AuthorHyman, Paul Lawrence.
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PublisherThe University of Arizona.
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AbstractRecombinational repair is a widespread mechanism for dealing with DNA damage. It is found in both prokaryotes and eukaryotes which implies that it is an ancient process which arose early in the evolutionary history of life on Earth. In addition, it has been implicated as a driving force in the evolution of sexual reproduction. In this dissertation I report experimental results which clarify the role of recombinational repair in bacteriophage (phage) T4. The Luria-Latarjet effect is an increase in resistance to DNA damage by phage T4 during infection. It has often been assumed to involve recombinational repair, but this has never been actually demonstrated. Using eleven phage T4 mutants, I have obtained evidence that the Luria-Latarjet effect is due to three repair pathways--excision repair, post-replication-recombinational-repair (PRRR) and multiplicity reactivation (MR), a form of recombinational repair. My results show that the Luria-Latarjet effect develops in two stages. The first stage starts soon after infection. Damage which occurs during the first stage can be repaired by excision repair or PRRR. The second stage appears to start after the first round of DNA replication is complete. Damage which occurs during this stage can apparently be repaired by MR as well as the other two repair pathways. I have also transferred the yeast RAD50 gene, which is required for recombinational repair, into an E. coli expression vector. After demonstrating expression of the protein, I used this construct to test for complementation by the RAD50 gene of E. coli and phage T4 mutants defective in recombinational repair. I was unable to demonstrate complementation in five different assays. Based on the results discussed above and what is known about the phage T4 life cycle, I propose a model for the Luria-Latarjet effect in phage T4. Further, I propose that recombinational repair has been selected to ensure progeny phage genomes are packaged with minimum damage. Since numerous other viruses also show a Luria-Latarjet effect type resistance to DNA damage, I suggest that the conclusions from this phage T4 study may have wide applicability to other viruses.
Degree ProgramMolecular and Cellular Biology