Hormesis Extends the Correlation between Stress Resistance and Life Span in Long-lived Mutants of Caenorhabditis Elegans

James Cypser, M.A., and

Thomas E. Johnson, Ph.D.

Institute for Behavioral Genetics

University of Colorado

1480 30th Street

Boulder CO 80303-0447

Tel: 303 492 0279

Fax: 303 492 8063

Email: johnsont@colorado.edu

In general we agree with Dr. Rattan's thesis. Mutations that increase both life span and resistance to environmental stress have been documented in model systems as diverse as Caenorhabditis elegans, Drosophila melanogaster, and Mus musculus1. The correlation between life extension (Age) and increased stress resistance is well established in C. elegans carrying single-gene mutations2. Indeed, an examination of fifteen C. elegans gerontogene mutants indicates that all display increased resistance to at least one stressor such as heat, reactive oxygen species, or ultra violet light (S. de Castro and T. Johnson, unpublished results). In identifying gerontogenes, however, one must be careful to look only at genetic changes that result in life-prolongation, since shortening of life can result from many deleterious interventions. The stress-resistant, long-lived phenotype of these Age mutants comes at an evolutionary cost, because all known gerontogene mutants also display other "primary" phenotypes such as lower fertility or delayed development, which would likely result in selection (under non-laboratory conditions) in favor of the wild type.

Both animals subjected to hormetic treatments and gerontogene mutants share important characteristics: increased stress resistance, life extension, and (possibly) reduced fitness. The reduction of fitness caused by thermally-induced hormesis manifests as decreased fertility3 (Cypser, unpublished). Thus gerontogene mutants phenotypically mirror multiple characteristics of genetically wild-type animals made long-lived via environmental manipulation.

None of the dozen or so cloned gerontogene mutants in C. elegans, code for genes previously known to "constitute various maintenance and repair pathways" (Rattan, this issue). Instead, many Age genes clearly code for proteins involved in intracellular signaling events, perhaps in response to exogenous stress (Murakami and Johnson, submitted) and certainly in response to crowding and lack of food4. The Age genes seem to function as modulators of multiple stress response pathways rather than as enzymatic effectors of such repair functions.

Genetic separation of Life Span and Thermotolerance

The striking correlation between increased stress resistance and life extension provokes the question of whether life extension requires increased stress resistance. Free radicals generated as metabolic by-products are an important endogenous source of stress theorized to be a cause of aging5. Coupling this concept to that of the general adaptive response to stress6,7 might easily lead to the conclusion that those individuals best able to resist stress in general would also be those best able to resist the stress of free radicals. Genetic study of the nematode is useful for disentangling the relationship between stress resistance and aging. Although increased stress resistance may be necessary to extend life, Lithgow3 showed that thermal stress resistance alone was not sufficient, as the C. elegans mutants daf-4 and daf-7 were stress resistant but not long-lived. Further investigation using C. elegans as a model of the hormetic response has revealed a possible genetic separation of induced stress resistance from hormetic life extension.

The daf-16 gene is required for expression of both the life extension and stress resistance phenotypes of gerontogene mutants in the dauer formation pathway as well as the increased life expectancy of several other gerontogene mutants (clk-1, spe-26, unc-4, unc-34) not known to be involved in dauer formation8,9,10. Wild-type worms pretreated with heat on the first day of adulthood become more resistant to heat than untreated controls. Animals carrying any of several mutant alleles of daf-16 perform as well after pretreatment as do pretreated, wild-type animals (Cypser, unpublished results). Thus daf-16 does not appear to be necessary for the induction of thermotolerance. It was natural to ask next whether daf-16 is necessary for the life-extending effect of hormesis. Wild-type animals given the same treatment that induces thermotolerance live approximately 20% longer than wild-type controls if returned to the ideal temperature of 200C11 (Cypser unpublished results). However, animals homozygous for a mutation in daf-16 do not derive an increased life span in response to the same pretreatment; rather, their life expectancy is decreased below even the slightly reduced life span of untreated daf-16 controls (Cypser and Johnson, in preparation). In short, daf-16 is not required for the induction of thermotolerance, but is required for induction of life extension. Thus we have another case wherein increased stress resistance is not sufficient to extend life span.

The thermotolerance induced by hormesis fades quickly, returning within three days of pretreatment (on the first day of adulthood) to that of untreated controls in a sterile strain with a wild-type life span. Additionally, the worm loses the ability to respond to thermal hormetic conditions with subsequent thermotolerance about the time of reproductive cessation. (Cypser and Johnson in preparation).

Hormesis as the Shadow of Dauer Formation

We have tested the ability of other dauer-defective mutants to extend their life span in response to hormetic conditions (heat pretreatment). We have found that in addition to daf-16, both daf-18 and daf-12 mutants do not display life extension in response to hormesis; and in general live shorter as a consequence of the same treatment that extends the life span of wild type (Cypser and Johnson in preparation). However, another dauer-defective mutant, daf-3, has yielded unusual results in tests of its induced thermotolerance. daf-3 animals respond differently to different treatment conditions. The standard conditions of pretreatment, two hours at 350C, confer thermotolerance comparable to that induced in wild-type. However, pretreatment of 24 hours at 270C may produce an increase in thermotolerance above and beyond the increased thermotolerance observed in wild-type animals treated with the same conditions. We are now testing the hormetic life extension abilities of mutants that tend to form dauers at 270C to determine whether the represented genes play critical roles in aging via hormesis.

It should be possible to establish what genes are required for hormetic life extension. Mutants displaying either reduced or enhanced hormetic abilities could be used in conjunction with micro array technology to identify these genes. For example, if daf-3 is found to be super-hormetic for life span, then a comparison of the transcripts of pretreated daf-3 and daf-16 mutants may reveal a class of genes transcriptionally regulated in opposite directions. Genes found to be critical to hormetic life extension in the nematode may also influence human aging. However, hormesis likely represents a trade-off of total fitness for short-term survival of the individual. Thus practical application may be hindered if pleiotropic effects, seen in the worm as reduced fertility, appear as side effects in humans.


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