HPV-16 E7 Interacts with the Endocytic Machinery via the AP2 Adaptor μ2 Subunit.

Human papillomavirus (HPV) E7 plays a major role in HPV-induced malignancy, perturbing cell cycle regulation, and driving cell proliferation. Major targets of cancer-causing HPV E7 proteins are the pRB family of tumor suppressors, which E7 targets for proteasome-mediated degradation and whose interaction is promoted through an acidic patch, downstream of the LXCXE motif in E7, that is subject to phosphorylation by casein kinase II (CKII). In this study we show that HPV-16 E7 targets the AP2-complex, which plays a critical role in cargo recognition in clathrin-mediated endocytosis. Intriguingly, HPV-16 E7 contains a specific amino acid sequence for AP2 recognition, and this overlaps the pRb LXCXE recognition sequence but involves completely different amino acid residues. HPV-16 E7 does this by binding to the AP2-μ2 adaptor protein subunit via residues 25-YEQL-28 within the LXCXE motif. Point mutations at Y25 within 22-LYCYE-26 suggest that the interaction of E7 with AP2-μ2 is independent from pRB binding. In cells, this interaction is modulated by acidic residues downstream of LXCXE, with the binding being facilitated by CKII-phosphorylation of the serines at positions 31 and 32. Finally, we also show that association of HPV-16 E7 with the AP2 adaptor complex can contribute to cellular transformation under low-nutrient conditions, which appears to be mediated, in part, through inhibition of AP2-mediated internalization of epidermal growth factor receptor (EGFR). This indicates that E7 can modulate endocytic transport pathways, with one such component, EGFR, most likely contributing toward the ability of E7 to induce cell transformation and malignancy. These studies define a new and unexpected role for HPV-16 E7 in targeting clathrin-mediated endocytosis.