Role of HIV-2 envelope in Lv2-mediated restriction

We have characterized envelope protein pseudotyped HIV-2 particles derived from two HIV-2 isolates termed prCBL23 and CBL23 in order to define the role of the envelope protein for the Lv2-mediated restriction to infection. Previously, it has been described that the primary isolate prCBL23 is restricted to infection of several human cell types, whereas the T cell line adapted isolate CBL23 is not restricted in these cell types. Molecular cloning of the two isolates revealed that the env and the gag gene are responsible for the observed phenotype and that this restriction is mediated by Lv2, which is distinct from Ref1/Lv1 (Schmitz, C., Marchant, D., Neil, S.J., Aubin, K., Reuter, S., Dittmar, M.T., McKnight, A., Kizhatil, K., Albritton, L.M., 2004. Lv2, a novel postentry restriction, is mediated by both capsid and envelope. J. Virol. 78 (4), 2006-2016). We generated pseudotyped viruses consisting of HIV-2 (ROD-ADeltaenv-GFP, ROD-ADeltaenv-RFP, or ROD-ADeltaenv-REN) and the prCBL23 or CBL23 envelope proteins as well as chimeric proteins between these envelopes. We demonstrate that a single amino acid exchange at position 74 in the surface unit of CBL23-Env confers restriction to infection. This single point mutation causes tighter CD4 binding, resulting in a less efficient fusion into the cytosol of the restricted cell line. Prevention of endosome formation and prevention of endosome acidification enhance infectivity of the restricted particles for GHOST/X4 cells indicating a degradative lysosomal pathway as a cause for the reduced cytosolic entry. The described restriction to infection of the primary isolate prCBL23 is therefore largely caused by an entry defect. A remaining restriction to infection (19-fold) is preserved when endosomal acidification is prevented. This restriction to infection is also dependent on the presence of the point mutation at position 74 (G74E).