Expression of E8^E2 Is Required for Wart Formation by Mouse Papillomavirus 1 In Vivo

Human papillomavirus (HPV) E1 and E2 proteins activate genome replication. E2 also modulates viral gene expression and is involved in the segregation of viral genomes. In addition to full-length E2, almost all papillomaviruses (PV) share the ability to encode an E8^E2 protein, which is a fusion of E8 with the C-terminal half of E2 that mediates specific DNA binding and dimerization. HPV E8^E2 acts as a repressor of viral gene expression and genome replication. To analyze the function of E8^E2 in vivo, we used the Mus musculus PV1 (MmuPV1) mouse model system. Characterization of the MmuPV1 E8^E2 protein revealed that it inhibits transcription from viral promoters in the absence and presence of E1 and E2 proteins and that this is partially dependent upon the E8 domain. MmuPV1 genomes in which the E8 ATG start codon was disrupted (E8(−)) displayed a 10- to 25-fold increase in viral gene expression compared to that of wild-type (wt) genomes in cultured normal mouse tail keratinocytes in short-term experiments. This suggests that the function and mechanism of E8^E2 is conserved between MmuPV1 and HPV. Surprisingly, challenge of athymic nude Foxn1(nu/nu) mice with MmuPV1 E8(−) genomes did not induce warts on the tail, in contrast to wt MmuPV1. Furthermore, viral gene expression was completely absent at E8(−) MmuPV1 sites 20 to 22 weeks after DNA challenge on the tail or quasivirion (QV) challenge in the vaginal vault. This reveals that expression of E8^E2 is necessary to form tumors in vivo and that this is independent from the presence of T cells. IMPORTANCE HPV encodes an E8^E2 protein which acts as repressors of viral gene expression and genome replication. In cultured normal keratinocytes, E8^E2 is essential for long-term episomal maintenance of HPV 31 (HPV31) genomes, but not for that of HPV16. To understand the role of E8^E2 in vivo, the Mus musculus PV1 (MmuPV1) mouse model system was used. This revealed that E8^E2’s function as a repressor of viral gene expression is conserved. Surprisingly, MmuPV1 E8^E2 knockout genomes did not induce warts in T cell-deficient mice. This shows for the first time that expression of E8^E2 is necessary for tumor formation in vivo, independent of T-cell immunity. This indicates that E8^E2 could be an interesting target for antiviral therapy in vivo.