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. 2012 Jun;25(3):193-203.
doi: 10.1089/vim.2011.0094. Epub 2012 May 2.

Antibodies to the central conserved region of respiratory syncytial virus (RSV) G protein block RSV G protein CX3C-CX3CR1 binding and cross-neutralize RSV A and B strains

Youngjoo Choi  1 , Caleb S Mason, Les P Jones, Jackelyn Crabtree, Patricia A Jorquera, Ralph A Tripp
Affiliations

Antibodies to the central conserved region of respiratory syncytial virus (RSV) G protein block RSV G protein CX3C-CX3CR1 binding and cross-neutralize RSV A and B strains

Youngjoo Choi et al. Viral Immunol. 2012 Jun.

Abstract

Respiratory syncytial virus (RSV) is a primary cause of severe lower respiratory tract disease in infants, young children, and the elderly worldwide, and despite decades of effort, there remains no safe and effective vaccine. RSV modifies the host immune response during infection by CX3C chemokine mimicry adversely affecting pulmonary leukocyte chemotaxis and CX3CR1+ RSV-specific T-cell responses. In this study we investigated whether immunization of mice with RSV G protein polypeptides from strain A2 could induce antibodies that block G protein-CX3CR1 interactions of both RSV A and B strains. The results show that mice immunized with RSV A2 G polypeptides generate antibodies that block binding of RSV A2 and B1 native G proteins to CX3CR1, and that these antibodies effectively cross-neutralize both A and B strains of RSV. These findings suggest that vaccines that induce RSV G protein-CX3CR1 blocking antibodies may provide a disease intervention strategy in the efforts to develop safe and efficacious RSV vaccines.

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Figures

FIG. 1.
FIG. 1.
Schematic of RSV G protein and location of G polypeptides. (A) The transmembrane region and cytoplasmic domain are indicated by TM and CD, respectively. The cysteine loop region is indicated. The locations of the RSV G polypeptides on the G protein (i.e., G1, G2, and G3) are also depicted. (B) Amino acid sequence of G2 polypeptide (PPT) is shown. For the purpose of comparison, the amino acid sequence of the cysteine loop region of the RSV B1 G protein (aa 169–198) is also depicted. The CX3C motifs in the RSV A2 and B1 G proteins are underlined, and the non-homologous amino acid residues are highlighted in bold.
FIG. 2.
FIG. 2.
RSV G polypeptide-specific IgG antibodies are reactive against native RSV/A2 and RSV/B1 G proteins. Purified IgG antibodies from the serum of G polypeptide (PPT) G1, G2, G3, or UV-inactivated RSV/A2 mice were evaluated for reactivity to native RSV/A2 (A), or native RSV/B1 (B) G proteins. The results represent three independent assays with three replicates per dilution per assay±standard error of the mean.
FIG. 3.
FIG. 3.
Purified IgG antibodies from G2 polypeptide-vaccinated mice inhibit RSV/A2 and RSV/B1 native G-protein binding to CX3CR1. IgG antibodies purified from the sera of G polypeptide- or UV-inactivated RSV A2-vaccinated mice were examined for their ability to inhibit binding of purified RSV/A2 or RSV/B1 native G protein to 293-CX3CR1 cells. Data represent the percent inhibition of RSV/A2 native G protein (A) or RSV/B1 native G protein (B) binding to 293-CX3CR1 cells. RSV G-protein-specific monoclonal antibody (clone 131-2G), and IgG purified from naïve mouse sera, were positive and negative controls, respectively. The percent inhibition was calculated using the formula: [1 – (percent Alexa 488-positive of 293-CX3CR1 treated with the antibody mixture/percent Alexa 488-positive of 293-CX3CR1 treated with G protein only)] (*p<0.01 when compared with naïve mouse IgG). The results represent three independent assays with three replicates per dilution per assay±standard error of the mean.
FIG. 4.
FIG. 4.
Neutralization of RSV A2 and RSV B1 by IgG antibodies purified from sera of RSV G polypeptide-vaccinated mice. Purified RSV G polypeptide (G1, G2, or G3)-specific IgG antibodies were examined for their ability to neutralize RSV A2 (A) and RSV B1 (B) by plaque reduction assay. Vero E6 cells were infected with 102 PFU of live RSV A2 or B1 stain that was previously incubated with 1 μg per well concentration of various RSV G protein-specific IgG antibodies. At day 5 post-infection, the cells were fixed with acetone:methanol (60:40). The plaques were immunostained using RSV anti-F monoclonal antibody 131-2A and counted. RSV F protein-specific monoclonal antibody from clone 131-2A and PBS were used as positive and negative controls, respectively. Data are represented as the percent RSV plaque reduction, and the values are expressed as percent plaque reduction relative to the negative control (*p<0.01 compared with naïve mouse IgG).
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