Functional antagonism between the retinoic acid receptor and the viral transactivator BZLF1 is mediated by protein-protein interactions.
The Epstein-Barr virus-encoded protein BZLF1 is a member of the basic leucine zipper (bZip) family of transcription factors. Like several other members of the bZip family, transcriptional activity of BZLF1 is modulated by retinoic acid receptors (RARs). We present evidence that the RAR alpha and BZLF1 can reciprocally repress each other's transcriptional activation by a newly discovered mechanism. Analysis of RAR alpha mutants in transfection studies reveals that the DNA binding domain is sufficient for inhibition of BZLF1 activity. Analysis of BZLF1 mutants indicates that both the coiled-coil dimerization domain and a region containing the transcriptional activation domain of BZLF1 are required for transrepression. Coimmunoprecipitation experiments demonstrate physical interactions between RAR alpha and BZLF1 in vivo. Furthermore, glutathione S-transferase-pulldown assays reveal that these protein-protein interactions are mediated by the coiled-coil dimerization domain of BZLF1 and the DNA binding domain of RAR alpha. While RAR alpha is unable to recognize BZLF1 binding sites, the RAR alpha can be tethered to the DNA by forming a heteromeric complex with BZLF1 bound to DNA. Tethering RARs via protein-protein interactions onto promoter DNA suggest a mechanism through which RARs might gain additional levels of transcriptional regulation.
Pfitzner
E
E
Institut für Experimentelle Krebsforschung, Klinik für Tumorbiologie, Universität Freiburg, Germany.
Becker
P
P
Rolke
A
A
Schüle
R
R
D016475:3T3 Cells; D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D002460:Cell Line; D002522:Chlorocebus aethiops; D004268:DNA-Binding Proteins / Q000096:biosynthesis / Q000302:isolation & purification / Q000378:metabolism*; D005982:Glutathione Transferase / Q000096:biosynthesis; D004854:Herpesvirus 4, Human / Q000235:genetics / Q000378:metabolism; D015151:Immunoblotting; D051379:Mice; D008969:Molecular Sequence Data; D018168:Receptors, Retinoic Acid / Q000096:biosynthesis / Q000302:isolation & purification / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000302:isolation & purification / Q000378:metabolism; D000072482:Retinoic Acid Receptor alpha; D015534:Trans-Activators / Q000096:biosynthesis / Q000302:isolation & purification / Q000378:metabolism*; D015533:Transcriptional Activation; D014162:Transfection; D014764:Viral Proteins / Q000378:metabolism
C063676:BZLF1 protein, Herpesvirus 4, Human; D004268:DNA-Binding Proteins; C000607071:RARA protein, human; C000607072:Rara protein, mouse; D018168:Receptors, Retinoic Acid; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; D000072482:Retinoic Acid Receptor alpha; D015534:Trans-Activators; D014764:Viral Proteins; D005982:Glutathione Transferase