Genetic Analysis of the Tumorigenic Properties of Human Adenovirus Serotypes 5 and 12

Abstract

<p>Different serotypes of human adenovirus (Ad) have differing oncogenic potentials in rodents. These properties include the resistance of cells transformed or infected by oncogenic serotypes to lysis by natural killer cells, and the suppression of the expression of the class I major histocompatibility complex molecule in oncogenically transformed cells. Two commonly studied Ad serotypes are Ad 5 (non-oncogenic) and Ad 12 (oncogenic). The reasons for their differing oncogenicities are not fully understood.</p> <p>The purpose of this thesis project was the construction of hybrid Ad 5/Ad 12 early region 1 (E1) plasmids and viruses in order to identify sequences which affect the tumorigenic phenotype. Hybrids were constructed by intra-plasmid homologous recombination in Escherichia coli, and rescued into infectious Ad 5 based virus. With one exception, all viruses with hybrid E1A regions were able to replicate efficiently in HeLa cells. The exception, T1036, contained a hybrid E1A transactivation region. Viruses with Ad 12 E1B sequences were impaired for replication in HeLa cells and MH12C2 cells, which express Ad 12 E1 proteins, suggesting that Ad 12 E1B cannot support the replication of the Ad 5 genome.</p> <p>When tested for their ability to transform primary baby rat kidney cells, hybrid E1 plasmids showed two discreet of transforming efficiencies, based on their sequence compositions. The amino terminus and conserved region 2 of E1A were found to colocalize with regions involved in transforming efficiency, but no differences were found in the abilities of these sequences to bind to cellular proteins p105Kb, p107, and p300. This suggested that transformation involves functions of E1A in addition to the binding of cellular proteins.</p> <p>The tumorigenicity of hybrid E1 plasmids was evaluated, and was shown to involve a region of Ad 12 E1A between conserved regions 2 and 3. This region, when introduced into Ad 5 E1A, could impart a low degree of tumorigenicity to the new construct. This suggests that while the region identified plays a role in the oncogenicity of Ad 12, it is not the only factor which influences the differential degrees of oncogenicity between Ad 5 and Ad 12.</p>