Staphylococcus aureus superantigens elicit redundant and extensive human Vbeta patterns

Abstract

Staphylococcus aureus can produce a wide variety of exotoxins, including toxic shock syndrome toxin 1 (TSST-1), staphylococcal enterotoxins, and staphylococcal enterotoxin-like toxins. These toxins share superantigenic activity. To investigate the beta chain (Vbeta) specificities of each of these toxins, TSST-1 and all known S. aureus enterotoxins and enterotoxin-like toxins were produced as recombinant proteins and tested for their ability to induce the selective in vitro expansion of human T cells bearing particular Vbeta T-cell receptors (TCR). Although redundancies were observed between the toxins and the Vbeta populations, each toxin induced the expansion of distinct Vbeta subsets, including enterotoxin H and enterotoxin-like toxin J. Surprisingly, the Vbeta signatures were not associated with a specific phylogenic group of toxins. Interestingly, each human Vbeta analyzed in this study was stimulated by at least one staphylococcal superantigen, suggesting that the bacterium derives a selective advantage from targeting the entire human TCR Vbeta panel.

FIG. 1.

CD69 expression by T lymphocytes upon S. aureus SAg challenge. CD69 expression was measured on T lymphocytes (CD3⁺) after 24 h of incubation with 1.0 μg/ml of SSAgs (black bars) and 0.1 μg/ml of SSAgs (white bars) of whole blood, Eagle's minimal essential medium (negative control; RPMI [gray bar]), or 10 μg/ml PHA (positive control; gray bar). Results are means ± standard deviations (n = 3).

FIG. 2.

Human Vβ expansion induced by S. aureus SAgs as detected with the IOTest Beta Mark kit. Purified recombinant toxins and PHA were studied for their ability to induce the selective expansion of T cells bearing particular TCR Vβ regions in PBMC culture. Vβ profiles were determined by flow cytometry using the IOTest Beta Mark kit completed with a Vβ6.7 antibody. Results are expressed as ratios of the percentage of TCR Vβ expansion induced by each toxin relative to that of PHA. The x axis label indicates Vβs. The data shown here are the representative ratios observed with each of the three blood donors (□, ⋄, ○) plus the medians of these ratios (−), while the horizontal dashed line represents a ratio of 1.

FIG. 2.

Human Vβ expansion induced by S. aureus SAgs as detected with the IOTest Beta Mark kit. Purified recombinant toxins and PHA were studied for their ability to induce the selective expansion of T cells bearing particular TCR Vβ regions in PBMC culture. Vβ profiles were determined by flow cytometry using the IOTest Beta Mark kit completed with a Vβ6.7 antibody. Results are expressed as ratios of the percentage of TCR Vβ expansion induced by each toxin relative to that of PHA. The x axis label indicates Vβs. The data shown here are the representative ratios observed with each of the three blood donors (□, ⋄, ○) plus the medians of these ratios (−), while the horizontal dashed line represents a ratio of 1.

FIG. 3.

Reconstitution of the phylogenetic tree of S. aureus SAgs. Amino acid sequences of the mature toxins were deduced from sequences obtained from GenBank and SignalP analysis. Alignment was performed with ClustalX software. Evolutionary distances were determined by the Kimura method, and the values were used to construct a dendrogram by means of the neighbor-joining method using SplitsTree4 software. Vβ expansion induced by each toxin is indicated. Vβ expansions observed in response to several SSAg phylogenic groups are boldfaced.