Worm Breeder's Gazette 15(1): 38 (October 1, 1997)
These abstracts should not be cited in bibliographies. Material contained herein should be treated as personal communication and should be cited as such only with the consent of the author.
Department of Biology/Toxicology, Ashland University, Ashland, Ohio 44805
Induction of heat shock proteins in response to thermal stress is a well-characterized phenomenon reported to occur in many organisms including C. elegans (Heschl and Baillie, DNA 8: 233-43, 1989). Other stressors, including chemical stressors, have also been shown to induce these proteins (D. Jones et al., Toxicology 109: 119-27, 1996). We have hypothesized that chemicals affecting energy metabolism will also induce the heat shock response. As a test compound, we chose sodium azide, an inhibitor of oxidative phosphorylation, that is also used as a nematode anesthetic. We have found that exposure to low levels of this chemical confers thermotolerance in the nematode. To test our hypothesis, we developed the following protocol: the wild type (N2) strain received a 1 hr pre-treatment with either azide or 33oC. After a 4 hr recovery at 22oC, nematodes were heat shocked at 37oC for 2 hrs. After heat shock, worms recovered again at 22oC for 4 hrs before the number of surviving worms was determined. A 4 hr recovery period was necessary because worms are not moving immediately after the 37oC heat shock. Additionally, we have observed that temperature fluctuation as little as 1.5oC can critically affect the outcome. For example, temperatures rising to 38.5oC for 4 minutes prove lethal to all worms. Azide concentrations of 5.0-20mM were chosen based on preliminary studies which indicated that concentrations >20mM were lethal. The control worms (receiving only the heat shock) had a 0.08 survival probability while animals receiving the 33oC pre-treatment had a 0.89 survival probability, illustrating the classic heat shock response. At all concentrations, the sodium azide pre-treated worms showed an increased survival probability when compared to the control. 10mM (0.07%) sodium azide, corresponding to the concentration typically used as an anesthetic, conferred maximal (0.70) survival probability. We are currently using Western Blots to test our hypothesis that the molecular mechanism of the response to azide is the same as the response to elevated temperatures. The hsps we are examining include hsp90, 70, 60, and 16 (hsp16 antibody kindly provided by Peter Candido). We have also been searching for hsp mutants. Elizabeth Malone and Jim Thomas reported at the worm meeting that daf-21 is really a hsp90 mutation. We are currently testing their temperature sensitive mutant (JT6130). Grown at the permissive temperature of 16oC, JT6130 survived heat shock after receiving pre-treatment with either azide or 33oC. However, these worms showed a reduction in survival probability when compared to N2. At the restricted temperature of 25oC, the worms pre-treated with either azide or 33oC did not survive the heat shock. These results strongly suggest that hsp90 is an important part of the worm's response to stress. We are searching ACEDB, as well as the literature, for information on mutants with defects in some aspect of energy metabolism. Like daf-21 and hsp90, we suspect that mutations in energy metabolism proteins may have already been identified by other names; therefore, we welcome any additional information concerning mutants that are thought to be involved in energy metabolism.