HIV-1 BF intersubtype recombinant Vpu second alpha helix plays an important role in the viral release and BST-2 degradation

DE CANDIA C; DUETTE G; ESPADA C; SALOMÓN H; CAROBENE M

Atlanta, Giorgia, USA

Conferencia; 20th Conference on Retroviruses and Opportunistic Infections; 2013

Background: Structural analysis of a BF intersubtype recombinant vpu variant with augmented capacity to enhance viral replication revealed that its transmembrane domain and α-helix-I from cytoplasmic domain corresponded to subtype B whereas α-helix-II corresponded to subtype F1. This work was aimed at evaluating the role of vpu α-helix-II on viral release enhancement and BST-2 and CD4 down-modulation from cell surface. Methods: VpuBF/B, where α-helix-II from the BF variant was replaced by the α-helix-II from the NL4-3 variant, and vpuB/F1, where α-helix-II from NL4-3 was replaced by the α-helix-II from the BF variant were generated. To evaluate viral production, pNL4-3 ∆vpu and pCG-vpuB, pCG-vpuF1, pCG-vpuBF, pCG-vpu B/F1, and pCG-vpu BF/B, were used to separately co-transfect HeLa cell cultures. pCG-GFP was used as negative control. Viral production was evaluated by p24 antigen. BST-2 and CD4 cell surface expression was evaluated by transfecting HeLa and HeLa CD4+ cells with vpu expression vectors separately; empty vector as a negative control. At 48 hours post transfection, BST-2 and CD4+ cell surface expression was evaluated by FACS. Statistical analysis was performed by t-student test. Results: Results showed that vpuB/F1 was more efficient in viral production than vpuB and vpuF1 (p >0.05). p24 production for the naturally occurring vpuBF was higher than for vpuBF/B (p >0.05). No difference in p24 production was observed between vpu BF/B and vpuB. VpuF1 p24 production was slightly but significantly higher than vpuB, and significantly lower than vpuBF (p