EBV Conference 2008 Guangzhou - Baylor College of Medicine

8 (RegID:1079; 1080; 1081)
Goran Gregorovic
Institution: Imperial College London
e-mail: g.gregorovic@imperial.ac.uk
CELL GENE REGULATION ASSOCIATED WITH EBER EXPRESSION
Goran Gregorovic, Daniel Charnock, Oliver Dittrich-Breiholz, Michael Kracht, Rainer Russ and Paul J.
Farrell
Oralabstract:
Microarray gene expression profiling has been used to investigate cell genes whose expression correlates
with expression of the Epstein-Barr virus EBER RNAs. New EBV mutants with deletions of EBER1 or
EBER2 were constructed using BAC mutagenesis in E. Coli. The EBV-BACs were transferred to 293
cells and multiple cell lines containing wild type B95-8 EBV BAC or the EBER1 or EBER2 deletion
mutants were cloned and characterised. RNA from 3 or 4 replica cell lines of each was used in
microarray gene expression profiling. Bioinformatic analysis of the data showed that the patterns of cell
gene expression of the wt and mutant virus lines could be distinguished by cluster analysis. Small
numbers of genes reproducibly differed in expression by greater than 5 fold in the different groups of cell
lines.
EBV lytic replication was induced in the 293 cell lines and the resulting virus (which carries a gfp marker)
was titred in Raji cells, counting gfp positive cells. Equal titres of the virus stocks were used to infect
primary human B cells isolated from peripheral blood or tonsils and lymphoblastoid cell lines (LCLs)
were readily obtained from the EBV mutants and wt virus. No substantial difference in the efficiency of
production of LCLs between the EBER mutants or wt BAC virus was observed in the conditions used.
RNA from the LCLs was used for further microarray gene expression profiling and the results were
compared to the 293 cell data. A panel of 30 genes selected from the microarray analysis was tested for
expression by RT-PCR and the differential expression of RNA according to EBER expression in the LCL
or 293 cell lines was confirmed. The results will be discussed in relation to potential roles for the EBER
RNAs in EBV infection.6
EBV Conference 2008 Guangzhou
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