Project Summary/Abstract Human papillomavirus (HPV) infections are responsible for a staggering 5% of all cancers worldwide. In the developed world, these infections cause an increasing number of oropharyngeal cancers. An estimated third of all oropharyngeal squamous cell carcinomas (OPSCC) are caused by HPV infection. Sexual behavior is a potent risk factor for HPV(+)-OPSCC, suggesting that a better understanding of viral transmission is critical. The viral lifecycle, specifically genome amplification and virion production, is intricately linked with cellular differentiation. We will characterize cellular genes that regulate the productive stages of the viral lifecycle. We will use HPV16 positive tonsillar epithelium equivalents grown in three-dimensional (3D) organotypic raft culture to recapitulate the effects of cellular differentiation on the viral lifecycle. Using this model system, we will use high-throughput approaches to identify cellular genes that regulate the productive viral lifecycle. Specifically, we aim to 1) identify how the expression of specific human genes co-varies with distinct phases in the viral lifecycle; 2) use a genetic screening approach to characterize cellular genes that are required for late gene transcription. The details regulating the late, productive stage of the viral lifecycle are not understood. Understanding the interplay between viral and cellular genes regulating cellular differentiation will lead to an improved basic biological knowledge of the papillomavirus life cycle and towards the development of possible interventions for HPV infections and associated cancers.