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J. Virol., 05 1997, 3466-3473, Vol 71, No. 5
CJ Chen, MD Kuo, LJ Chien, SL Hsu, YM Wang and JH Lin
The mechanism of replication of the flavivirus Japanese encephalitis virus
(JEV) is not well known. The structures at the 3' end of the viral genome
are highly conserved among divergent flaviviruses, suggesting that they may
function as cis-acting signals for RNA replication and, as such, might
specifically bind to cellular or viral proteins. UV cross-linking
experiments were performed to identify the proteins that bind with the JEV
plus-strand 3' noncoding region (NCR). Two proteins, p71 and p110, from
JEV-infected but not from uninfected cell extracts were shown to bind
specifically to the plus-strand 3' NCR. The quantities of these binding
proteins increased during the course of JEV infection and correlated with
the levels of JEV RNA synthesis in cell extracts. UV cross-linking coupled
with Western blot and immunoprecipitation analysis showed that the p110 and
p71 proteins were JEV NS5 and NS3, respectively, which are proposed as
components of the RNA replicase. The putative stem-loop structure present
within the plus-strand 3' NCR was required for the binding of these
proteins. Furthermore, both proteins could interact with each other and
form a protein-protein complex in vivo. These findings suggest that the 3'
NCR of JEV genomic RNA may form a replication complex together with NS3 and
NS5; this complex may be involved in JEV minus-strand RNA synthesis.
Copyright © 1997, American Society for Microbiology
RNA-protein interactions: involvement of NS3, NS5, and 3' noncoding regions of Japanese encephalitis virus genomic RNA
Institute of Preventive Medicine, National Defense Medical Center, Taipei, Taiwan, Republic of China.
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