Dimerization of the Epstein-Barr virus ZEBRA protein in the yeast two-hybrid system. Comparison of a ZEBRA variant with the B95-8 form

Epstein-Barr virus (EBV) is a herpes virus associated with several human tu mors. The EBV protein, ZEBRA, is a transactivator of the basic leucine zipp er family (bZip). It binds to specific sequences on DNA and is able to inte ract with cellular proteins such as p53. The interaction of the ZEBRA prote in with its cognate DNA sequences is stable as long as the dimerization dom ain is functional. Recent work from this laboratory identified a ZEBRA vari ant (Z206) with a single amino acid change at residue 206. An alanine is su bstituted for a serine, and this replacement is present in 72% of nasophary ngeal carcinoma from Europe and North Africa. As amino acid 206 lies within the dimerization domain it could be instrumental in interactions with othe r proteins. The yeast two-hybrid system was used to study ZEBRA-protein int eractions. As ZEBRA by itself is a transactivator in yeast, it cannot be us ed directly in this assay. This paper describes modifications in ZEBRA amin o acid sequences, rendering it usable in the yeast two-hybrid assay. We com pared the dimerization capacity of the Z206 variant to that of ZEBRA from B 95-8 (Z95) and observed that reporter gene activity with Z206 was consisten tly lower than that of Z95 (P < 0.05). Furthermore, no interaction was foun d to occur between either form of ZEBRA (Z206 or Z95) and the tumor suppres sor, p53 in the yeast two-hybrid system. (C) 2000 Societe francaise de bioc himie et biologie moleculaire / Editions scientifiques et medicales Elsevie r SAS.