Localization of CD4+ T cell epitope hotspots to exposed strands of HIV envelope glycoprotein suggests structural influences on antigen processing

Abstract

The spectrum of immunogenic epitopes presented by the H2-IA(b) MHC class II molecule to CD4(+) T cells has been defined for two different (clade B and clade D) HIV envelope (gp140) glycoproteins. Hybridoma T cell lines were generated from mice immunized by a sequential prime and boost regime with DNA, recombinant vaccinia viruses, and protein. The epitopes recognized by reactive T cell hybridomas then were characterized with overlapping peptides synthesized to span the entire gp140 sequence. Evidence of clonality also was assessed with antibodies to T cell receptor Valpha and Vbeta chains. A total of 80 unique clonotypes were characterized from six individual mice. Immunogenic peptides were identified within only four regions of the HIV envelope. These epitope hotspots comprised relatively short sequences ( approximately 20-80 aa in length) that were generally bordered by regions of heavy glycosylation. Analysis in the context of the gp120 crystal structure showed a pattern of uniform distribution to exposed, nonhelical strands of the protein. A likely explanation is that the physical location of the peptide within the native protein leads to differential antigen processing and consequent epitope selection.

Figures

Figure 1

The Th epitopes are located in distinct hotspots. The antigenic peptides defined in Table 1 are restricted to four distinct regions of the env sequence. The hotspots are identified as regionsA–D through the gp120 (A–C) and gp41 (D) sequence. Note that immunogen sequences initiated at the junction of env signal sequence (leader) and gp120 coding sequence.

Figure 2

Anatomical location of the hotspots to exposed, nonhelical loops and strands of the env protein. The Th hotspots are highlighted (red) on the crystal structure (34, 37) of gp120 (blue) complexed with CD4 (gray). The structure can be oriented by the position of the N- and C-terminal residues of gp120 and the CD4 binding site. The regionsA–C in this figure correspond toA–C in Fig. 1 with the omission of V2 and V3 sequences, absent from the crystal. The composite of all three hotspots is shown from front (D) and side (E) views.