SEQUENCES WITHIN THE POLIOVIRUS INTERNAL RIBOSOME ENTRY SEGMENT CONTROL VIRAL-RNA SYNTHESIS

The 5' untranslated region of poliovirus RNA has been reported to poss ess two functional elements: (i) the 5' proximal 88 nucleotides form a cloverleaf structure implicated in positive-strand RNA synthesis duri ng viral replication, and (ii) nucleotides 134 to at least 556 functio n as a highly structured internal ribosome entry segment (IRES) during cap-independent, internal initiation of translation. We show here tha t the IRES itself is bifunctional and contains sequences necessary for viral RNA synthesis per se. For this purpose, we used a dicistronic p oliovirus RNA in which the translation of the viral non-structural (re plication) proteins is uncoupled from the poliovirus IRES. In this sys tem, RNA synthesis is readily detectable in transfected cells, even wh en the poliovirus IRES is inactivated by point mutation. However, dele tion of the major part of the poliovirus IRES renders viral-specific R NA synthesis undetectable. Using the same system, we show that a three nucleotide deletion at position 500 in the 5' untranslated region dra stically affects both translation efficiency and RNA synthesis. Furthe rmore, disruption of the secondary structure of the IRES around nucleo tide 343 has minimal effects on IRES function, but dramatically reduce s viral RNA replication. Taken together, these results provide direct evidence that sequences essential for viral RNA synthesis are located in the 3' region of the poliovirus IRES.