Cap-independent translation conferred by the 5 ' leader of tobacco etch virus is eukaryotic initiation factor 4G dependent

The 5' leader of tobacco etch virus (TEV) genomic RNA directs efficient tra nslation from the naturally uncapped viral mRNA. Two distinct regions withi n the TEV 143-nucleotide leader confer cap-independent translation in vivo even when present in the intercistronic region of a discistronic mRNA, indi cating that the TEV leader contains an internal ribosome entry site (IRES). In this study, the requirements for TEV IRES activity were investigated. T he TEV IRES enhanced translation of monocistronic or dicistronic mRNAs in v itro under competitive conditions, i.e., at high RNA concentration or in ly sate partially depleted of eukaryotic initiation factor 4F (eIF4F) and eIFi so4F, the two cap binding complexes in plants. The translational advantage conferred by the TEV IRES under these conditions was lost when the lysate r educed in eIF4F and eIFiso4F was supplemented with eIF4F (or, to a lesser e xtent, eIFiso4F) but not when supplemented with eIF4E, eIFiso4E, eIF4A, or eIF4B. eIF4G, the large subunit of eIF4F, was responsible for the competiti ve advantage conferred by the TEV IRES. TEV IRES activity was enhanced mode rately by the poly(A)-binding protein. These observations suggest that the TEV IRES directs cap-independent translation through a mechanism that invol ves eIF4G specifically.