The role of virus structure and host cell membrane trafficking as well as signalling factors in Human Papillomavirus Type 16 endocytosis

• Antragsteller: Professor Dr. Mario Schelhaas
• Fachliche Zuordnung: Virologie
• Förderung: Förderung von 2008 bis 2015
• Projektkennung: Deutsche Forschungsgemeinschaft (DFG) - Projektnummer 83146105

Several human papillomaviruses (HPV) are the central cause for cervical cancer. The major HPV subtype found in cervix carcinomas is HPV-16. Little is known about the entry of HPVs into host cells except that it occurs via endocytosis. Our results indicate that HPV-16 enters host cells by a clathrin- and caveolin-independent mechanism that features some but not all characteristics of macropinocytosis, and that likely represents a novel endocytic route into cells. We propose to further investigate the cellular entry of HPV-16 by endocytosis. This project has the following aims: - To identify most of the host cell factors involved in HPV-16 endocytosis- To elucidate the role of host cell factors and signalling in the dynamics of HPV-16 endocytosis- To understand the molecular role of sulfated carbohydrates in binding of HPV-16- To identify structural changes in HPV-16 virions that serve as functional ‚cues’ for endocytic entry and trafficking as well as for uncoating of the virus.The main methods to be applied include various techniques for live cell microscopy as well as cell biological, morhological, genetic, and biochemical techniques. To categorise the requirements for HPV entry a genome-wide siRNA-mediated gene silencing screen will be performed in an automated high through-put assay. To gain temporal and spatial information on the events in HPV endocytosis, several advanced light microscopic techniques (wide field, confocal, and total internal reflection microscopy) will be used to follow single HPVs and their association with specific markers for endocytic organelles in living cells. To determine the carbohydrates that serve most specifically for binding of HPV-16 to host cells we will screen a carbohydrate microarrays. To elucidate structural changes in virus particles during endocytosis we will use isolation, biochemical, and ultrastructural analysis of virus particles from infected cells. The results are expected to contribute both to the understanding of HPV entry into host cells, and to the analysis of endocytic traffic in mammalian cells.

• DFG-Verfahren: Emmy Noether-Nachwuchsgruppen
• Großgeräte: Aufrüstung des im S2 Labor vorhandenen TIRF Mikroskops
• Gerätegruppe: 5080 Optisches Mikroskopzubehör