Mechanisms for enveloped virus budding: can some viruses do without an ESCRT?

Abstract

Many enveloped viruses complete their replication cycle by forming vesicles that bud from the plasma membrane. Some viruses encode "late" (L) domain motifs that are able to hijack host proteins involved in the vacuolar protein sorting (VPS) pathway, a cellular budding process that gives rise to multivesicular bodies and that is topologically equivalent to virus budding. Although many enveloped viruses share this mechanism, examples of viruses that require additional viral factors and viruses that appear to be independent of the VPS pathway have been identified. Alternative mechanisms for virus budding could involve other topologically similar process such as cell abscission, which occurs following cytokinesis, or virus budding could proceed spontaneously as a result of lipid microdomain accumulation of viral proteins. Further examination of novel virus-host protein interactions and characterization of other enveloped viruses for which budding requirements are currently unknown will lead to a better understanding of the cellular processes involved in virus assembly and budding.

Figure 1. Vesicularization pathways

(A) Cellular VPS pathway and MVB biogenesis. Receptors on the plasma membrane destined for degradation (e.g. transferrin receptors) are internalized into vesicles by endocytosis which fuse with other vesicles to form endosomes en route to the lysosome. Ubiquitination of cargo proteins recruits the ESCRT complexes. To form MVBs, Vps4 AAA-ATPase activity is required to remove ESCRT-III proteins from the membrane and induce vesicularization. (B) Enveloped virus budding from the plasma membrane. For some enveloped viruses, viral proteins recruit components of the ESCRT complex through L domains, redirecting the MVB formation machinery from endosomes to sites of virus budding. (C) Abscission. To complete cytokinesis, abscission requires the recruitment of ESCRT proteins Tsg101 and AIP1/Alix by Cep55 to the midbody, and the activity of Vps4. The membrane topology of abscission, when the daughter cell pinches off, is equivalent to both MVB formation and virus budding.

Figure 2. ESCRT complexes and vesicle formation

ESCRT complexes I, II, and III are recruited to the endosomal membrane by ubiquitinated cargo proteins and inter-complex interactions. The AAA-ATPase Vps4A/B facilitates MVB vesicle formation by removing ESCRT-III subunits from the endosomal surface. Viral L domains found in viral proteins interact with components of the VPS pathway, redirecting the complexes to the site of virus budding on the plasma membrane.

Figure 3. Influenza VLP morphology as shown by EM

Influenza virions, VLPs containing all VLP proteins (WT VLP), and VLPs lacking M1 protein (M1-VLP) were prepared by infecting or transfecting 293T cells. Purified virions and VLPs were immunostained with a monoclonal anti-HA antibody followed by IgG conjugated to 15 nm gold and then negative stained. Bar, 100 nm. From Chen et al. (2007) with permission from the American Society for Microbiology.

Figure 4. Budding of HIV-1, influenza VLPs, PIV5 VLPs, and VSV in the presence of dominant-negative Vps4

Budding of HIV-1 in the presence of dominant-negative Vps4A-KQ or Vps4B-KQ (A), or budding of influenza VLPs (B), PIV5 VLPs (C), or VSV (D) in the presence of increasing amounts of dominant-negative Vps4A-EQ was tested. HIV-1 and PIV5 VLP budding was dramatically reduced whereas VSV and influenza VLP budding was not inhibited. Adapted from Chen et al. (2007) with permission from the American Society for Microbiology.

Figure 5. Computer simulation of spontaneous membrane vesicularization

In coarse-grained computer simulations modeling membrane vesicularization, capsids placed in the simulated lipid environment drove vesicle budding through attractive and cooperative forces. (A)-(F) shows a series of simulation snapshots over a time course of approximately 0.3 ms. Adapted from Reynwar et al. (2007) with permission from the Nature Publishing Group.