The present work investigated the mechanism of induced radiation resistance. Experiments were designed to identify the nature of lesions that induce this mechanism and the cellular components which regulated its expression in the yeast Saccharomyces cerevisiae and the green alga Chlamydomonas reinhardtii. DNA recombinational repair capacity induced by DNA damage was believed to be the system that conferred resistance to killing by radiation in yeast. The nature of the radiation generated DNA lesions that induce this mechanism have been examined using the two different endpoints of resistance to cell killing and suppression chemical mutation. In yeast, DNA lesions produced by low-LET (Linear Energy Transfer) $\gamma$-rays were more efficient at inducing radioresistance and mutation suppression than lesions produced by high-LET neutrons. It was inferred that DNA single strand breaks were important inducing lesions. Oxygen modifiable lesions and hydroxyl radicals (OH$\cdot)$ were particularly efficient inducers of this repair mechanism. Radiation induction of DNA repair depends not only on a variable induction response to different DNA lesions, but also on the availability of the induced repair system to deal with subsequent damage. Yeast mutants deficient in topoisomerase I, in topoisomerase II, or in DNA polymerase I were used to further investigate the involvement of these enzymes in the mechanism of induced thermal tolerance or radiation resistance. The systems that confer increased resistance to heat or radiation were independent of either topoisomerase activity or DNA polymerase function but topoisomerases may have a regulatory role during the signalling of these mechanisms. Maintenance of correct DNA topology may prevent induction of the heat shock response, and heat shock induction of a component of the full radiation resistance in yeast may be the consequence of topoisomerase I inactivation. DNA polymerase I did not seem to have a role in the thermal tolerance or radioresistance mechanism in yeast. Synchronized G$\sb1$ or G$\sb2$ C. reinhardtii cells were neither radiosensitized nor did they become radioresistant in response to heat shock. However, synchronized G$\sb1$ or G$\sb2$ algal cells had a normal heat-induced thermotolerance response. Neither radiation, heat shock, nor cycloheximide suppressed chemical mutations in algae. The DNA repair capacity of C. reinhardtii was not altered by heat shock and in this organism chemical mutations were not a consequence of an induced error-prone repair system. Heat shock was capable of inducing only partial radioresistance in wild type yeast cells while radiation induced full radioresistance. Dot blot analysis of the RAD52 epistasis group revealed that heat shock increased expression of only RAD52 transcripts whereas radiation increased expression of RAD52, 54, and 57 therefore demonstrating differential induction of repair genes by heat shock or radiation. Yeast mutants deficient in recombinational repair (rad52) were radiosensitive and neither heat shock nor radiation exposure could induce radioresistance. Nonetheless these mutants exhibited enhanced expression of RAD51, rad52, RAD54, 55, and 57 transcripts after either heat shock or radiation exposure. It was inferred that the inducible, non-functional rad52 gene product was incapable of conferring radioresistance (repairing DNA damage) and, as a consequence, additional repair genes may have subsequently been induced in an unsuccessful attempt to repair the residual damage. (Abstract shortened by UMI.)
| Identifer | oai:union.ndltd.org:uottawa.ca/oai:ruor.uottawa.ca:10393/5627 |
| Date | January 1990 |
| Creators | Boreham, Douglas R. |
| Publisher | University of Ottawa (Canada) |
| Source Sets | Université d’Ottawa |
| Detected Language | English |
| Type | Thesis |
| Format | 158 p. |
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