New Cellular Targets of HPV-16 E7

Cervical cancer is the third most common cancer in women, accounting for 9.8% of all new cancer cases worldwide. Human papillomaviruses (HPV s) play a central role in the etiology of cervical neoplasia. E7 is the major transforming protein of HPV s. The transforming activity of E7 has been attributed in part to its interaction with the retinoblastoma (pRb) tumour suppressor; however, the pRb interaction alone is not sufficient for transformation by E7. In a screen for cellular targets of HPV-16 E7, I identified the Ski interacting Qrotein, Skip, as a new interacting partner of E7. I show that HPV-16 E7 directly associates with Skip via sequences in its carboxy terminal region, and the evolutionarily conserved praline rich sequences (PRS) of the SNW domain of Skip. Skip has been found to bind to the highly conserved region of v-Ski that is required for its transforming activity, although the mechanism is not yet elucidated. I first investigated the biochemical significance of the Ski- Skip interaction in more detail. Here, .I have mapped the interaction site between Ski and Skip. I show that Skip acts as a generalised transcriptional co-activator, using a variety of different promoters. In addition, ectopic expression of Skip and Ski results in synergistic co-activation, which is dependent upon the Skip-Ski protein interaction. The Transcriptional trans-activating activities of Skip were mapped to the evolutionarily conserved core SNW domain of the protein. Moreover, this region was found to be essential for its ability to interact with cSki, and this data therefore provides a function for the Skip SNW domain. Taken together, these studies suggest that one of the ways in which Ski brings about transformation is by binding and cooperating with the SNW domain of Skip in transcriptional activation. Interestingly, both E7 and Ski oncoproteins bind to the same conserved SNW domain, suggesting that this is the key functional domain of Skip. HPV-16 E7 functionally targets Skip in viva and inhibits its transcriptional activation activity. However, E7 does not induce degradation of Skip. Two transformation defective mutants of E7 were identified that failed both to bind Skip and to inhibit its transcriptional activity. These results suggest that inhibition of Skip function may contribute to cell transformation by HPV-16 E7.