The application of an Epstein-Barr Virus specific antisense ribozyme for the in vitro suppression of EBNA-1 and LMP-1 expression
The application of an Epstein-B~lrr virus specific antisense ribozyme for the in vitro suppression of EBNA-l and LMP-l expression
Cheung Mei Sze
For the degree of Master of Philosophy at the University of Hong Kong
in March 2002
The Epstein Barr Virus (EBV) is a member of the human herpesvirus family and is one of the few naturally occurring human tumor viruses. It is associated with many human malignancies and leads to an uncontrollable proliferation of the infected cells. Two EBV encoded proteins, EBV nuclear antigen 1 (EBNA-l) and EBV encoded latent membrane protein 1 (LMP-l), are essential for the proliferation and immortalization of the infected cells. While EBNA-l is expressed in all EBV-infected cells and is important for the replication of the viral genome, LMP-I is expressed in some BBY-infected cells that are oncogenic. LMP-I is one of the most important mediators in EBV -induced cell proliferation and plays an essential role in cell immortalization. Thus, the downregulation of EBNA-I or LMP-l in EBY-immortalized cells may inhibit EBY related malignancies. Ribozyme is a small RNA molecule that is able to catalyze the cleavage of targeted RNA at specific sites. Earlier studies have reported its use to inhibit gene expression in various model systems. In this study, antisense ribozyme is used as a paradigm to down-regulate the expression ofEBNA-l and LMP-I in EBNA-l and LMPI expressing cell lines respectively. Two EBNA-l antisense ribozyme constructs and three LMP-l antisense ribozyme constructs were produced according to the thermodynamic folding of their mRNAs. Transfection of the ribozymes into an EBNA-l expressing embryonic kidney cell line 293c18 and a LMP-l expressing rat embryonic fibroblast cell line REF-LMP-l were performed to test the function of the ribozymes.
Obvious down-regulation of EBNA-I and LMP-l expressIOns were observed in the ribozyme-transfected cell lines. The suppression of EBNA-l in ceUline 293c18 led to a decrease in cell proliferation rate as well. These studies demonstrated the ability of the EBNA-l and LMP-l specific antisense ribozymes to down-regulate the expressions of EBNA-l and LMP-l. These ribozyme may be useful as therapeutic agents in the antiviral therapy of EBV in the future.
School:The University of Hong Kong
School Location:China - Hong Kong SAR
Source Type:Master's Thesis
Keywords:epstein barr virus genetics antisense dna catalytic rna genetic regulation gene expression
Date of Publication:01/01/2003