We have investigated the possible role of -acting factors interacting with the untranslated regions (UTRs) of coxsackievirus B3 (CVB3) RNA. We show here that polypyrimidine tract-binding protein (PTB) binds specifically to both 5′ and 3′ UTRs, but with different affinity. We have demonstrated that PTB is a bona fide internal ribosome entry site (IRES) -acting factor (ITAF) for CVB3 RNA by characterizing the effect of partial silencing of PTB in HeLa cells. Furthermore, IRES activity in BSC-1 cells, which are reported to have a very low level of endogenous PTB, was found to be significantly lower than that in HeLa cells. Additionally, we have mapped the putative contact points of PTB on the 5′ and 3′ UTRs by an RNA toe-printing assay. We have shown that the 3′ UTR is able to stimulate CVB3 IRES-mediated translation. Interestingly, a deletion of 15 nt at the 5′ end or 14 nt at the 3′ end of the CVB3 3′ UTR reduced the 3′ UTR-mediated enhancement of IRES activity significantly, and a reduced interaction was shown with PTB. It appears that the PTB protein might help in circularization of the CVB3 RNA by bridging the ends necessary for efficient translation of the viral RNA.


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