Complementation of diverse HIV-1 Env defects through cooperative subunit interactions: a general property of the functional trimer

Salzwedel, Karl; Berger, Edward A.

doi:10.1186/1742-4690-6-75

Cited by 19 publications

(22 citation statements)

References 61 publications

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“…S4A shows that complementation was readily detectable with the pseudotype assay, although the efficiency of the mixed trimers was ∼100-fold less than that observed with wild-type constructs based on the 50% tissue culture infectious dose (TCID 50 ). This lower efficiency presumably accounts for our previous lack of detection of mixed trimer complementation by virions using the less sensitive multinuclear-activation galactosidase indicator (MAGI) infectivity assay (15). Fig.…”

Section: Demonstration Of Env Complementation With Hiv Virion Particlesmentioning

confidence: 97%

“…Our previous analyses of subunit complementation (i.e., generation of functional mixed trimers upon coexpression of two Envs with different inactivating mutations) used the Env-mediated cell fusion assay (14,15), which is highly sensitive for detecting Env function. Here we wished to apply the Env pseudotype virion infection system to test for complementation with virus particles.…”

Section: Demonstration Of Env Complementation With Hiv Virion Particlesmentioning

confidence: 99%

“…We devised a functional strategy based on a previously described Env complementation system (14,15) to distinguish between cis versus trans mechanisms of epitope masking. The results are discussed in terms of various proposed trimer models based on structural analyses and molecular modeling.…”

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confidence: 99%

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Intraprotomer masking of third variable loop (V3) epitopes by the first and second variable loops (V1V2) within the native HIV-1 envelope glycoprotein trimer

Liu

Cimbro²,

Lusso³

et al. 2011

Proc. Natl. Acad. Sci. U.S.A.

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Within the trimeric HIV-1 envelope (Env) spike, the first and second variable loops (V1V2 region) and the third variable loop (V3) of the gp120 subunit play dual roles in antibody recognition, because they contain neutralization epitopes and also participate in epitope masking. The spatial relationships between V1V2 and V3 and the associated mechanisms of epitope masking remain unclear. Here we investigated interactions between these domains using two monoclonal antibodies recognizing distinct conserved linear epitopes that are subject to masking in the functional trimer, which limits their neutralizing activities. Using Env pseudotype virus infection assays, we found that deleting the V1V2 region greatly enhanced neutralization by both antibodies, leading us to consider two alternative models: V1V2 on one gp120 protomer masks V3 on the same protomer (intraprotomer or cis masking) versus on an adjacent protomer (interprotomer or trans masking). Our experimental approach exploited a previously described complementation system wherein two variant Envs harboring different inactivating mutations (one in gp120, the other in gp41) are coexpressed in the same cell; functional Env results only from cooperative interactions within mixed trimers, thereby enabling selective examination of mixed trimer activity. We introduced additional mutations that either promoted (V1V2 deletion, i.e., unmasking) or prevented (GPGR to GPGQ mutation, i.e., epitope destruction) interaction with the antibodies. The observed neutralization sensitivities of mixed trimers produced from various combinations of constructs support the intraprotomer (cis) model of V1V2 masking of V3 epitopes.envelope glycoprotein trimer | functional complementation | Env trimer structure | cis versus trans T he envelope glycoprotein (Env) of HIV-1 and the related simian immunodeficiency virus (SIV) is the molecular machine that drives virion entry into the target cell (1). The Env spikes displayed on the surface of virions and infected cells are homotrimers of gp120/gp41 heterodimers, derived by proteolytic maturation of the gp160 precursor. Upon initiation of HIV-1 infection, gp120 binds first to the "primary" cellular receptor CD4, then to the "coreceptor," i.e., chemokine receptor CCR5 or CXCR4. These sequential events are associated with dramatic conformational changes in both the gp120 and gp41 subunits, leading to insertion of the N-terminal fusion peptide of gp41 into the membrane of the target cell, followed by gp41 refolding and direct fusion of the virion and plasma membranes. The end result is virion entry. Because of its role in the initial step of HIV infection and its prominent display on the virion surface, the Env spike is the primary target of neutralizing antibodies during natural infection and for vaccine development (2). Moreover, the preferential reactivity of an Env for CCR5 and/or CXCR4 dictates the tropism of HIV-1 variants for different CD4-expressing target cell types, a critical determinant of HIV transmission and pathogenesis (3). Entry...

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Section: Demonstration Of Env Complementation With Hiv Virion Particlesmentioning

confidence: 97%

Section: Demonstration Of Env Complementation With Hiv Virion Particlesmentioning

confidence: 99%

See 1 more Smart Citation

Intraprotomer masking of third variable loop (V3) epitopes by the first and second variable loops (V1V2) within the native HIV-1 envelope glycoprotein trimer

Liu

Cimbro²,

Lusso³

et al. 2011

Proc. Natl. Acad. Sci. U.S.A.

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“…The assembly and virion incorporation of such heterotrimers has been confirmed previously (Herrera et al, 2006;Kuhmann et al, 2000;Salzwedel and Berger, 2009). We produced these chimeric pseudotyped viruses, which are capable of only single-round infection, as previously described (Zhang et al, 2004).…”

Section: Generating Chimeric Virusmentioning

confidence: 67%

“…The production of heterotrimers, consisting of wild type and mutant Env subunits, has been confirmed previously (Herrera et al, 2006;Kuhmann et al, 2000;Salzwedel and Berger, 2009). To demonstrate that the amount of DNA used in the transfections directly corresponds to the amount of gp120 present in virions, we ran Western blots on virus preparations produced with different amounts of plasmid DNA.…”

Section: Confirming Protein Production and Incorporationmentioning

confidence: 80%

Stoichiometric parameters of HIV-1 entry

Zarr

Siliciano

2015

Virology

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During HIV type 1 (HIV-1) entry, trimers of gp120 bind to CD4 and either the CCR5 or CXCR4 coreceptor on the target cell. The stoichiometric parameters associated with HIV-1 entry remain unclear. Important unanswered questions include: how many trimers must attach to CD4 molecules, how many must bind coreceptors, and how many functional gp120 subunits per trimer are required for entry? We performed single round infectivity assays with chimeric viruses and compared the experimental relative infectivity curves with curves generated by mathematical models. Our results indicate that HIV-1 entry requires only a small number of functional spikes (one or two), that Env trimers may function with fewer than three active subunits, and that there is no major difference in the stoichiometric requirements for CCR5 vs. CXCR4 mediated HIV-1 entry into host cells.

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Membrane Fusion Mediated by Human Immunodeficiency Virus Envelope Glycoprotein

Melikyan

2011

Current Topics in Membranes

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Complementation of diverse HIV-1 Env defects through cooperative subunit interactions: a general property of the functional trimer

Cited by 19 publications

References 61 publications

Intraprotomer masking of third variable loop (V3) epitopes by the first and second variable loops (V1V2) within the native HIV-1 envelope glycoprotein trimer

Intraprotomer masking of third variable loop (V3) epitopes by the first and second variable loops (V1V2) within the native HIV-1 envelope glycoprotein trimer

Stoichiometric parameters of HIV-1 entry

Membrane Fusion Mediated by Human Immunodeficiency Virus Envelope Glycoprotein

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