The Large Marseillevirus Explores Different Entry Pathways by Forming Giant Infectious Vesicles

Abstract

Triggering the amoebal phagocytosis process is a sine qua non condition for most giant viruses to initiate their replication cycle and consequently to promote their progeny formation. It is well known that the amoebal phagocytosis process requires the recognition of particles of >500 nm, and most amoebal giant viruses meet this requirement, such as mimivirus, pandoravirus, pithovirus, and mollivirus. However, in the context of the discovery of amoebal giant viruses in the last decade, Marseillevirus marseillevirus (MsV) has drawn our attention, because despite its ability to successfully replicate in Acanthamoeba, remarkably it does not fulfill the >500-nm condition, since it presents an ϳ250-nm icosahedrally shaped capsid. We deeply investigated the MsV cycle by using a set of methods, including virological, molecular, and microscopic (immunofluorescence, scanning electron microscopy, and transmission electron microscopy) assays. Our results revealed that MsV is able to form giant vesicles containing dozens to thousands of viral particles wrapped by membranes derived from amoebal endoplasmic reticulum. Remarkably, our results strongly suggested that these giant vesicles are able to stimulate amoebal phagocytosis and to trigger the MsV replication cycle by an acidification-independent process. Also, we observed that MsV entry may occur by the phagocytosis of grouped particles (without surrounding membranes) and by an endosome-stimulated pathway triggered by single particles. Taken together, not only do our data deeply describe the main features of MsV replication cycle, but this is the first time, to our knowledge, that the formation of giant infective vesicles related to a DNA virus has been described. IMPORTANCETriggering the amoebal phagocytosis process is a sine qua non condition required by most giant viruses to initiate their replication cycle. This process requires the recognition of particles of >500 nm, and many giant viruses meet this requirement. However, MsV is unusual, as despite having particles of ϳ250 nm it is able to replicate in Acanthamoeba. Our results revealed that MsV is able to form giant vesicles, containing dozens to thousands of viral particles, wrapped in membranes derived from amoebal endoplasmic reticulum. Remarkably, our results strongly suggest that these giant vesicles are able to stimulate phagocytosis using an acidification-independent process. Our work not only describes the main features of the MsV replication cycle but also describes, for the first time to our knowledge, the formation of huge infective vesicles in a large DNA viruses. T he discovery of the amoebal giant virus Acanthamoeba polyphaga mimivirus (APMV) in 2003 (1) raised new and exciting questions regarding the virosphere and boosted the hunt for new giant viruses. Owing to these efforts, an increasing number of remarkable giant viruses have been described (2-6).Recent data suggest that giant viruses initiate their replication cycles after being phagocytosed by amoebas or other phagocytic cells (1, 7). Th...

Morphological and Taxonomic Properties of <i>Tokyovirus</i>, the First <i>Marseilleviridae</i> Member Isolated from Japan

“…revealed that it was larger than the amoeba cell nucleus, in many cases occupying 1/3 of the amoeba cytoplasm (Fig. 1C, 2, S3, Table S4 and S5), as observed for other Marseilleviridae members (5, 8). …”

“…Tokyovirus infection was accompanied by the large-scale development of VF in the amoeba cytoplasm, similar to that observed with other Marseilleviridae members (8, 10). A recent study reported that Marseillevirus possesses unique mechanisms for entry into Acanthamoeba cells, forming giant infectious vesicles surrounded by membranes, which differs from the entry mechanisms of other giant viruses (5). Tokyovirus was also found to display several morphological types in Acanthamoeba cells, such as giant vesicles including many viral particles surrounded by membranes and single particles in the amoeba cytoplasm (Fig.…”

Extracellular vesicles are the Trojan horses of viral infection

“…Indeed even smaller vesicles like exosomes (∼100 nm) may have the capacity to fit multiple HAV or HEV particles. This phenomena may not be limited to just RNA viruses as vesicular trafficking of viral populations has also been recently reported for Marseille virus, a large DNA virus of capsid size ∼250 nm, that appears to travel in populations of thousands within giant micrometer size vesicles [32]. …”

Lipid Tales of Viral Replication and Transmission

“…Even smaller vesicles, such as exosomes which are typically ~100nm in diameter, have the capacity to traffic multiple HAV or HEV viral particles [63]. While this novel type of viral transmission was first reported with (+) ssRNA viruses [3], recently a giant DNA virus, Marseille virus, has been discovered to also traffic between cells within micrometer vesicles, with each vesicle carrying hundreds of viral particles at once [95]. …”