In the present study, we have isolated CD4+ CTLs that recognize an epitope from EBV nuclear Ag 2 in association with two different HLA-DQ Ags, DQA1*0501/DQB1*0201 (DQ2) or DQA1*0501/DQB1*0301 (DQ7). Both the HLA-DQ2 and HLA-DQ7 alleles displayed a similar efficiency in the endogenous and exogenous presentation of this epitope. Since earlier studies have shown that the EBV-associated malignancy, Burkitt's lymphoma (BL), escapes class I-restricted immune recognition by down-regulating the expression of peptide transporter genes, we have explored the possibility that these tumor cells can process class II-restricted CTL epitopes. The data presented in this study clearly demonstrate that BL cells were recognized efficiently by CD4+, MHC class II-restricted EBV-specific CTLs following infection with recombinant vaccinia encoding EBV nuclear Ag 2. Analysis of surface MHC class II expression on BL cells revealed high levels of HLA-DR and HLA-DQ molecules, and most of these molecules were negative for the invariant chain peptide, referred to as CLIP. Moreover, these tumor cells also showed normal levels of HLA-DMB gene expression, which has been shown previously to be an essential component of the class II processing pathway. The present finding of efficient processing function through the class II pathway in BL cells provides a novel mechanism for immune targeting of EBV-positive malignancies.