Occluding the mannose moieties on human immunodeficiency virus type 1 gp120 with griffithsin improves the antibody responses to both proteins in mice

Abstract

To assess the influence of mannosylated glycans on the immunogenicity of human immunodeficiency virus type 1 (HIV-1) Env proteins, we immunized mice with monomeric gp120 in the presence and absence of the mannose-binding protein, griffithsin (GRFT). For comparison, other groups of mice received the nonglycosylated HIV-1 Gag protein, with and without GRFT. Coimmunization with GRFT increased the anti-gp120 IgG reactivity significantly, but had no effect on the anti-Gag response. We also investigated the IgG response to GRFT and found that gp120, but not Gag, enhanced its immunogenicity. For both proteins, IgG1 antibodies dominated the IgG response, with IgG2b as the next most prevalent subclass. We conclude that gp120-GRFT complexes are more immunogenic than the free proteins, for both components, and that occluding the mannose moieties on monomeric gp120 can improve the humoral immune response to this protein.

FIG. 1.

Partial block of gp120 binding to human DC-SIGN by griffithsin (GRFT). The binding to gp120 of HIV-Ig, 2G12, and DC-SIGN-Fc (upper left panel), 17b±sCD4 (upper right panel), VRC01, b12, and CD4-IgG2 (lower left panel), and 39F and PA1 (lower right panel) in the presence of varied concentrations of GRFT (x-axis) is normalized to that in the absence of GRFT (defined as 100%). The data are representative of two to three experiments. The means±SEM of two replicates in one ELISA are shown.

FIG. 2.

Effect of GRFT on the IgG responses to gp120 and Gag. The anti-gp120 (A) and anti-Gag (B) IgG responses are shown for three time points after immunization as background-corrected OD₄₉₀ values. The animals received either gp120 or Gag with or without GRFT, as indicated. The data points represent the means±SEM for the five mice in each group.

FIG. 3.

Isotype profile of the IgG response to gp120 and Gag in the presence and absence of GRFT. The IgG1, IgG2a, IgG2b, and IgG3 subclass responses at week 6 are shown as background-corrected OD₄₉₀ values. (A) Anti-gp120; (B) anti-Gag. The data points represent the means (±SEM) for the five mice in each group. Note that the scales on the y-axes differ between the diagrams for IgG1, which is the dominant IgG subclass, and the minor subclasses, IgG2a, IgG2b, and IgG3.

FIG. 4.

IgG responses to GRFT in mice coimmunized with gp120 or Gag. The anti-GRFT IgG endpoint titers at the indicated time points were analyzed by GRFT-GFP direct-coating assay. The median values among the mice are indicated by horizontal bars.

FIG. 5.

IgG subclass responses to GRFT in mice coimmunized with gp120 or Gag. The anti-GRFT IgG endpoint titers were analyzed by direct-coating GFP-GRFT ELISA. Only results from week 8, when total IgG reactivity with GRFT was strongest, are shown.

FIG. 6.

Antigenic basis for enhanced IgG responses in gp120–GRFT coimmunized mice. Recognition of gp120–GRFT complexes by week-6 sera from mice immunized with gp120 alone (left panel) or with gp120 plus GRFT (right panel). The data points show average OD₄₉₀ after background subtraction±SEM as a function of log serum concentration. Serum reactivity with gp120 captured alone is represented by gray squares and with gp120–GRFT complexes by black triangles. Note that the scales on the y-axes differ in order to demonstrate the variations within the weaker responses (left-hand diagram, immunization without GRFT) and the stronger ones (right-hand diagram, immunization with GRFT).