Epstein-Barr virus latency in vivo and in vitro

In this thesis, I studied 1) The latency situation of Epstein- Barr virus (EBV) in bone marrow transplanted (BMT) patients and healthy virus carriers. 2) The role of EBNA1 and cellular transcriptional factors Oct and Grg/TLE family in EBV latency switch. B lymphocytes have been identified as the main reservoir of latent Epstein-Barr virus (EBV) in healthy virus carriers. We have established a semi-quantitative PCR method (SQ-PCR) to estimate the EBV genome load in the blood B-cell subpopulation in healthy individuals. EBV DNA was detected in subfractionated IgM-, IgG- and IgA-positive B cells. Between 80%. and 90% of the viral DNA was found in the lgA-positive compared with the lgA-negative fraction. We used SQ-PCR to monitor the blood levels of EBV-DNA in 9 patients receiving allogeneic BMT. Four of 5 recipients of HLA-mismatched T-cell-depleted grafts showed a 4- to 5-log increase of EBV-DNA within 1 to 3 months after BMT. Administration of 2 to 4 infusions of 107 EBV-specific cytotoxic T-lymphocytes (CTLs)/M2 starting from the time of maximal virus load resulted in a 2- to 3-log decrease of virus titers in 3 patients. A moderate increase of virus titers was also detected in 3 of 4 patients receiving unmanipulated HLA-matched grafts, whereas one patient with Wiskott-Aldrich syndrome (WAS) reached a 5-log increase of EBVDNA load within 70 days after BMT. Our results suggest that a rapid increase of circulating EBV-DNA occurs in the absence of EBV-specific T-cell precursors or in the presence of congenital immune defects that prevent the reestablishment of virus-specific immunity. Prophylactic administration of EBV-CTI-s early after 13MT appears to provide the most effective protection against the development of EBV-associated lymphoproliferative disease. We identified that Cp could also can be activated by octamer-binding factor (OCT) proteins. Physical binding to the FR by the cellular transcription factors OCT1 and OCT2 was demonstrated by using electrophoretic mobility shift assay (EMSA). Furthermore, OCT2 alone or OCTI in combination with co-regulator Bob-1could drive transcription of a heterologous thymidine kinase promoter liked to the FR both in B cells and epithelial cells. Cp controlled by the FR also activated by binding of OCT2 to FR. OCT proteins can recruit the Grg/TLE - farmly of transcription factor regulatory proteins (co-factors) and those proteins repress OCT2 promoter activity through FR. Although all Grg/TLE variants could repress the