Emerging arthropod-borne viruses, such as alphaviruses and bunyaviruses, represent a serious threat to human and animal health worldwide, and for most of them, vaccines and specific treatments are
unavailable. Viral host cell entry can be divided into several entry checkpoints, and the most important checkpoints for low pH-dependent
enveloped viruses, such as bunyaviruses and alphaviruses, include receptor binding at the cell surface and, followed by endocytosis, low pH
dependent membrane fusion from within intracellular compartments. A more thorough understanding of the detailed mechanisms allowing the viruses to pass these checkpoints is a pre-requisite for the design of viral entry inhibitors.
This thesis reports the in vitro analysis of native alphavirus-receptor interactions, with the help of electron cryo-microscopy and icosahedral
reconstruction of virus-recaptor complexes, using the prototypic alphavirus Semliki Forest virus (SFV) and the C-type lectin DC-SIGN. Together with
results from collaborative work on SFV glycosylation, this study provides progress in defining the binding sites of DC-SIGN at the surface of SFV.
Second, an in vitro system for phlebovirus fusion was developed using standard fluorometry, and has been characterized with the help of electron
cryo-microscopy. It was discovered that negatively charged phospholipids with a conical shape, including the late endosomal phospholipid BMP,
allow efficient phlebovirus fusion in vitro, thereby providing a possible rationale for phlebovirus fusion in late endosomes. Furthermore, electron
cryo-microscopy of phlebovirus-liposome complexes allowed the capture of early stage fusion intermediates and laid the basis for possible future
higher resolution studies of these fusion intermediates.This thesis is not currently available on ORA