Global Analysis of Murine Cytomegalovirus Open Reading Frames Using Yeast Two-Hybrid and Growth Phenotype Analysis

Human cytomegalovirus (HCMV), a beta-herpesvirus, is an important opportunistic pathogen that primarily affects individuals with compromised or immature immune systems. It is of great significance in AIDS patients where it can cause serious morbidity through retinitis-associated blindness, and other complications, such as pneumonia and enteritis. In developed nations, it is a leading viral cause of congenital disease, where in-utero infection manifests in mental and behavioral disorders. In order to control infection and HCMV associated disease, new compounds and novel strategies must be developed. Understanding the role viral proteins play during the course of infection will help elucidate the mechanisms of HCMV pathogenesis and provide important information on potential targets for new treatments. However, the strict species specificity of HCMV prevents any studies into the pathogenesis of the virus in an animal host. This limitation can be overcome through the use of murine cytomegalovirus (MCMV). MCMV, like HCMV, is a betaherpesvirus that exhibits similar pathogenesis in mice to HCMV infection in the human host. The genetic structure of MCMV contains significant sequence homology to HCMV AD169 in at least 78 ORFs and can thereby be used as an important tool in elucidating the functions of these ORFs in a complete in vivo system. In our study, we have conducted a comprehensive YTH screen to identify potential interactions between approximately 170 MCMV ORFs. Growth phenotype analysis were also conducted using five different cell lines potentially involved in various aspects of CMV infection. Between these 170 predicted proteins we have identified 94 potential interactions that exhibit varying levels of essentiality depending on the type of cell infected. We aim to understand the nature of the interactions between the viral particle and proteins encoded by the virus in order to elucidate potential mechanisms by which these proteins help to assemble and create new progeny viruses. The interactions that we have identified in this study provide a framework to predict the functions of uncharacterized viral proteins. And understanding the importance of each protein in the context of infection can further help to determine the nature of these unknown viral proteins. Together using information about known viral proteins that interact with these unknown elements, we can develop a better understanding of how all of these components contribute to viral infection which can be used to determine more effective methods to treat or prevent CMV associated diseases