Figure 8. Model of VACV intracellular enveloped virion (IEV) formation and egress.

The formation of VACV IEVs proceeds through one of two pathways: Cisternae (CIS)-based wrapping (top), or as described in this report multivesicular body (MVB)–based wrapping (bottom). In both cases, single-membrane intracellular mature virions (IMVs) bud into virus-modified cellular membranes. For CIS wrapping, IMV is enveloped in a tight-fitting, double-membrane cisternae derived from the TGN or early endosome resulting in the formation of the triple-membrane IEV. During MVB-based wrapping, IMV(s) bud into the lumen of the MVB resulting in the acquisition of a tight second membrane, with the limiting membrane of the MVB effectively becoming the third IEV membrane. Although the mechanism and cellular factors that regulate closure of CIS-IEVs is unknown, we show that the formation—and presumably closure—of MVB-IEVs depends on cellular endosomal sorting complexes required for transport machinery. Upon formation, both CIS- and MVB-IEVs transit to the plasma membrane where they undergo fusion leaving behind the outermost membrane to become double-membrane extracellular enveloped virions. We found that MVB-based wrapping accounts for half of all VACV wrapping events and subsequent extracellular enveloped virion formation.