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. 2021 Nov 6;13(11):2236.
doi: 10.3390/v13112236.

HIV-1 Envelope Glycoproteins Proteolytic Cleavage Protects Infected Cells from ADCC Mediated by Plasma from Infected Individuals

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HIV-1 Envelope Glycoproteins Proteolytic Cleavage Protects Infected Cells from ADCC Mediated by Plasma from Infected Individuals

Jérémie Prévost et al. Viruses. .

Abstract

The HIV-1 envelope glycoprotein (Env) is synthesized in the endoplasmic reticulum as a trimeric gp160 precursor, which requires proteolytic cleavage by a cellular furin protease to mediate virus-cell fusion. Env is conformationally flexible but controls its transition from the unbound "closed" conformation (State 1) to downstream CD4-bound conformations (States 2/3), which are required for fusion. In particular, HIV-1 has evolved several mechanisms that reduce the premature "opening" of Env which exposes highly conserved epitopes recognized by non-neutralizing antibodies (nnAbs) capable of mediating antibody-dependent cellular cytotoxicity (ADCC). Env cleavage decreases its conformational transitions favoring the adoption of the "closed" conformation. Here we altered the gp160 furin cleavage site to impair Env cleavage and to examine its impact on ADCC responses mediated by plasma from HIV-1-infected individuals. We found that infected primary CD4+ T cells expressing uncleaved, but not wildtype, Env are efficiently recognized by nnAbs and become highly susceptible to ADCC responses mediated by plasma from HIV-1-infected individuals. Thus, HIV-1 limits the exposure of uncleaved Env at the surface of HIV-1-infected cells at least in part to escape ADCC responses.

Keywords: ADCC; CD4 mimetics; Env glycoprotein; HIV+ plasma; HIV-1; Temsavir; furin cleavage site; nnAbs.

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Conflict of interest statement

The authors declare no competing interest.

Figures

Figure 1
Figure 1
Proteolytic cleavage stabilizes Env in its “closed” conformation. (A) Sequence alignment of the HIV-1 Env furin cleavage site region from primary viruses CH058 (GenBank accession number JN944940) and CH077 (GenBank accession number JN944941) with the HXB2 reference strain (GenBank accession number K03455). Putative furin cleavage sequences are highlighted by black boxes. Positively charged residues (arginine and lysine) are shown in blue. Residue numbering is based on the HXB2 strain. Identical residues are shaded in dark gray, and conserved residues are shaded in light gray. (BE) 293T cells were transfected with primary IMCs (B,C) CH058, (D,E) CH077 WT or their cleavage-deficient (Cl−) variants and metabolically labeled with [35S]-methionine and [35S]-cysteine. (B,D) Cell lysates and supernatants were immunoprecipitated with plasma from HIV-1-infected individuals. The precipitated proteins were loaded onto SDS-PAGE gels and analyzed by autoradiography and densitometry to calculate their processing indexes. The processing index is a measure of the conversion of the mutant gp160 Env precursor to mature gp120, relative to the wild-type Env trimer. (C,E) Shown is the average of processing indexes calculated in 3 independent experiments. (FI) Cell-surface staining of (F,G) IMC transfected 293T cells (H,I) or primary CD4+ T cells infected with IMCs (F,H) CH058 and (G,I) CH077 WT or their cleavage-deficient (Cl−) variants using a panel of anti-Env bNAbs (PGT126, PG9, PGT151, VRC34, VRC03) and nnAbs (19b, F240, 17b, A32, C11). Shown are the mean fluorescence intensities (MFI) using the different antibodies normalized to the signal obtained with the conformation-independent 2G12 mAb. MFI values were measured on the transfected or infected (p24+) population for staining obtained in at least 3 independent experiments. Error bars indicate the mean ± SEM. Statistical significance was tested using an unpaired t-test (* p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001).
Figure 2
Figure 2
Virions displaying uncleaved Env are better recognized by nnAbs. (A) Viral infectivity was assessed by incubating TZM-bl target cells with HIV-1 CH058 virions expressing the wild-type (WT) or cleavage-deficient (Cl−) Env glycoprotein for 48 h. Viral preparations were normalized according to reverse transcriptase activity. (B) VSV-G-pseudotyped viral particles encoding the luciferase gene (Luc+) and bearing HIV-1 CH058 Env wildtype (WT) or its cleavage-deficient mutant (Cl−) were used to infect 293T cells to determine their infectivity in a single-round infection. (C,D) These recombinant pseudovirions were further tested for virus capture by (C) a panel of anti-Env bNAbs (PGT126, PG9, PGT151, VRC34, VRC03) and nnAbs (19b, F240, 17b, A32, C11) or (D) HIV-IG. RLU values obtained using the different antibodies were normalized to the signal obtained with the conformation-independent 2G12 mAb. Data shown are the mean ± SEM from at least three independent experiments. Statistical significance was tested using an unpaired t-test or a Mann–Whitney U-test based on statistical normality (* p  <  0.05; ** p  <  0.01; *** p  <  0.001; **** p < 0.0001; ns, nonsignificant).
Figure 3
Figure 3
Env cleavage protects HIV-1-infected cells from ADCC mediated by HIV+ plasma. (A,B) Cell surface staining of primary CD4+T cells infected with primary HIV-1 viruses (A) CH058 and (B) CH077 WT or their cleavage-deficient (Cl−) variants using plasma from 15 different HIV-1-infected individuals in the presence of 50 µM of CD4mc BNM-III-170, conformational blocker Temsavir or an equivalent volume of the vehicle (DMSO). The graphs show the MFI obtained on the infected (p24+) cell population. (C,D) Primary CD4+ T cells infected with (C) CH058 and (D) CH077 viruses were also used as target cells, and autologous PBMCs were used as effector cells in a FACS-based ADCC assay. The graphs shown represent the percentages of ADCC mediated by 15 different HIV+ plasma in the presence of 50 µM of CD4mc BNM-III-170, attachment inhibitor Temsavir or an equivalent volume of the vehicle (DMSO). All results were obtained using cells from at least three different donors. Error bars indicate the means ± SEM. Statistical significance was tested using a repeated measures one-way ANOVA with a Holm–Sidak post-test (* p  <  0.05; ** p  <  0.01; *** p  <  0.001; **** p  <  0.0001; ns, nonsignificant).

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References

    1. Earl P.L., Doms R.W., Moss B. Oligomeric structure of the human immunodeficiency virus type 1 envelope glycoprotein. Proc. Natl. Acad. Sci. USA. 1990;87:648–652. doi: 10.1073/pnas.87.2.648. - DOI - PMC - PubMed
    1. Earl P.L., Moss B., Doms R.W. Folding, interaction with GRP78-BiP, assembly, and transport of the human immunodeficiency virus type 1 envelope protein. J. Virol. 1991;65:2047–2055. doi: 10.1128/jvi.65.4.2047-2055.1991. - DOI - PMC - PubMed
    1. Kantanen M.L., Leinikki P., Kuismanen E. Endoproteolytic cleavage of HIV-1 gp160 envelope precursor occurs after exit from the trans-Golgi network (TGN) Arch. Virol. 1995;140:1441–1449. doi: 10.1007/BF01322670. - DOI - PubMed
    1. Qi M., Williams J.A., Chu H., Chen X., Wang J.J., Ding L., Akhirome E., Wen X., Lapierre L.A., Goldenring J.R., et al. Rab11-FIP1C and Rab14 direct plasma membrane sorting and particle incorporation of the HIV-1 envelope glycoprotein complex. PLoS Pathog. 2013;9:e1003278. doi: 10.1371/journal.ppat.1003278. - DOI - PMC - PubMed
    1. Kirschman J., Qi M., Ding L., Hammonds J., Dienger-Stambaugh K., Wang J.J., Lapierre L.A., Goldenring J.R., Spearman P. HIV-1 Envelope Glycoprotein Trafficking through the Endosomal Recycling Compartment Is Required for Particle Incorporation. J. Virol. 2018;92 doi: 10.1128/JVI.01893-17. - DOI - PMC - PubMed

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