1 RNAi. Blots were probed with anti-myc and anti-α-tubulin antibodies (upper panel). Quantification of band intensity is shown in lower panel (N=3, **P˂0.001). (E) A maa-1 loss-of-function mutation does not further increase the lifespan of long-lived vhl-1(ok161) mutants (P=0.0692 for vhl-1(ok161) vs maa-1(ok2033); vhl-1(ok161), P=0.8449 for maa-1(ok2033) vs maa-1(ok2033); vhl-1(ok161)). (F) Downregulation of maa-1 does not affect the lifespan of long-lived transgenic animals overexpressing HIF-1(P621G)::myc (hif-1 OE). (P=0.0678 for hif-1 OE on control vs maa-1 RNAi). P values were calculated using the log-rank (Mantel-Cox) method. Replicate experiments are shown in Table S1. Additional statistical analysis is shown in Table S2.