Papillomaviruses infect squamous epithelial tissues, and their replicative life cycle is tightly linked to the differentiation status of the infected epithelial tissues. HPVs initially infect dividing basal epithelial cells, which populate the bottom layer of the epithelium. There, HPVs establish a persistent infection, and the viral genome replicates in concert with the host cell DNA. Production of infectious viral progeny, however, is restricted to terminally differentiated epithelial cells. These cells are growth-arrested and, hence, they no longer produce the factors that are necessary to support HPV genome synthesis. HPVs encode up to three accessory proteins, E5, E6, and E7, that reprogram terminally differentiated epithelial cells to support large scale viral genome synthesis. The mechanistic contributions of the E5, E6, and E7 proteins have been extensively studied only for one specific HPV, HPV16, a member of the alpha genus. Given that all HPVs have to establish a replication-competent milieu in terminally differentiated, growth-arrested cells to produce viral progeny, it has been reasonably assumed that the insights gained from studies with HPV16 would apply to most if not all of the more than 440 phylogenetically different HPVs. More recent studies with beta genus HPVs, however, have revealed that this was an erroneous assumption. The beta HPVs only encode two accessory proteins, E6 and E7. They lack E5s and the molecular targets and biological activities of the E6 and E7 proteins are quite distinct. Unlike HPV16 E6, which binds and targets p53 for degradation, the beta HPV E6 proteins inhibit NOTCH and TGF-beta signaling by binding to MAML and SMAD proteins, respectively. The gamma genus HPVs include over 300 members, and similar to the beta HPVs, they do not encode E5 proteins. Gamma HPVs are almost entirely unstudied because most infections have not been linked to clinically relevant lesions. The members of the gamma 6 species, however, stand out because, unlike most gamma HPVs that infect cutaneous epithelia, they have all been isolated from anogenital tract mucosal epithelia. Similar to HPV16, they are likely sexually transmitted and have been linked to medically relevant cervical lesions. Moreover, the gamma 6 HPVs stand out by their unique genome organization. Unlike all other HPVs, the gamma 6 HPVs have no E6 proteins but instead encode unique, tiny hydrophobic transmembrane proteins, designated E10. We will determine the biochemical targets and biological activities of the gamma 6 HPV101 E10 and E7 proteins. The proposed work will provide fascinating insights that will be entirely novel to the HPV field. The studies will reveal how the HPV101 E7 and E10 proteins cooperate to permit the viral lifecycle in differentiated epithelial cells and how this may result in cervical lesions.