Calicivirus VP2 forms a portal to mediate endosome escape

Abstract

To initiate the infectious process, many viruses enter their host cells by triggering endocytosis following receptor engagement. The mechanism by which non-enveloped viruses, such as the caliciviruses, escape the endosome is however poorly understood. The Caliciviridae include many important human and animal pathogens, most notably norovirus, the cause of winter vomiting disease. Here we show that VP2, a minor capsid protein encoded by all caliciviruses, forms a large portal assembly at a unique three-fold symmetry axis following receptor engagement. This feature surrounds an open pore in the capsid shell. We hypothesise that the VP2 portal complex is the means by which the virus escapes the endosome, penetrating the endosomal membrane to release the viral genome into the cytoplasm. Cryogenic electron microscopy (cryoEM) and asymmetric reconstruction were used to investigate structural changes in the capsid of feline calicivirus (FCV) that occur when the virus binds to its cellular receptor junctional adhesion molecule-A (fJAM-A). Near atomic-resolution structures were calculated for the native virion alone and decorated with soluble receptor fragments. We present atomic models of the major capsid protein VP1 in the presence and absence of fJAM-A, revealing the contact interface and conformational changes brought about by the interaction. Furthermore, we have calculated an atomic model of the portal protein VP2 and revealed the structural changes in VP1 that lead to pore formation. While VP2 was known to be critical for the production of infectious virus, its function has been hitherto undetermined. Our finding that VP2 assembles a portal that is likely responsible for endosome escape represents a major step forward in our understanding of both the Caliciviridae and icosahedral RNA containing viruses in general.

Dynamics in the murine norovirus capsid revealed by high-resolution cryo-EM

“…However, in line with other norovirus structures, we cannot locate VP2 in any of the structures reported here. Based on recent work with feline calicivirus, it has been proposed that VP2 may bind the capsid interior at a single icosahedral 3-fold axis [29]. Whilst it has been shown through mutational analysis that VP1 N-termini are not required for coprecipitation with VP2 [15], the ability of VP1 N-termini to organise differently may provide an interface for 'recognition' of the 3-fold axis (formed by B-and C-type VP1 subunits) either directly or indirectly, e.g., VPg, which in turn guides VP2.…”

Structural basis of rotavirus RNA chaperone displacement and RNA annealing

“…To improve visualization of the RNA density, we performed focused classification using C4 symmetry-expanded data with a mask applied to a single RNA-binding face of NSP2 (22)(23)(24).…”

Collection, pre-processing and on-the-fly analysis of data for high-resolution, single-particle cryo-electron microscopy

“…A low magnification atlas is then taken of the grid to help identify regions of appropriate ice thickness for data collection (Step 26). The areas for automated collection are then selected (27)(28)(29)(30).…”