Nonsense mutations in close proximity to the initiation codon fail to trigger full nonsense-mediated mRNA decay.
Nonsense-mediated mRNA decay (NMD) is a surveillance mechanism that degrades mRNAs containing premature translation termination codons. In mammalian cells, a termination codon is ordinarily recognized as "premature" if it is located greater than 50-54 nucleotides 5' to the final exon-exon junction. We have described a set of naturally occurring human beta-globin gene mutations that apparently contradict this rule. The corresponding beta-thalassemia genes contain nonsense mutations within exon 1, and yet their encoded mRNAs accumulate to levels approaching wild-type beta-globin (beta(WT)) mRNA. In the present report we demonstrate that the stabilities of these mRNAs with nonsense mutations in exon 1 are intermediate between beta(WT) mRNA and beta-globin mRNA carrying a prototype NMD-sensitive mutation in exon 2 (codon 39 nonsense; beta 39). Functional analyses of these mRNAs with 5'-proximal nonsense mutations demonstrate that their relative resistance to NMD does not reflect abnormal RNA splicing or translation re-initiation and is independent of promoter identity and erythroid specificity. Instead, the proximity of the nonsense codon to the translation initiation AUG constitutes a major determinant of NMD. Positioning a termination mutation at the 5' terminus of the coding region blunts mRNA destabilization, and this effect is dominant to the "50-54 nt boundary rule." These observations impact on current models of NMD.
Inácio
Angela
A
Centro de Genética Humana, Instituto Nacional de Saúde Dr. Ricardo Jorge, Av. Padre Cruz, 1649-016 Lisbon, Portugal.
Silva
Ana Luísa
AL
Pinto
Joana
J
Ji
Xinjun
X
Morgado
Ana
A
Almeida
Fátima
F
Faustino
Paula
P
Lavinha
João
J
Liebhaber
Stephen A
SA
Romão
Luísa
L
D000818:Animals; D002460:Cell Line; D003062:Codon; D018387:Codon, Initiator; D018389:Codon, Nonsense; D018388:Codon, Terminator; D004247:DNA / Q000737:chemistry; D005091:Exons; D005799:Genes, Dominant; D005914:Globins / Q000737:chemistry / Q000235:genetics; D006367:HeLa Cells; D006801:Humans; D007438:Introns; D051379:Mice; D009154:Mutation; D009841:Oligonucleotides / Q000235:genetics; D016366:Open Reading Frames; D010957:Plasmids / Q000378:metabolism; D011132:Polyribosomes / Q000737:chemistry / Q000378:metabolism; D011401:Promoter Regions, Genetic; D014176:Protein Biosynthesis; D017434:Protein Structure, Tertiary; D012326:RNA Splicing; D012333:RNA, Messenger / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012260:Ribonucleases / Q000737:chemistry; D013395:Sucrose / Q000494:pharmacology; D013997:Time Factors; D014162:Transfection
D003062:Codon; D018387:Codon, Initiator; D018389:Codon, Nonsense; D018388:Codon, Terminator; D009841:Oligonucleotides; D012333:RNA, Messenger; D013395:Sucrose; D005914:Globins; D004247:DNA; D012260:Ribonucleases