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The terminal fragments (EJ and EM) of the XbaI-E transforming segment of human cytomegalovirus can independently induce the tumorigenic conversion of immortalized cells. To study their interaction, Rat-2 cells were transfected singly or with a combination of cloned EJ and EM DNAs. Large transformed foci were induced at a 10-fold higher frequency by EJ plus EM than by either DNA fragment alone. Focus-derived lines transformed by EJ plus EM produced tumors in syngeneic rats at a much faster rate (5 to 7 days) than did cell lines transformed by EJ or EM alone (25 to 35 days). Southern hybridizations showed that EM-homologous DNA was retained, exhibiting a complex pattern of multiple and amplified bands in EJ-plus-EM lines compared to a simple pattern in EM-induced lines. EJ DNA was not detected in the single or double transformants. The levels of p29, a 29-kilodalton transformation-sensitive marker in Rat-2 cells, were decreased 10to 100-fold in cell lines transformed by EJ or EM fragment alone. Synthesis of p29 was shut off in EJ-plus-EM transformants. These data demonstrate that two unlinked transforming regions of human cytomegalovirus can cooperate to produce an aggressive tumorigenic phenotype.

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