C. elegans has been critical in identifying mediators of programmed cell death. However, to our knowledge, hypoxic death has not been studied in C. elegans despite the fact that heart attacks and strokes, the results of cellular hypoxia, cause the majority of human deaths. Towards the goal of developing a forward genetic model of hypoxic death, we have screened through 71,438 mutagenized genomes and 74 existing mutant strains and found several mutants resistant to hypoxic death (Hyp - hyp oxic death abnormal). The most Hyp strains were found among previously isolated mutants of the
daf-2 gene, which codes for an insulin/insulin growth factor receptor homolog. The hypoxia resistance was profound; for example, 5.9 +/- 2 % of
daf-2(
e1370) were dead versus 92.8 +/- 2% of N2 after a 24 hr recovery from hypoxic incubation. With shorter hypoxic times, all of the N2 live but have permanent behavioral defects, suggestive of neuronal death.
e1370 behaves normally even after incubation times that kill all N2. The
e1370 animals are paralyzed while in the hypoxia chamber demonstrating that they were indeed hypoxic, and gaseous anesthetics affect
e1370 with the same time course as N2 indicating the cuticle does not exclude gas exchange.
daf-2(rf) 's Hyp phenotype was recessive, markedly allele specific, and did not correlate with
daf-2 's other phenotypes including Age, Daf-c, and thermal tolerance. Sequencing of all of the strong Hyp alleles showed that like
e1370 the 3 other strong Hyp alleles carried missense mutations in the tyrosine kinase domain of
daf-2 whereas mutations in the ligand-binding domain produced little or no Hyp phenotype. Utilizing
daf-2(
e1370) strains transformed with
daf-2(+) driven by tissue specific promoters (thanks to Catherine Wolkow - Ruvkun lab), we found partial rescue of hypoxia resistance by
daf-2 (+) expression in neurons or muscle but not in intestinal cells. The Hyp phene of
e1370 is completely suppressed by
daf-16(null) and
daf-18(null) , but not
daf-12(null) .
age-1(null) and
pdk-1(null) are only weakly Hyp, suggesting different pathways mediate the Hyp and Age phenes of
daf-2(rf). Ced mutants are not Hyp.