Translational control by an upstream open reading frame in the HER-2/neu transcript

Overexpression of the HER-2 (neu, erbB-2) receptor results in cellular tran sformation and is associated with a variety of human cancers. Multiple mech anisms, including gene amplification and transcriptional, posttranscription al, and translational controls contribute to the regulation of HER-S expres sion, One of the components of these regulatory mechanisms is a short upstr eam open reading frame (uORF) in the HER-2 mRNA that represses downstream t ranslation in a variety of cell types. Here we explore the mechanism by whi ch this uORF exerts its inhibitory effect. As judged by comparisons of protein and mRNA abundance and by polysomal dis tribution analyses, the uORF represses translation of the HER-2 cistron or of a heterologous reporter gene. Despite its conservation among mammalian s pecies, the peptide sequence of the uORF is not required for this inhibitor y effect. Rather, the majority of ribosomes that load on the HER-S mRNA mos t likely translate the uORF and are then unable to reinitiate at the downst ream AUG codon, in part due to the short intercistronic spacing. A minority of ribosomes gain access to the HER-P initiation codon either by leaky sca nning past the upstream AUG codon or by reinitiating after having translate d the uORF despite the short intercistronic region. These results suggest t hat the HER-2 uORF controls synthesis of this oncoprotein by limiting ribos omal access to downstream initiation sites.