Two conserved amino acids differentiate the biology of high-risk and low-risk HPV E5 proteins.

The high-risk alpha human papillomaviruses (HPVs) are responsible for 99% of cervical cancers. While the biological functions of the HPV E6 and E7 oncoproteins are well-characterized, the function of E5 has remained elusive. Here, we examined gene expression changes induced by E5 proteins from high-risk HPV-16 and low-risk HPV-6b in multiple pools of primary human keratinocytes. Surprisingly, microarray analysis revealed that over 700 genes were significantly regulated by HPV-6b E5, while only 25 genes were consistently and significantly regulated by HPV-16 E5 in three biological replicates. However, we observed that more than thousand genes were altered in individual sample compared with vector. The gene expression profile induced by 16E5 in primary genital keratinocytes was very different from what has been previously published using immortalized HaCaT cells. Genes altered by HPV-16 E5 were unaffected by HPV-6b E5. Our data demonstrate that E5 proteins from the high- and low-risk HPVs have different functions in the HPV-host cell. Interestingly, conversion of two amino acids in HPV-16 E5 to the low-risk HPV-6b sequence eliminated the induction of high-risk related cellular genes.