ИСТИНА

There are two major modes of translation initiation in eukaryotic cells: the cap-dependent scanning mechanism and the mechanism of internal ribosome entry which does not require the cap. It is commonly suggested that a monocistronic mRNA predominantly uses only one of these mechanisms under certain conditions. Here we decsribe an mRNA that can be translated by both mechanisms, and with simple changes of the mRNA composition we are able to vary contribution of both mechanisms into the overall translation efficiency of the mRNA. The case of the Hepatitis C virus is one of the most studied examples of Internal Ribosome Entry Sites (IRESes). It is commonly accepted that HCV IRES covers nucleotides 42-342 of the HCV mRNA. However, issue of influence on IRES activity by sequences laying outside this region remains controversial. 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 m7G-capping, while that of the 5'-trancated 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 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. As a conclusion, we report the case of mRNA which is able to direct translation by the both 5'-end-dependent and IRES-dependent modes.