Sendai Virus Y Proteins Are Initiated by a Ribosomal Shunt

Abstract

The vast majority of eukaryotic mRNAs are monocistronic and express a single open reading frame (ORF) which initiates from the ATG codon nearest the capped 5Ј end. This initiation site is chosen by a scanning mechanism in which initiation factors, 40S ribosomal subunits, and initiator Met-tRNA bind to the capped 5Ј end of the mRNA and linearly scan the nucleotide sequence for the first start codon (the 5Ј scanning model; for a review, see reference 29). Some eukaryotic viral mRNAs, like the Sendai virus (SeV) P/C mRNA, however, express two ORFs which overlap (Fig. 1). Such mRNAs therefore also initiate proteins at start codons which are not 5Ј proximal. To account for initiation on these bicistronic mRNAs, a modified scanning model which allows for leaky scanning, i.e., scanning in which the scanning complex can bypass the first start codon to some extent when it is not in an optimal context for initiation and thereby can initiate at a downstream start codon, was proposed (23). This model followed from the realization that the nucleotide sequence surrounding the start codon (the context) was important for the efficiency with which ribosomes initiate protein synthesis, with the strongest determinant for efficiency being a purine at position Ϫ3, followed by a G at position ϩ4.Eukaryotic mRNAs known to initiate proteins at two start codons are nevertheless uncommon, and those that initiate at more than two start codons are rare. The most extreme example of an mRNA known to initiate proteins at multiple start sites remains the SeV P/C mRNA, which uses five start codons located 81 to 201 nucleotides (nt) from the 5Ј end and a start codon located more than 1500 nt from the 5Ј end to generate eight primary translation products (CЈ, P, C, Y1, Y2, V, W, and X) (Fig. 1). The second start site (and 5Ј-proximal ATG codon) generates three proteins (P, V, and W) containing the same N-terminal 317 amino acids (aa) but different C-terminal regions, as this mRNA is cotranscriptionally edited by the SeV polymerase via the programmed insertion of zero, one, or two G residues (45). The X protein, which represents approximately the C-terminal 95 aa of the 568-aa-long P protein, is thought to be initiated in a cap-dependent but scanning-independent manner (5). The first ribosomal start site (an unusual ACG start codon) and the third, fourth, and fifth start sites (all ATG codons) generate a nested set of four C proteins (termed CЈ, C, Y1, and Y2, of 215, 204, 183, and 175 aa, respectively) with a common C terminus, irrespective of whether the mRNA is edited (6,18,33). The Y proteins do not appear to be generated by proteolytic cleavage of the C or CЈ protein, because a recombinant SeV which does not express either C or CЈ (due to mutation of their start codons) overexpresses the Y proteins (25a). The available evidence suggests that whereas the first three start sites (for the CЈ, P, and C proteins) are accessed by leaky scanning, it is unlikely that the last two start sites (for the Y proteins) are accessed by a mechanism involvi...

Differential expression of Myc1 and Myc2 isoforms in cells transformed by eIF4E: evidence for internal ribosome repositioning in the human c-myc 5′UTR

“…There is no ®rm evidence to conclude that scanning is a linear, continuous process, as opposed to a discontinuous 5' to 3' migration and alignment of initiation complexes with structural elements along the mRNA 5'UTR. Indeed, a very similar mechanism, with very similar experimental evidence, was recently proposed for the translation of the Y Sendai virus proteins (Latorre et al, 1998), although this mechanism has never before been shown for the translation of a cellular mRNA, like c-myc.…”

Two Alternative Translation Mechanisms Are Responsible for the Expression of the HCV ARFP/F/Core+1 Coding Open Reading Frame

“…Different mechanisms that allow escape from an upstream initiator codon and direct initiation from internal codons have been described; these include context-dependent leaky-scanning (13,17,18,19,20,21), ribosome shunting (17,22,23,24,25,26,27), and IRES-mediated initiation (28). The internal initiation of ARFP/F/coreϩ1 translation in vivo might include features of one of these mechanisms.…”

Tethering of eIF4G to adenoviral mRNAs by viral 100k protein drives ribosome shunting

“…In CaMV, ribosome shunting is mediated by a short upstream open reading frame (ORF) and a downstream stable hairpin structure that together act to accumulate and reinitiate 40S ribosome subunits at a downstream ORF. Ribosome shunting has also been described for adenovirus (Ad) late mRNAs (Yueh andSchneider 1996, 2000), the Sendai virus Y mRNAs (Curran and Kolakofsky 1988;Latorre et al 1998;de Breyne et al 2003), papillomavirus E1 mRNA (Remm et al 1999), and possibly for several mammalian mRNAs (Yueh and Schneider 2000;Rogers et al 2004). The general mechanism for ribosome shunting in Ad and CaMV involves loading of 40S ribosome subunits onto the 5Ј end of the capped mRNA, entry of ribosome subunits a short distance into the mRNA, possibly by limited 5Ј scanning, followed by direct translocation of 40S subunits to a downstream initiation codon, directed by cisacting RNA shunting elements.…”