CONTROL OF EBV LYTIC GENE EXPRESSION DURING LATENCY

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This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. The major goal of this research is the elucidation of the mechanisms controlling the switch from latency to viral replication in Epstein-Barr virus (EBV) infected B lymphocytes. More specifically, this research focuses on the regulation of two linked EBV genes, BSLF1 and BRLF1, which are integrally involved in this switch. A detailed understanding of how viral reactivation is controlled is essential for understanding maintenance of latency, and may reveal strategies for interfering with viral persistence in the host. During the pervious funding period, significant progress has been made in identifying the critical cis-elements involved in regulating the BZLF1 gene promoter (Zp), and the cellular factors that bind to these sites. In addition, two calcium response pathways have been identified that can trigger viral reactivation. However, we only have a partial picture of how transcription of the BZLF1 gene is regulated. Furthermore, regulation of the BRLF1 gene has not been carefully analyzed. Thus, to identify and further define the cis-elements, cellular transcription factors and signaling pathways involved regulating the BRLF1/BZLF1 gene locus we propose the following aims: Aim 1. Functional analysis of cis-elements involved in regulating ZP and RP: 1a. Investigate the interdependence of the BRLF1 and BZLF1 genes; 1b. Identify and characterize cis-elements involved in regulating induction of Rp; 1c. Characterize role of Sp1, Sp3 and MEF2D binding to the Z1 domains in Zp; 1d. Characterize and clone the cellular factor(s) binging to the negative cis-element ZIIR in Zp; Aim 2. Generation of EBV mutants in the BRLF1/BZLF1 locus: 2a. Characterize the role of Zp and Zta in the induction of the viral lytic cycle; 2b. Assess the impact of selected mutations in critical cis-elements involved in regulating Zp and Rp.