The Mechanism of Budding of Retroviruses from Cell Membranes
Parallel pathways in ASV and HIV-1 Gag budding. Retroviruses recruit components of the ESCRT machinery to build a budding complex for particle release. (1) The dominant L domains for HIV-1 and ASV Gag bind to Tsg101 and Nedd4, respectively. Whether this initial interaction takes place in the cytosol or at the plasma membrane remains to be defined. (2) Nedd4 mediates ubiquitination of ASV Gag. HIV-1 Gag is ubiquitinated by an unidentified E3 ligase. Some evidence suggests that Nedd4L may play a role since its overexpression rescues budding of HIV-1 Gag/PTAP. (3) Gag oligomerization in the cytosol increases membrane avidity and rapidly targets Gag to sites of assembly/budding on the plasma membrane. ASV Gag assembles on N-Rh-PE-positive, endosome-derived membranes. HIV-1 Gag assembles on N-Rh-PE-negative membranes. (4) During the budding process, Gag may recruit additional ESCRT factors eventually leading to ESCRT-III polymerization at the base of a budding particle. (5) ESCRT-III subunits recruit the AAA ATPase, Vps4, to mediate the disassembly of membrane-bound ESCRT complexes and to provide the energy for membrane fission. (6) VLPs are released from cellular membranes. Covalently linking ESCRT proteins to the C-terminus of Gag bearing L domain-mutations restores budding at different stages. Tethering Tsg101 to ASV Gag/p2b (p2b-Tsg101) or HIV-1 Gag/P7L (P7L-Tsg101) rescues budding through an HIV-like pathway (ESCRT-I-dependent, N-Rh-PE-negative membranes). Tethering EAP20 to ASLV Gag/p2b (p2b-Eap20) rescues budding through an ASLV-like pathway (ESCRT-II-dependent, N-Rh-PE-positive membranes). Black arrows indicate wild-type Gag budding pathways. Gray arrows indicate reconstituted budding pathways of ASLV Gag/p2b-ESCRT fusions and HIV-1 Gag/P7L-ESCRT fusions.
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