We have analyzed the expression of the three major known growth transformation-associated Epstein-Barr virus (EBV) proteins, EBNA-1, EBNA-2, and latent membrane protein (LMP), in a series of somatic cell hybrids derived from the fusion of EBV-carrying Burkitt lymphoma (BL) lines with EBV-positive or EBV-negative B-cell lines. Independently of the cell phenotype, EBNA-1 was invariably coexpressed in all EBV-carrying hybrids. In hybrids between EBV-carrying, LMP-positive and LMP-negative Burkitt lymphoma lines, LMP was expressed, indicating positive control. Two EBV-negative lymphoma lines, Ramos and BJAB, differed in their ability to express LMP after B95-8 virus-induced conversion and after hybridization with Raji cells. BJAB was permissive while Ramos was nonpermissive for LMP, although both expressed EBNA-2. The EBNA-2-deleted P3HR-1 virus gave the same pattern of LMP expression in these two cells. Our findings indicate that the expression of EBNA-1, EBNA-2, and LMP is regulated by independent mechanisms.