A nuclear export signal within the structural Gag protein is required for prototype foamy virus replication

Area of Science:

• Virology
• Molecular Biology
• Cell Biology

Background:

• Retrovirus Gag polyproteins are essential for replication, utilizing cellular machinery.
• Prototype foamy virus (PFV) Gag proteins traffic to the nucleus before returning to the cytoplasm for assembly and egress.
• The function of nuclear Gag trafficking and its export mechanism remain unclear.

Purpose of the Study:

• To investigate the molecular mechanisms governing Prototype Foamy Virus (PFV) Gag nuclear export.
• To identify functional elements within PFV Gag responsible for its nuclear-cytoplasmic trafficking.
• To elucidate the role of nuclear export in the PFV replication cycle.

Main Methods:

• Identification and mutagenesis of a nuclear export sequence (NES) in PFV Gag.
• Leptomycin B (LMB) sensitivity assays to assess NES function.
• Dominant-negative mutant analysis and trans-complementation experiments using HIV-1 Rev NES.

Main Results:

• A leptomycin B (LMB)-sensitive nuclear export sequence (NES) was identified in the N-terminus of PFV Gag, crucial for late-stage replication.
• Mutations in the PFV Gag NES caused nuclear accumulation of Gag, blocking virus egress.
• NES-defective PFV Gag acted as a dominant-negative mutant, sequestering wild-type Gag in the nucleus.
• While heterologous HIV-1 Rev NES could rescue cytoplasmic localization, it did not restore PFV infectivity.

Conclusions:

• PFV Gag-Gag interactions are critical for cytoplasmic functions like capsid assembly and virus release.
• Nuclear Gag-Gag interactions are implicated in the nuclear export of Gag and viral RNA.
• PFV RNA export likely involves two sequential, complementary mechanisms during replication.