Mus musculus papillomavirus 1 E8

High-risk human papillomaviruses (PV) account for approximately 600,000 new cancers per year. The early protein E8^E2 is a conserved repressor of PV replication, whereas E4 is a late protein that arrests cells in G2 and collapses keratin filaments to facilitate virion release. While inactivation of the Mus musculus PV1 (MmuPV1) E8 start codon (E8-) increases viral gene expression, surprisingly, it prevents wart formation in FoxN1nu/nu mice. To understand this surprising phenotype, the impact of additional E8^E2 mutations was characterized in tissue culture and mice. MmuPV1 and HPV E8^E2 similarly interact with cellular NCoR/SMRT-HDAC3 co-repressor complexes. Disruption of the splice donor sequence used to generate the E8^E2 transcript or E8^E2 mutants (mt) with impaired binding to NCoR/SMRT-HDAC3 activates MmuPV1 transcription in murine keratinocytes. These MmuPV1 E8^E2 mt genomes also fail to induce warts in mice. The phenotype of E8^E2 mt genomes in undifferentiated cells resembles productive PV replication in differentiated keratinocytes. Consistent with this, E8^E2 mt genomes induced aberrant E4 expression in undifferentiated keratinocytes. In line with observations for HPV, MmuPV1 E4-positive cells displayed a shift to the G2 phase of the cell cycle. In summary, we propose that in order to enable both expansion of infected cells and wart formation in vivo, MmuPV1 E8^E2 inhibits E4 protein expression in the basal keratinocytes that would otherwise undergo E4-mediated cell cycle arrest. IMPORTANCE Human papillomaviruses (PVs) initiate productive replication, which is characterized by genome amplification and expression of E4 protein strictly within suprabasal, differentiated keratinocytes. Mus musculus PV1 mutants that disrupt splicing of the E8^E2 transcript or abolish the interaction of E8^E2 with cellular NCoR/SMRT-HDAC3 co-repressor complexes display increased gene expression in tissue culture but are unable to form warts in vivo. This confirms that the repressor activity of E8^E2 is required for tumor formation and genetically defines a conserved E8 interaction domain. E8^E2 prevents expression of E4 protein in basal-like, undifferentiated keratinocytes and thereby their arrest in G2 phase. Since binding of E8^E2 to NCoR/SMRT-HDAC3 co-repressor is required to enable expansion of infected cells in the basal layer and wart formation in vivo, this interaction represents a novel, conserved, and potentially druggable target.

Deregulation of host gene expression by HPV16 E8

Productive replication of human papillomaviruses (HPV) only takes place in differentiating keratinocytes. The HPV16 E8^E2 protein acts as a repressor of viral gene expression and genome replication and HPV16 E8^E2 knock-out (E8-) genomes display enhanced viral late protein expression in differentiated cells. Global transcriptome analysis of differentiated HPV16 wild-type and E8-cell lines revealed a small number of differentially expressed genes which are not related to cell cycle, DNA metabolism or keratinocyte differentiation. The analysis of selected genes suggested that deregulation requires cell differentiation and positively correlated with the expression of viral late, not early transcripts. Consistent with this, the additional knock-out of the viral E4 and E5 genes, which are known to enhance productive replication, attenuated the deregulation of these host cell genes. In summary, these data reveal that productive HPV16 replication modulates host cell transcription.

Functions of Papillomavirus E8

Papillomaviruses (PV) replicate in undifferentiated keratinocytes at low levels and to high levels in differentiated cells. The restricted replication in undifferentiated cells is mainly due to the expression of the conserved viral E8^E2 repressor protein, a fusion protein consisting of E8 and the hinge, DNA-binding, and dimerization domain of E2. E8^E2 binds to viral genomes and represses viral transcription and genome replication by recruiting cellular NCoR/SMRT-HDAC3 corepressor complexes. Tissue culture experiments have revealed that E8^E2 modulates long-term maintenance of extrachromosomal genomes, productive replication, and immortalization properties in a virus type-dependent manner. Furthermore, in vivo experiments have indicated that Mus musculus PV1 E8^E2 is required for tumor formation in immune-deficient mice. In summary, E8^E2 is a crucial inhibitor whose levels might determine the outcome of PV infections.