YY1 BINDS TO AND REGULATES CIS-ACTING NEGATIVE ELEMENTS IN THE EPSTEIN-BARR-VIRUS BZLF1 PROMOTER

A 48-bp cis-acting negative element in the Epstein-Barr virus BZLF1 ge ne P1 promoter has been described previously. By DNase I footprinting experiments, two regions were identified as the protein-binding sites (previously designated site I and Site II). In this report, the cellul ar transcription factor YY1 has been identified as a protein which bin ds to both of these elements, now designated ZIVA and ZIVB. Both ZIVA and ZIVB conferred cis-acting negative regulation on an enhancerless s imian virus 40 promoter. In cotransfection experiments, overexpression of YY1 caused further repression of the enhancerless simian virus 40 promoter containing either the ZIVA or ZIVB element. Cotransfection of a plasmid expressing antisense to YY1 increased the expression of the heterologous promoter containing ZIVA but not ZIVB. In similar experi ments carried out with the P1 promoter, overexpression of YY1 caused d ownregulation of P1 whereas antisense RNA to YY1 caused a slight incre ase in expression. Analyses of various p1 mutant constructions reveale d additional YY1 sites downstream of ZIVB. Overexpression of YY1 also caused downregulation of a P1 mutant with no apparent YY1-binding site s. TPA treatment of Raji cells caused a temporal loss of YY1-binding a ctivity but had no effect on the intracellular levels of YY1 protein. Serum induction of quiescent B cells also caused loss of YY1 binding t o the ZIVB site, which was found to be a weak serum response element. In contrast, anti-immunoglobulin G treatment of Akata cells had no eff ect on either the YY1-binding activity or protein levels. The binding of YY1 to the cis-acting negative elements in infected B cells may pla y a pivotal role in the maintenance of Epstein-Barr virus latency.