Rapid selection for an N-linked oligosaccharide by monoclonal antibodies directed against the V3 loop of human immunodeficiency virus type 1

One mechanism of immune evasion utilized by human immunodeficiency virus (HIV) and simian immunodeficiency virus (SIV) envelope glycoproteins is the presence of a dense carbohydrate shield. Accumulating evidence from in vitro and in vivo experiments suggests that alterations in N-linked glycosylation of SIV gp120 can enhance host humoral immune responses that may be involved in immune control. The present study was designed to determine the ability of glycosylation mutant viruses to redirect antibody responses to shielded envelope epitopes. The influence of glycosylation on the maturation and specificity of antibody responses elicited by glycosylation mutant viruses containing mutations of specific N-linked sites in and near the V1 and V2 regions of SIVmac239 gp120 was determined. Results from these studies demonstrated a remarkably similar maturation of antibody responses to native, fully glycosylated envelope proteins. However, analyses of antibodies to defined envelope domains revealed that mutation of glycosylation sites in V1 resulted in increased antibody recognition to epitopes in V1. In addition, we demonstrated for the first time that mutation of glycosylation sites in V1 resulted in a redirection of antibody responses to the V3 loop. Taken together, these results demonstrate that N-linked glycosylation is a determinant of SIV envelope B-cell immunogenicity in addition to in vitro antigenicity. In addition, our results demonstrate that the absence of N-linked carbohydrates at specific sites can influence the exposure of epitopes quite distant in the linear sequence.

mentioning

confidence: 68%

Removal of N-Linked Glycosylation Sites in the V1 Region of Simian Immunodeficiency Virus gp120 Results in Redirection of B-Cell Responses to V3

Cole¹,

Steckbeck

Rowles

et al. 2004

“…It has been reported previously that glycans shield antibody epitopes (29,32,36,45). Very recently, Wei et al (45) showed that NAbs directed towards HIV gp160 exert sufficient viral inhibitory activity to force the virus to replace the neutralization-sensitive virus by successive populations of resistant virus.…”

Section: Vol 78 2004 Hiv Resistance To Mannose-specific Plant Lectimentioning

confidence: 99%

Profile of Resistance of Human Immunodeficiency Virus to Mannose-Specific Plant Lectins

Balzarini

Laethem

Hatse

et al. 2004

The mannose-specific plant lectins from the Amaryllidaceae family (e.g., Hippeastrum sp. hybrid and Galanthus nivalis) inhibit human immunodeficiency virus (HIV) infection of human lymphocytic cells in the higher nanogram per milliliter range and suppress syncytium formation between persistently HIV type 1 (HIV-1)-infected cells and uninfected CD4 ؉ T cells. These lectins inhibit virus entry. When exposed to escalating concentrations of G. nivalis and Hippeastrum sp. hybrid agglutinin, a variety of HIV-1(III B ) strains were isolated after 20 to 40 subcultivations which showed a decreased sensitivity to the plant lectins. Several amino acid changes in the envelope glycoprotein gp120, but not in gp41, of the mutant virus isolates were observed. The vast majority of the amino acid changes occurred at the N glycosylation sites and at the S or T residues that are part of the N glycosylation motif. The degree of resistance to the plant lectins was invariably correlated with an increasing number of mutated glycosylation sites in gp120. The nature of these mutations was entirely different from that of mutations that are known to appear in HIV-1 gp120 under the pressure of other viral entry inhibitors such as dextran sulfate, bicyclams (i.e., AMD3100), and chicoric acid, which also explains the lack of cross-resistance of plant lectin-resistant viruses to any other HIV inhibitor including T-20 and the blue-green algae (cyanobacteria)-derived mannose-specific cyanovirin. The plant lectins represent a welldefined class of anti-HIV (microbicidal) drugs with a novel HIV drug resistance profile different from those of other existing anti-HIV drugs.

“…However, no reduction of CCR5 usage was observed when the same residue was mutated on SF13 and SF162 (M-tropic) (34) or ConB (M-tropic) (51). Additionally, the corresponding mutation had no effect on infectivity of other T-tropic stains (LAI, BRU, and HXB2 [2,27,43]). These apparent discrepancies are likely due to specific differences in amino acid sequences in the V3 loop for different isolates, which together with carbohydrate residues create a V3 loop …”

Section: Discussionmentioning

confidence: 96%

N-Linked Glycosylation Sites Adjacent to and within the V1/V2 and the V3 Loops of Dualtropic Human Immunodeficiency Virus Type 1 Isolate DH12 gp120 Affect Coreceptor Usage and Cellular Tropism

Ogert

Lee

Ross

et al. 2001

128

The envelope glycoprotein of human immunodeficiency virus type 1 (HIV-1) is extensively glycosylated, containing approximately 23 asparagine (N)-linked glycosylation sites on its gp120 subunit. In this study, specific glycosylation sites on gp120 of a dualtropic primary HIV-1 isolate, DH12, were eliminated by sitedirected mutagenesis and the properties of the resulting mutant envelopes were evaluated using a recombinant vaccinia virus-based cell-to-cell fusion assay alone or in the context of viral infections. Of the glycosylation sites that were evaluated, those proximal to the V1/V2 loops (N135, N141, N156, N160) and the V3 loops (N301) of gp120 were functionally critical. The glycosylation site mutations near the V1/V2 loop compromised the use of CCR5 and CXCR4 equally. In contrast, a mutation within the V3 loop preferentially inhibited the usage of CCR5; although this mutant protein completely lost its CCR5-dependent fusion activity, it retained 50% of the wild-type fusion activity with CXCR4. The replication of a virus containing this mutation was severely compromised in peripheral blood mononuclear cells, MT-4 cells, and primary monocyte-derived macrophages. A revertant virus, which acquired second site changes in the V3 loop that resulted in an increase in net positive charge, was isolated. The revertant virus fully recovered the usage of CXCR4 but not of CCR5, thereby altering the tropism of the parental virus from dualtropic to T-tropic. These results suggest that carbohydrate moieties near the V1/V2 and the V3 loops play critical roles in maintaining proper conformation of the variable loops for optimal interaction with receptors. Our results, combined with those of previously reported studies, further demonstrate that the function of individual glycans may be virus isolate dependent. Human immunodeficiency virus type 1 (HIV-1) primarily infects CD4ϩ T lymphocytes and cells of monocyte-macrophage lineage. The cellular tropism of HIV-1 is determined largely at the level of virus entry, which depends on a series of interactions between viral envelope glycoprotein and cellular receptors. The gp120 surface glycoprotein subunit is thought to interact first with CD4 (the primary receptor) and then with one (or more) of the recently identified coreceptors, which include chemokine receptors CXCR4 and CCR5 (for reviews, see references 15, 16, 26, and 32). These interactions are thought to trigger conformational changes in the complex multimeric viral envelope glycoprotein structure, allowing the hydrophobic domain of transmembrane glycoprotein gp41 subunit to interact with the cellular membrane and induce viruscell fusion.While the exact mechanism of membrane fusion is still unclear, the interactions between gp120 and cellular receptors are slowly beginning to be understood. The detailed molecular nature of the interactions between gp120 and CD4 has been elucidated from analysis of a crystal structure of gp120 core complexed with CD4 (two N-terminal domains) and the 17b neutralizing monoclonal antibody (Fab fra...