Angiomotin functions in HIV-1 assembly and budding

Area of Science:

• Virology
• Cell Biology
• Molecular Biology

Background:

• Retroviral Gag proteins utilize PPXY motifs to recruit NEDD4 E3 ubiquitin ligases for virus release.
• HIV-1 release can be stimulated by NEDD4L, even without a PPXY motif on Gag, suggesting an adaptor protein's role.

Purpose of the Study:

• To investigate the mechanism by which NEDD4L stimulates HIV-1 release in the absence of a PPXY motif.
• To identify and characterize the adaptor protein involved in NEDD4L-mediated HIV-1 release.

Main Methods:

• Co-immunoprecipitation assays to demonstrate binding between Angiomotin (AMOT), NEDD4L, and HIV-1 Gag.
• Overexpression and depletion studies of AMOT to assess its effect on HIV-1 release and infectivity.
• Site-directed mutagenesis to create AMOT mutants unable to bind NEDD4L.
• Electron microscopy to visualize virion assembly in the presence and absence of AMOT.

Main Results:

• Angiomotin (AMOT) binds to both NEDD4L and HIV-1 Gag.
• AMOT overexpression stimulates HIV-1 release and infectivity, while AMOT depletion inhibits them.
• AMOT mutants deficient in NEDD4L binding fail to rescue virus release.
• Absence of AMOT leads to incomplete spherical enveloped particle formation during virion assembly.

Conclusions:

• Angiomotin (AMOT) functions as a crucial adaptor protein linking NEDD4L and HIV-1 Gag.
• AMOT is essential for the final stages of immature virion assembly, prior to ESCRT-mediated budding.
• AMOT and NEDD4L collaborate to enhance HIV-1 release and infectivity.