Sequence variation within the dominant amino terminus epitope affects antibody binding and neutralization of human immunodeficiency virus type 1 Tat protein

Abstract

Tat is among the required regulatory genes of human immunodeficiency virus type 1 (HIV-1). Tat functions both within infected cells as a transcription factor and as an extracellular factor that binds and alters bystander cells. Some functions of extracellular Tat can be neutralized by immune serum or monoclonal antibodies. In order to understand the antibody response to Tat, we are defining antibody epitopes and the effects of natural Tat sequence variation on antibody recognition. The dominant Tat epitope in macaque sera is within the first 15 amino acids of the protein amino terminus. Together with a subdominant response to amino acids 57 to 60, these two regions account for most of the macaque response to linear Tat epitopes and both regions are also sites for the binding of neutralizing antibodies. However, the dominant and subdominant epitope sequences differ among virus strains, and this natural variation can preclude antibody binding and Tat neutralization. We also examined serum samples from 31 HIV-positive individuals that contained Tat binding antibodies; 23 of the 31 sera recognized the amino terminus peptide. Similar to binding in macaques, human antibody binding to the amino terminus was affected by variations at positions 7 and 12, sequences that are distinct for clade B compared to other viral clades. Tat-neutralizing antibodies to the dominant amino terminus epitope are affected by HIV clade variation.

FIG. 1.

Peptide competition of Tat toxoid immunized monkey sera binding to Tat. Monkey sera at dilutions between 1:100 and 1:1,000 were preincubated with no peptide (♦) or peptides corresponding to the N-terminal region (▪), the basic domain (▴), both peptides (×), or scrambled peptides corresponding to both regions (•). The optical density at 405 nm (x axis) was measured after an ELISA on plates coated with an 86-amino-acid Tat protein. Animals are identified by number.

FIG. 2.

Peptide inhibition of Tat neutralization by monoclonal antibody TR1. The optical density at 450 nm (OD₄₅₀) was measured by a p24 ELISA 72 h after the addition of samples to a HeLa cell line with an integrated provirus lacking Tat.