5′ UTR length regulates the production of alternative N-terminal protein isoforms in health and disease

The 5′ untranslated region (5′ UTR) of an mRNA is classically viewed as a regulatory region that controls the amount of protein production, but not the resulting protein sequence. Here, we demonstrate that 5′ UTR length also plays a direct role in alternative N-terminal protein isoform production by controlling start codon selection. We find that very short 5′ UTRs enhance leaky scanning, thereby promoting the production of truncated alternative N-terminal protein isoforms. Changes in 5′ UTR length due to alternative transcription initiation can tune the relative abundance of alternative N-terminal isoforms from the same gene. In addition, we identify mutations in rare genetic diseases that alter 5′ UTR length, including a deletion in the VHL 5′ UTR in von Hippel–Lindau disease that shifts translation toward the shorter VHLp19 isoform. Together, our results implicate 5′ UTR length as a determinant of alternative N-terminal isoform production and reveal an underappreciated mechanism by which noncoding mutations can reshape the proteome.