Neutralizing Epitopes in the Membrane-Proximal External Region of HIV-1 gp41 Are Influenced by the Transmembrane Domain and the Plasma Membrane

The membrane-proximal external region (MPER) of the HIV-1 envelope glycoprotein (Env) bears epitopes of broadly neutralizing antibodies (bnAbs) from infected individuals; it is thus a potential vaccine target. We report an NMR structure of the MPER and its adjacent transmembrane domain in bicelles that mimic a lipid-bilayer membrane. The MPER lies largely outside the lipid bilayer. It folds into a threefold cluster, stabilized mainly by conserved hydrophobic residues and potentially by interaction with phospholipid headgroups. Antigenic analysis and comparison with published images from electron cryotomography of HIV-1 Env on the virion surface suggest that the structure may represent a prefusion conformation of the MPER, distinct from the fusion-intermediate state targeted by several well-studied bnAbs. Very slow bnAb binding indicates that infrequent fluctuations of the MPER structure give these antibodies occasional access to alternative conformations of MPER epitopes. Mutations in the MPER not only impede membrane fusion but also influence presentation of bnAb epitopes in other regions. These results suggest strategies for developing MPER-based vaccine candidates.

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Evaluation of a novel multi-immunogen vaccine strategy for targeting 4E10/10E8 neutralizing epitopes on HIV-1 gp41 membrane proximal external region

Virology ◽

10.1016/j.virol.2017.02.015 ◽

2017 ◽

Vol 505 ◽

pp. 113-126 ◽

Cited By ~ 10

Author(s):

Saikat Banerjee ◽

Heliang Shi ◽

Marisa Banasik ◽

Hojin Moon ◽

William Lees ◽

...

Keyword(s):

Vaccine Strategy ◽

External Region ◽

Membrane Proximal External Region ◽

Hiv 1 ◽

Neutralizing Epitopes

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Cholesterol-Dependent Membrane Fusion Induced by the gp41 Membrane-Proximal External Region-Transmembrane Domain Connection Suggests a Mechanism for Broad HIV-1 Neutralization

Journal of Virology ◽

10.1128/jvi.02151-14 ◽

2014 ◽

Vol 88 (22) ◽

pp. 13367-13377 ◽

Cited By ~ 28

Author(s):

B. Apellaniz ◽

E. Rujas ◽

P. Carravilla ◽

J. Requejo-Isidro ◽

N. Huarte ◽

...

Keyword(s):

Membrane Fusion ◽

Transmembrane Domain ◽

External Region ◽

Membrane Proximal External Region ◽

Hiv 1

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The Atomic Structure of the HIV-1 gp41 Transmembrane Domain and Its Connection to the Immunogenic Membrane-proximal External Region

Journal of Biological Chemistry ◽

10.1074/jbc.m115.644351 ◽

2015 ◽

Vol 290 (21) ◽

pp. 12999-13015 ◽

Cited By ~ 31

Author(s):

Beatriz Apellániz ◽

Edurne Rujas ◽

Soraya Serrano ◽

Koldo Morante ◽

Kouhei Tsumoto ◽

...

Keyword(s):

Atomic Structure ◽

Transmembrane Domain ◽

External Region ◽

Membrane Proximal External Region ◽

Hiv 1

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Contribution of the Cytoplasmic Determinants of Vpu to the Expansion of Virus-Containing Compartments in HIV-1-Infected Macrophages

Journal of Virology ◽

10.1128/jvi.00020-19 ◽

2019 ◽

Vol 93 (11) ◽

Cited By ~ 4

Author(s):

Olivier Leymarie ◽

Leslie Lepont ◽

Margaux Versapuech ◽

Delphine Judith ◽

Sophie Abelanet ◽

...

Keyword(s):

Plasma Membrane ◽

Viral Particle ◽

Transmembrane Domain ◽

Immune Surveillance ◽

Restriction Factor ◽

Viral Particles ◽

Protein Functions ◽

Specific Accumulation ◽

Model Cell ◽

Hiv 1

ABSTRACTHIV-1 infection of macrophages leads to the sequestration of newly formed viruses in intracellular plasma membrane-connected structures termed virus-containing compartments (VCCs), where virions remain infectious and hidden from immune surveillance. The cellular restriction factor bone marrow stromal cell antigen 2 (BST2), which prevents HIV-1 dissemination by tethering budding viral particles at the plasma membrane, can be found in VCCs. The HIV-1 accessory protein Vpu counteracts the restriction factor BST2 by downregulating its expression and removing it from viral budding sites. Numerous studies described these Vpu countermeasures in CD4+T cells or model cell lines, but the interplay between Vpu and BST2 in VCC formation and HIV-1 production in macrophages is less explored. Here, we show that Vpu expression in HIV-1-infected macrophages enhances viral release. This effect is related to Vpu’s ability to circumvent BST2 antiviral activity. We show that in absence of Vpu, BST2 is enriched in VCCs and colocalizes with capsid p24, whereas Vpu expression significantly reduces the presence of BST2 in these compartments. Furthermore, our data reveal that BST2 is dispensable for the formation of VCCs and that Vpu expression impacts the volume of these compartments. This Vpu activity partly depends on BST2 expression and requires the integrity of the Vpu transmembrane domain, the dileucine-like motif E59XXXLV64and phosphoserines 52 and 56 of Vpu. Altogether, these results highlight that Vpu controls the volume of VCCs and promotes HIV-1 release from infected macrophages.IMPORTANCEHIV-1 infection of macrophages leads to the sequestration of newly formed viruses in virus-containing compartments (VCCs), where virions remain infectious and hidden from immune surveillance. The restriction factor BST2, which prevents HIV-1 dissemination by tethering budding viral particles, can be found in VCCs. The HIV-1 Vpu protein counteracts BST2. This study explores the interplay between Vpu and BST2 in the viral protein functions on HIV-1 release and viral particle sequestration in VCCs in macrophages. The results show that Vpu controls the volume of VCCs and favors viral particle release. These Vpu functions partly depend on Vpu’s ability to antagonize BST2. This study highlights that the transmembrane domain of Vpu and two motifs of the Vpu cytoplasmic domain are required for these functions. These motifs were notably involved in the control of the volume of VCCs by Vpu but were dispensable for the prevention of the specific accumulation of BST2 in these structures.

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Destabilization of Highly Rigid Bilayers Enriched in Cholesterol by the Membrane-Proximal External Region of HIV-1 gp41

Biophysical Journal ◽

10.1016/j.bpj.2010.12.3646 ◽

2011 ◽

Vol 100 (3) ◽

pp. 634a

Author(s):

Beatriz Apellaniz ◽

Andrey Ivankin ◽

David Gidalevitz ◽

Jose L. Nieva

Keyword(s):

External Region ◽

Membrane Proximal External Region ◽

Hiv 1

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Identification of SERINC5-001 as the Predominant Spliced Isoform for HIV-1 Restriction

Journal of Virology ◽

10.1128/jvi.00137-17 ◽

2017 ◽

Vol 91 (10) ◽

Cited By ~ 23

Author(s):

Xianfeng Zhang ◽

Tao Zhou ◽

Jie Yang ◽

Yumei Lin ◽

Jing Shi ◽

...

Keyword(s):

Plasma Membrane ◽

Transmembrane Domain ◽

The Other ◽

Stable Expression ◽

Membrane Localization ◽

Plasma Membrane Localization ◽

Immunodeficiency Virus ◽

Alternatively Spliced ◽

Anti Hiv ◽

Hiv 1

ABSTRACT Among the five serine incorporator (SERINC) family members, SERINC5 (Ser5) was reported to strongly inhibit HIV-1 replication, which is counteracted by Nef. Ser5 produces 5 alternatively spliced isoforms: Ser5-001 has 10 putative transmembrane domains, whereas Ser5-004, -005, -008a, and -008b do not have the last one. Here, we confirmed the strong Ser5 anti-HIV-1 activity and investigated its isoforms' expression and antiviral activities. It was found that Ser5-001 transcripts were detected at least 10-fold more than the other isoforms by real-time quantitative PCR. When Ser5-001 and its two isoforms Ser5-005 and Ser5-008a were expressed from the same mammalian expression vector, only Ser5-001 was stably expressed, whereas the others were poorly expressed due to rapid degradation. In addition, unlike the other isoforms, which are located mainly in the cytoplasm, Ser5-001 is localized primarily to the plasma membrane. To map the critical determinant, Ser5 mutants bearing C-terminal deletions were created. It was found that the 10th transmembrane domain is required for Ser5 stable expression and plasma membrane localization. As expected, only Ser5-001 strongly inhibits HIV-1 infectivity, whereas the other Ser5 isoforms and mutants that do not have the 10th transmembrane domain show very poor activity. It was also observed that the Nef counteractive activity could be easily saturated by Ser5 overexpression. Thus, we conclude that Ser5-001 is the predominant antiviral isoform that restricts HIV-1, and the 10th transmembrane domain plays a critical role in this process by regulating its protein stability and plasma membrane targeting. IMPORTANCE Human immunodeficiency virus (HIV) and simian immunodeficiency virus (SIV) express a small protein, Nef, to enhance viral pathogenesis in vivo. Nef has an important in vitro function, which is to make virus particles more infectious, but the mechanism has been unclear. Recently, Nef was reported to counteract a novel anti-HIV host protein, SERINC5 (Ser5). Ser5 has five alternatively spliced isoforms, Ser5-001, -004, -005, -008a, and -008b, and only Ser5-001 has an extra C-terminal transmembrane domain. We now show that the Ser5-001 transcripts are produced at least 10-fold more than the others, and only Ser5-001 produces stable proteins that are targeted to the plasma membrane. Importantly, only Ser5-001 shows strong anti-HIV-1 activity. We further demonstrate that the extra transmembrane domain is required for Ser5 stable expression and plasma membrane localization. These results suggest that plasma membrane localization is required for Ser5 antiviral activity, and Ser5-001 is the predominant isoform that contributes to the activity.

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The interaction between the membrane-proximal external region and the N-trimer region of HIV-1 gp41: Involvement in viral fusion

Science Bulletin ◽

10.1007/s11434-009-0280-6 ◽

2009 ◽

Vol 54 (10) ◽

pp. 1707-1712

Author(s):

Jing Li ◽

Lu Lu ◽

Fan Wu ◽

Xi Chen ◽

Ben Niu ◽

...

Keyword(s):

Viral Fusion ◽

External Region ◽

Membrane Proximal External Region ◽

Hiv 1

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Anti-V3/Glycan and Anti-MPER Neutralizing Antibodies, but Not Anti-V2/Glycan Site Antibodies, Are Strongly Associated with Greater Anti-HIV-1 Neutralization Breadth and Potency

Journal of Virology ◽

10.1128/jvi.00129-15 ◽

2015 ◽

Vol 89 (10) ◽

pp. 5264-5275 ◽

Cited By ~ 23

Author(s):

Rajesh Abraham Jacob ◽

Thandeka Moyo ◽

Michael Schomaker ◽

Fatima Abrahams ◽

Berta Grau Pujol ◽

...

Keyword(s):

Neutralizing Antibodies ◽

Prediction Interval ◽

Vaccine Design ◽

Broadly Neutralizing Antibodies ◽

External Region ◽

Membrane Proximal External Region ◽

Site Recognition ◽

Anti Hiv ◽

Glycan Site ◽

Hiv 1

ABSTRACTThe membrane-proximal external region (MPER), the V2/glycan site (initially defined by PG9 and PG16 antibodies), and the V3/glycans (initially defined by PGT121–128 antibodies) are targets of broadly neutralizing antibodies and potential targets for anti-HIV-1 antibody-based vaccines. Recent evidence shows that antibodies with moderate neutralization breadth are frequently attainable, with 50% of sera from chronically infected individuals neutralizing ≥50% of a large, diverse set of viruses. Nonetheless, there is little systematic information addressing which specificities are preferentially targeted among such commonly found, moderately broadly neutralizing sera. We explored associations between neutralization breadth and potency and the presence of neutralizing antibodies targeting the MPER, V2/glycan site, and V3/glycans in sera from 177 antiretroviral-naive HIV-1-infected (>1 year) individuals. Recognition of both MPER and V3/glycans was associated with increased breadth and potency. MPER-recognizing sera neutralized 4.62 more panel viruses than MPER-negative sera (95% prediction interval [95% PI], 4.41 to 5.20), and V3/glycan-recognizing sera neutralized 3.24 more panel viruses than V3/glycan-negative sera (95% PI, 3.15 to 3.52). In contrast, V2/glycan site-recognizing sera neutralized only 0.38 more panel viruses (95% PI, 0.20 to 0.45) than V2/glycan site-negative sera and no association between V2/glycan site recognition and breadth or potency was observed. Despite autoreactivity of many neutralizing antibodies recognizing MPER and V3/glycans, antibodies to these sites are major contributors to neutralization breadth and potency in this cohort. It may therefore be appropriate to focus on developing immunogens based upon the MPER and V3/glycans.IMPORTANCEPrevious candidate HIV vaccines have failed either to induce wide-coverage neutralizing antibodies or to substantially protect vaccinees. Therefore, current efforts focus on novel approaches never before successfully used in vaccine design, including modeling epitopes. Candidate immunogen models identified by broadly neutralizing antibodies include the membrane-proximal external region (MPER), V3/glycans, and the V2/glycan site. Autoreactivity and polyreactivity of anti-MPER and anti-V3/glycan antibodies are thought to pose both direct and indirect barriers to achieving neutralization breadth. We found that antibodies to the MPER and the V3/glycans contribute substantially to neutralization breadth and potency. In contrast, antibodies to the V2/glycan site were not associated with neutralization breadth/potency. This suggests that the autoreactivity effect is not critical and that the MPER and the V3/glycans should remain high-priority vaccine candidates. The V2/glycan site result is surprising because broadly neutralizing antibodies to this site have been repeatedly observed. Vaccine design priorities should shift toward the MPER and V3/glycans.

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