Epstein-Barr virus (EBV) is a ubiquitous human herpesvirus that persistently infects 85% of the adult population worldwide. In this report, we examine the proliferative response and cytokine secretion profile of CD4 ؉ T lymphocytes from a panel of unrelated EBV-positive donors against two EBV latent antigens, EBNA1 and EBNA3C. Substantial proliferative responses by CD4؉ lymphocytes were demonstrated to both antigens in multiple, randomly selected donors. Surprisingly, we observed a striking and consistent difference in cytokine response to EBNA1 and EBNA3C. EBNA1-specific CD4 ؉ T lymphocytes from multiple unrelated donors preferentially produced type 2-like cytokines in response to antigenic stimulation, while the response to EBNA3C was a characteristic type 1 response. The implications of these findings for EBV persistence and the development of EBV-associated malignancies are discussed.Epstein-Barr virus (EBV) is a ubiquitous human herpesvirus that infects more than 85% of the adult population worldwide and establishes a persistent infection for the lifetime of the host. The EBV carrier state (latency) is characterized by expression of a limited set of EBV genes and by sporadic reactivation of the lytic cycle in latently infected cells, leading to viral replication. Several lines of evidence implicate latently infected B lymphocytes as the major EBV reservoir. Upon reactivation in tonsillar B lymphocytes, EBV can productively infect oropharyngeal epithelium, resulting in infectious virus production and transmission (29,49).The importance of EBV as a human pathogen is evinced by its etiologic role in the infectious mononucleosis syndrome. The potential significance of EBV in malignancy is suggested by its B-cell-transforming ability in vitro and by its strong association with a number of human malignancies, including Burkitt's lymphoma, nasopharyngeal carcinoma, Hodgkin's disease, and immunoblastic lymphomas in immunocompromised patients. Despite the oncogenic potential of the virus, the vast majority of EBV-infected individuals remain asymptomatic. A considerable amount of evidence suggests that cell-mediated immune response, particularly CD8 ϩ cytotoxic T-lymphocyte (CTL) recognition of the EBV latent proteins in persistently infected cells, is critical in suppressing EBV replication in latently infected individuals (reviewed in references 49 and 50).Studies of EBV latent gene expression in persistently infected cells and EBV-positive tumors including both spontaneous and induced EBV-transformed B-lymphoblastoid cell lines (LCL) suggest that up to eight EBV proteins including six nuclear antigens (i.e., EBV nuclear antigen 1 [EBNA1], -2, -3A, -3B, -3C, and -LP) and two membrane proteins (latent membrane protein 1 [LMP1] and 2 [LMP2]) may be differentially expressed in cells exhibiting various forms of EBV latency. Significantly, EBNA1 is the only EBV latent antigen consistently expressed in all patterns of EBV latent gene expression, including EBV-positive malignancies; this reflects its essential function in...