Inhibition of Epstein-Barr virus replication by small interfering RNA targeting the Epstein-Barr virus protease gene.
BACKGROUND
The Epstein-Barr virus (EBV) protease (PR), coded by the BVRF2 gene, is essential for the maturation of the viral capsid and viral DNA packaging during the late stage of the EBV lytic cycle. Like the other herpesvirus serine PRs, EBV PR could be a target for the inhibition of EBV replication. To date, no data have been reported on the inhibition of EBV PR messenger RNA (mRNA) by small interfering RNA (siRNA).


METHODS
In this study, siRNAs targeting EBV PR were delivered to the epithelial 293 cell line stably transfected with the complete B95-8 EBV episome. EBV DNA and PR mRNA were quantified by real-time PCR in cells and supernatant, protein expression was assessed by immunoblotting, and production of EBV infectious particles in the culture medium was measured by Raji cell superinfection.


RESULTS
The EBV PR mRNA within the cells was reduced by 73%, the PR protein by 35% and the amount of virus in the cell supernatant was drastically decreased by 86% or 95%, depending on the method.


CONCLUSIONS
The strong effect of the siRNA targeting EBV PR on EBV replication attests to the crucial role played by EBV PR in the production of infectious particles and suggests that targeting this enzyme can be a new strategy against EBV-associated diseases where virus replication occurs.
Larrat
Sylvie
S
UMI 3265, UJF-EMBL-CNRS, Unit of Virus Host Cell Interactions, Grenoble, France.
Morand
Patrice
P
Bas
Ariane
A
Vigne
Solenne
S
Crance
Jean-Marc
JM
Boyer
Véronique
V
Nicod
Sandrine
S
Grossi
Laurence
L
Buisson
Marlyse
M
Burmeister
Wim P
WP
Seigneurin
Jean-Marie
JM
Germi
Raphaële
R
D002460:Cell Line; D015967:Gene Expression Regulation, Viral; D004854:Herpesvirus 4, Human / Q000187:drug effects / Q000201:enzymology / Q000235:genetics / Q000502:physiology*; D006801:Humans; D010447:Peptide Hydrolases / Q000235:genetics / Q000378:metabolism*; D012333:RNA, Messenger / Q000096:biosynthesis / Q000378:metabolism; D034741:RNA, Small Interfering / Q000378:metabolism / Q000494:pharmacology; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014764:Viral Proteins / Q000235:genetics / Q000378:metabolism; D014779:Virus Replication / Q000187:drug effects*
D012333:RNA, Messenger; D034741:RNA, Small Interfering; D014764:Viral Proteins; D010447:Peptide Hydrolases