Direct killing of Epstein-Barr virus (EBV)–infected B cells by CD4 T cells directed against the EBV lytic protein BHRF1

Abstract

We show here that T-cell responses to this particular BHRF1 epitope are shared by most EBV-infected DR*0401 ؉ individuals, as BHRF1-specific CD4 T cells could be sorted out from all the DRB*0401 T-cell lines analyzed, using magnetic beads coated with recombinant BHRF1/DR*0401 complexes. Sorting with these peptide MHC class II multimers was very efficient, as the yield of recovery of BHRF1-specific T cells was nearly 100%. Functional analysis of a large number of clones responding to BHRF1/DR*0401 demonstrated their cytolytic action against autologous and allogeneic DR*0401 ؉ EBVtransformed B-lymphoblastoid cell lines (B-LCLs), with 40% to 80% killing efficiency and potent interferon ␥ production, thus suggesting that this CD4 T-cell population contributes to the control of EBV replication. B-LCL lysis by these T-cell clones was DR*0401 dependent, EBV dependent, and was not merely due to bystander killing. Taken together, these data provide the first demonstration that a lytic antigen can induce a direct cytolytic response against EBV-infected cells.

Emergence of Simian Immunodeficiency Virus-Specific Cytotoxic CD4+ T Cells and Increased Humoral Responses Correlate with Control of Rebounding Viremia in CD8-Depleted Macaques Infected with Rev-Independent Live-Attenuated Simian Immunodeficiency Virus

“…It is now apparent that a subset of CD4 + T cells expresses both perforin and granzymes (59)(60)(61), and that these cells are capable of killing both in vitro and in vivo. Direct killing in vivo by CD4 + T cells has been shown in several viral infections, including lymphocytic choriomeningitis virus, EBV, and gammaherpesvirus (62)(63)(64), and the presence of CD4 + T cells with a cytotoxic phenotype has been described in HIV-1-infected individuals (65). In fact, studies with CD4 + T cells obtained from HIV-1-infected individuals and healthy controls demonstrated that such CD4 + T cells not only harbor granules containing perforin and granzyme B, but were also able to release them upon specific stimulation with CMV peptides, resulting in lysis of the target cells ex vivo (66).…”

Selection of high-avidity CD8 T cells correlates with control of hepatitis C virus infection

“…Protein-secretion inhibitor brefeldin-A (1 mM) was added after 2 hours' incubation. Intracellular staining assays were performed as previously described 18 with anti-IFN-␥ or anti-tumor necrosis factor alpha (TNF-␣) mAb (100 ng/mL, BD Pharmingen, Biosciences).…”

EBV-Specific CD4+ T Cell Clones Exhibit Vigorous Allogeneic Responses

“…Cytotoxicity was tested by standard 4-h chromium-51 release assay, at defined E:T ratios, as described previously (24). When used, synthetic peptides were directly added to chromium-51-labeled targets and incubated for 1 h before excess unbound peptide was washed off.…”

“…ϩ T cell clones EBV-specific CD4 ϩ T cell clones were derived from PBL from healthy virus carriers or from synovial fluid from patients suffering from rheumatoid arthritis, as described previously (24,25). T cell clones were maintained in RPMI 1640 supplemented with 10% pooled human serum, 1 mM L-glutamine, and 150 IU/ml rIL-2.…”

“…In contrast, there is little evidence on the influence of antiviral CD4 ϩ T cells on the alloreactive repertoire (22,23). In this study, we screened for cross-recognition of allogeneic EBV-transformed B cell lines (lymphoblastoid cell lines (LCLs)) by EBV-specific CD4 ϩ T cell clones (24,25) and report several examples of cross-reactivity leading to potent killing of allogeneic LCLs expressing a particular MHC class II allele.…”