DIRECT BRLF1 BINDING IS REQUIRED FOR COOPERATIVE BZLF1 BRLF1 ACTIVATION OF THE EPSTEIN-BARR-VIRUS EARLY PROMOTER, BMRF1/

Disruption of Epstein - Barr virus (EBV) latency is mediated through t he activation of the viral immediate-early proteins, BZLF1 (Z) and BRL F1 (R).i.; (Chevallier-Greco, A., et al., (1 986) EMBO J., 5, 3243 - 9 ; Countryman, and Miller, G. (1985) Proc. Natl. Acad. Sci. USA, 82, 40 85 - 4089). We have previously demonstrated that these proteins cooper atively activate the EBV early promoter BMRF1 in lymphoid cells but no t in epithelial cells. Although cooperative transactivation by these p roteins has been demonstrated with a number of EBV promoters, the mech anism of this interaction is not well understood. We now show that the cooperative activation of the BMRF1 promoter by Z-plus-R requires an intact R binding site and at least one functional Z response element ( ZRE). Despite the presence of an R binding site, the BMRF1 promoter is only moderately responsive to R alone in either HeLa or Jurkat cells. Efficient activation of the BMRF1 promoter by Z alone in HeLa cells r equires two ZREs (located at - 59 and - 106), whereas two additional Z binding sites (located at - 42 and - 170) contribute very little to Z -induced activation. In the absence of ZREs, Z acted as a repressor of R-induced transactivation. These observations, along with observation s made by other investigators (Giot, J.F. et al., (1991) Nucleic Acids Res., 19, 1251 - 8), suggest that Z-plus-R cooperative activation is dependent upon 1) direct binding by R and Z to responsive promoter ele ments and 2) contributions by cell-specific factors.