The Human Papillomavirus Type 16 E5 Oncoprotein Inhibits Epidermal Growth Factor Trafficking Independently of Endosome Acidification

作者：Frank A. Suprynowicz, Ewa Krawczyk, Jess D. Hebert, Sawali R. Sudarshan, Vera Simić, Christopher M. Kamonjoh, Richard Schlegel · 发表于：Journal of Virology · 年份：2010 · DOI：10.1128/jvi.00831-10 · 被引用次数：73 · 研究领域：Cervical Cancer and HPV Research、interferon and immune responses、Virus-based gene therapy research

The human papillomavirus type 16 E5 oncoprotein (16E5) enhances acute, ligand-dependent activation of the epidermal growth factor receptor (EGFR) and concomitantly alkalinizes endosomes, presumably by binding to the 16-kDa "c" subunit of the V-ATPase proton pump (16K) and inhibiting V-ATPase function. However, the relationship between 16K binding, endosome alkalinization, and altered EGFR signaling remains unclear. Using an antibody that we generated against 16K, we found that 16E5 associated with only a small fraction of endogenous 16K in keratinocytes, suggesting that it was unlikely that E5 could significantly affect V-ATPase function by direct inhibition. Nevertheless, E5 inhibited the acidification of endosomes, as determined by a new assay using a biologically active, pH-sensitive fluorescent EGF conjugate. Since we also found that 16E5 did not alter cell surface EGF binding, the number of EGFRs on the cell surface, or the endocytosis of prebound EGF, we postulated that it might be blocking the fusion of early endosomes with acidified vesicles. Our studies with pH-sensitive and -insensitive fluorescent EGF conjugates and fluorescent dextran confirmed that E5 prevented endosome maturation (acidification and enlargement) by inhibiting endosome fusion. The E5-dependent defect in vesicle fusion was not due to detectable disruption of actin, tubulin, vimentin, or cytokeratin filaments, suggesting that membrane fusion was being directly affected rather than vesicle transport....