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Much research is being conducted into the causes of human cancer. A number of human autosomal recessive diseases such as Xeroderma Pigmentosum are characterized at least in part by a defect or aberration in one or more forms of DNA repair and at the same time an elevated incidence of cancer. Also, carcinogens cause mutations in DNA and the greater the carcinogenicity, the greater the mutagenicity. As a result, much attention has been focused on DNA repair and its relationship to cancer incidence. The HCR of V antigen formation by UV-irradiated Adenovirus type 2 (Ad 2) was examined using apparently normal human fibroblasts, tumor cells (HeLa CCL2), and cells transformed by Ad 5 DNA (293, 293 N3S). A decrease in the HCR of V antigen formation was found for HeLa CCL2 cells as compared to apparently normal human fibroblasts, but not for the transformed cells. These results are discussed in terms of the characteristics of the cell types. Herpes simplex virus type I encodes a polymerase and thymidine kinase (tk) activity which are involved in viral DNA synthesis. Paa ᷇ 5, an HSV-1 mutant containing one or more mutations in the polymerase gene is an antimutator. If these are also involved in viral DNA repair, then the HSV-1 polymerase, tk activity, and mutant polymerases conferring altered mutation rates should provide excellent tools with which to probe cellular DNA repair processes and mutagenesis. The study of the HCR of plaque forming ability of HSV-1 KOS wild type (WT), Paa ᷇ 5 and PTK3B (lacking thymidine kinase activity ) using VERO cells revealed a decrease in the HCR of Paa ᷇ 5 and increase of surviving fractions of PTK3B with respect to that of HSV-1 KOS WT. Similar studies using apparently normal human fibroblasts, Xeroderma Pigmentosum and Cockayne Syndrome cells also implicated the HSV-1 polymerase in viral DNA repair. The results are discussed in terms of the function of the HSV-1 polymerase and the DNA repair abilities of XP and CS cells. / Thesis / Master of Science (MSc)

The purpose of this study was to assess the effect of early region 1 functions of Ad5 on the ability of viruses containing a selectable marker in early region 3 (E3) to transform mammalian cells. To do this I have constructed and characterized five recombinant viruses containing the thymidine kinase (tk) gene from Herpes Simplex Virus type 1 (HSV1) inserted in E3. The biochemical transformation assay performed using these recombinant viruses allowed the separation of the selection process (incorporation and expression of tk) from the requirement for expression of E1 functions. The method of isolating the desired recombinant viruses was in vivo recombination following mixed infection of 293 cells. The parental viruses used were: hr 1 which expresses a truncated version of the E1a 243R product lacking amino acids 166 to 2437 pm975 which fails to express the E1a 289R product d1312 from which the majority of the E1a coding region has been deleted: hr6 which fails to express a wild type E1b 496R protein; and wild type Ad 5. Each coinfection was done with d1E1,3tk, a previously constructed recombinant virus (Haj-Ahmad and Graham, 1986). Using the resulting recombinant viruses, semipermissive tk- Rat 2 cells were infected and selected for conversion to the tk+ phenotype, as well as being assayed for viability post infection. Comparisons were made of tk transformation frequencies with and without correction for differential cell viability measured after infection with different viruses. Correction for differential cell viability greatly reduced the differences in transformation frequencies observed directly. However hr1tk remained able to induce tk+ transformation at a significantly greater frequency than hr6tk, pm975tk, or Ad5tk. The mutants d131,2tk and d1E1,3tk gave statistically indistinguishable results corresponding to an intermediate level of transformation, while hr6tk, pm975tk, and Ad5tk were grouped together as being least efficient at transformation. The infected Rat2 cell viability assays provide evidence of a correlation involving expression of the early region la (Ela) 289 residue product, efficient viral DNA replication, and cell death. Recombination frequencies obtained during the isolation of the recombinant viruses varied greatly depending on the combination of infecting parental viruses. The following factors appeared to affect recombination frequency: 1. the input ratio of the coinfecting viruses; 2. interference in the replication of d1E1,3tk relative to the other virus present; and 3. the presence of small numbers of mismatched base pairs (seven) near the left terminus of some of the viruses used in coinfection with d1E1,3tk. / Thesis / Master of Science (MS)