Cardioviral internal ribosomal entry site is functional in a genetically engineered dicistronic poliovirus

Abstract

HIGH mutation rates have driven RNA viruses to shorten their genomes to the minimum possible size¹. Mammalian (+)-strand RNA viruses and retroviruses have responded by reducing the number of cis-acting regulatory elements, a constraint that has led to the emergence of the polyprotein². Poliovirus is a (+)-stranded picornavirus whose polyprotein, encoded by an open reading frame spanning most of the viral RNA³, is processed by virus-encoded proteinases^4,5. Despite their genetic austerity, picor-naviruses have retained long 5' untranslated regions^6–8, which harbour cis-acting elements that promote initiation of translation independently of the uncapped 5' end of the viral messenger RNA^9–12. These elements are termed 'internal ribosomal entry sites'¹⁰ and are formed from highly structured RNA segments^13–15 of at least 400 nucleotides¹⁶. How these elements function is not known, but special RNA-binding proteins may be involved¹⁷. The ribosome or its 40S subunit probably binds at or near a Y_nX_m AUG motif (where Y is a pyrimidine and X is a purine) at the 3' border of the internal ribosomal entry site¹⁷, which either provides the initiating codon^16,18 or enables the ribosome to translocate to one downstream (E.W. et al., submitted). Initiation from most eukary-otic messenger RNAs usually occurs by ribosomal recognition of the 5' and subsequent scanning to the AUG codon¹⁹. Here we describe a genetic strategy for the dissection of polyproteins which proves that an internal ribosomal entry site element can initiate translation independently of the 5' end.