The canonical UPF1-dependent nonsense-mediated mRNA decay is inhibited in transcripts carrying a short open reading frame independent of sequence context

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Abstract
Nonsense-mediated mRNA decay (NMD) is a surveillance mechanism that degrades mRNAs carrying premature translation termination codons. Generally, NMD is elicited if translation terminates >50–54 nucleotides (nt) upstream of an exon–exon junction. We have previously reported that human β-globin mRNAs carrying 5′-proximal nonsense mutations (e.g., β15) accumulate to normal levels, suggesting an exception to the “50–54-nt boundary rule.” In the present report, we demonstrate that the strength of the UPF1-dependent NMD of mutant β-globin mRNAs is specifically determined by the proximity of the nonsense codon to the initiation AUG. This conclusion is supported by a parallel effect of the short ORF size on NMD of nonsense-containing α-globin mRNAs. To determine whether the short-ORF effect on NMD response is conserved in heterologous transcripts, we assessed its effects on a set of β-globin/triosephosphate isomerase (TPI) hybrid mRNAs and on the TPI mRNA. Our data support the conclusion that nonsense mutations resulting in a short ORF are able to circumvent the full activity of the canonical UPF1-dependent NMD pathway.