Epstein-Barr virus nuclear antigen 1, Oct and Groucho/TLE in control of promoter regulation

Abstract: The human herpes virus Epstein-Barr virus (EBV) is carried by approximately 95% of the adult population. It can form a lifelong latent infection in the B-cells by avoiding our immune system. A latent EBV infection is usually asymptomatic but a few EBV associated cancer forms has been described (e.g. Burkitt's lymphoma (BL)). EBV can avoid the immune system by restricting expression of its latent genes. Based on variation in viral gene expression several different latency forms have been described, e.g. latency 111 where all latency genes are expressed and latency 1 (e.g. BL) where only the nuclear protein EBNA l is expressed. EBNA l has three important functions, replication and maintenance of the viral genome and to regulate transcription. In latency 111 cells EBNA l is transcribed from the C promoter (Cp) together with six other proteins (EBNA l -6). The more restricted gene expression pattern in latency 1 cells is associated with a down regulation of the Cp and activity from the Q promoter (Qp) which leads to selective EBNA1 gene transcription. A switch between Cp and Qp usage might be instrumental in driving the host cell between latency forms. This thesis concerns the upstream control of Cp regulation with an emphasis on the interplay between cellular transcription factors and viral proteins. An EBNA1 responsive element called Family of Repeats (FR) is situated upstream of the Cp. EBNA1 can bind to each of the 20 repeats found in FR, thereby activating transcription. We have used various methods to show that the two cellular transcription factors Oct-1 and Oct-2 can bind to the FR sequence in vivo and in vitro and that the binding varies between the different repeats. We also show that binding has an impact on promoter activity in which Oct-2 alone and Oct-1 together with Bob.1 can substitute for the effect of EBNA1 on FR or further enhance the effect of EBNA1. We also describe the finding of a corepressor for Oct-proteins, namely the cellular protein Grg/TLE. The repression was shown to be highly dependent on the sequence to which Oct binds. This finding also applied to FR in EBV. All full-length Grg/TLE proteins as well as the truncated version Grg-5 can repress FR dependent Oct-2 activity. Binding of Grg/TLE to FR was also demonstrated both in vivo and in vitro. Repression by Grg/TLE could be cancelled by EBNA l, as well as the EBNA l induced activity could be repressed by Oct-2 + Grg/TLE.