Recombinant HIV type 1 gp160

Phosphatidylinositol 4,5-bisphosphate (PIP2) is critical for HIV-1 virus assembly. The viral membrane is enriched in PIP2, suggesting that the virus assembles at PIP2-rich microdomains. We showed previously that in model membranes PIP2 can form nanoscopic clusters bridged by multivalent cations. Here, using purified proteins we quantitated the binding of HIV-1 Gag-related proteins to giant unilamellar vesicles containing either clustered or free PIP2. Myristoylated MA strongly preferred binding to clustered PIP2. By contrast, unmyristoylated HIV-1 MA, RSV MA, and a PH domain all preferred to interact with free PIP2. We also found that HIV-1 Gag multimerization promotes PIP2 clustering. Truncated Gag proteins comprising the MA, CA, and SP domains (MACASP) or the MA and CA domains (MACA) induced self-quenching of acyl chain-labeled fluorescent PIP2 in liposomes, implying clustering. However, HIV-1 MA itself did not induce PIP2 clustering. A CA inter-hexamer dimer interface mutation led to a loss of induced PIP2 clustering in MACA, indicating the importance of protein multimerization. Cryo-electron tomography of liposomes with bound MACA showed an amorphous protein layer on the membrane surface. Thus, it appears that while protein-protein interactions are required for PIP2 clustering, formation of a regular lattice is not. Protein-induced PIP2 clustering and multivalent cation-induced PIP2 clustering are additive. Taken together, these results provide the first evidence that HIV-1 Gag can selectively target pre-existing PIP2-enriched domains of the plasma membrane for viral assembly, and that Gag multimerization can further enrich PIP2 at assembly sites. These effects could explain the observed PIP2 enrichment in HIV-1. (C) 2020 The Authors. Published by Elsevier Ltd.

To explore the molecular epidemiological status of human immunodeficiency virus type 1 (HIV-1) in Ganzhou, China, eight HIV-1-positive outpatients were recruited from July 5 to 21, 2018. Six HIV-1 near-full-length sequences were amplified and sequenced from the plasma samples that were collected before the patients' antiretroviral treatments. Phylogenetic and bootscan analyses revealed that one of the sequences was CRF01_AE, one was CRF55_01B, and two were CRF07_BC. Notably, one of the sequences was a unique recombinant form, and one of them was a second-generation recombinant form of CRF07_BC and subtype C. These results revealed that multiple HIV-1 subtypes are circulating in Ganzhou, China. Systematic surveys with large sample sizes are urgently needed to explore the exact molecular epidemiological characteristics and to trace the origins of HIV-1 in Ganzhou, China.