ИСТИНА

Two principal modes of translation initiation in eukaryotic cells are cap-dependent scanning mechanism and internal ribosome entry. It is commonly suggested that a single mRNA predominantly uses only one of these mechanisms. Here we desсribe the case of an mRNA that can be translated by the both, and simple changes of the mRNA composition affects their contribution into the overall translation efficiency of the mRNA. Mechanism of Hepatitis C mRNA translation initiation is one of the most thoroughly studied examples of Internal Ribosome Entry Sites (IRESes). HCV IRES covers nucleotides 42-342 of the virus mRNA, but how sequences laying outside this region influence IRES activity remains a controversial issue. Here, using reporter monocistronic mRNAs we show that deletion of nts. 1-42 stimulates translation 2-4 times both in vitro and in vivo. We found that the 5'-truncated HCV IRES (nts. 42-342) has the same affinity to the 40S ribosomal subunit or eIF3 as the wild-type IRES. Interestingly, translation directed by the complete HCV 5' UTR can be stimulated by m 7 G-capping, while that of the 5'-truncated IRES variant can't be. We found that the 5'-cap stimulates translation of any HCV mRNA variant if it only has any native or artificial sequence upstream of domain II of the HCV IRES. In order to explain the fact of different cap-dependence we discriminate between the two modes of translation initiation by introducing an additional in-frame AUG codon into the HCV coding region downstream from the authentic one, and by creating the mutated IRES variants. In accordance with our expectations, the IRES directs translation from the authentic AUG only, while the 5'-end initiation mode contributes to the translation from both AUGs, irrespectively of the presence of the 5'-cap.