This project is concerned with the functions of the products of transcriptional early region 4 (E4) of human adenoviruses types 2 and 5. E4 covers about 3000 base pairs near the right end of the adenoviral genome, and E4 products are known to play roles in the expression of viral late genes (at the level of the synthesis of viral late mRNA), the replication of viral DNA, the assembly of virus particles, and the inhibition of host cell protein synthesis after infection. Both genetic and biochemical approaches will be employed to examine E4 function. Genetic studies will include completion of the detailed characterization of a series of E4 deletion mutants to determine the roles in the viral life cycle and the locations of genes and sites in E4, the construction of specific E4 point mutants to probe the functions of gene products already implicated in regulatory events, and the isolation of second-site revertants of E4 deletion mutants to identify viral sites and gene products that interact with E4 proteins. Biochemical studies will concentrate on the roles played in late protein synthesis by E4 products. In Vitro systems will first be developed that duplicate the disruption of late gene expression observed in E4 mutant infections. Those systems will be used to precisely define the defect, and in conjunction with E4 products overproduced by recombinant DNA techniques, will be used in studies of the interaction of E4 proteins with RNA, other viral gene products, and the components of the host cell genetic expression machinery. The effects of E4 mutations on viral DNA replication, fiber synthesis, and transformation will also be investigated in greater detail in an effort to determine how E4 products are involved in those processes. This project will provide information of the regulatory that occur in cells infected with human adenoviruses. Emphasis will be placed on analyzing a region of the viral genome known as E4 that regulates viral late gene expression, DNA replication, assembly of virus particles, and inhibition of host cell metablism. Dr. Ketner has developed a host cell line that contains and expresses the viral E4 region. As a result, viral E4 mutants, which would otherwise be lethal, can be propagated and analyzed. The results of these provide an understanding of the regulatory functions of the different genes within the E4 region.
