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

eIF4E is essential for translation initiation, but its overexpression cause s malignant transformation. Recent work demonstrated that eIF4E/F participa tes in exposing and locating alternate translation start codons during scan ning. Translation initiation of several important protooncogenes and growth -regulators, such as Myc and FGF-2, can start at CUG start codon(s) upstrea m of the normal open reading frame (ORF), The resulting amino-terminal exte nsion alters the properties of these proteins and their intracellular distr ibution, In cells overexpressing eIF4E, c-myc is overexpressed and particul arly the larger, CUG-initiated form (Myc1), Recent reports suggest that syn thesis of Myc2, the normally expressed AUG-initiated form, is mediated by a n IRES. To determine,what role eIF4E might play in c-myc expression, the c- myc 5' untranslated region (UTR) was fused in-frame to CAT reporters, and s everal more derivative constructs were made. In vitro translation experimen ts (with and without eIF4E/F); expression in CHO cells transformed with eIF 4E; and deletion/mutation analysis demonstrated that Myc1 is translated by a scanning mechanism, while Myc2 is translated by Internal Ribosome Reposit ioning. Moreover, the existence of a true IRES in the 5'UTR was contradicte d by its failure to direct translation of a circular transcript, in contras t to hsp70, The c-myc 5'UTR also failed to engage in translation in the abs ence of functional eIF4F, after cleavage of the eIF4G component with CVB4 p rotease-2A, The Internal Repositioning Element (IRPE) in c-myc 5'UTR was de limited to nucleotides (nt) 394-440 from the P1 transcription start site.