Nonsense-mediated mRNA decay (NMD) blockage promotes nonsense mRNA stabilization in protein 4.1R deficient cells carrying the 4.1R Coimbra variant of hereditary elliptocytosis.
We describe a new approach to stabilize nonsense mRNA, based on the inhibition of the NMD mechanism, by combining cycloheximide-mediated inhibition of translation, and caffeine-mediated inhibition of UPF1 phosphorylation. This approach aimed to identify the impact of a 4.1R splicing mutation. This mutation is involved in a partial deficiency of 4.1R in the homozygous state in a patient with hereditary elliptocytosis and a moderated hemolytic anemia. We show that, in addition to two known minor shortened and stable spliceoforms, the mutation activates an intronic cryptic splice site, which results in a nonsense mRNA major isoform, targeted to degradation in intact cells by NMD. This accounts for the main cause of 4.1R partial deficiency. In a general perspective, blocking the NMD mechanism would help to identify a missing isoform, and pave the path for a molecular targeting strategy to circumvent a deleterious splicing pathway in favor of a therapeutic splicing pathway.
Morinière
Madeleine
M
mRNA Metabolism in Normal and Pathological Cells, CGMC, CNRS, Université Lyon 1, Villeurbanne, France.
Delhommeau
François
F
Calender
Alain
A
Ribeiro
Leticia
L
Delaunay
Jean
J
Baklouti
Faouzi
F
D017398:Alternative Splicing / Q000235:genetics; D018389:Codon, Nonsense; D003598:Cytoskeletal Proteins / Q000172:deficiency / Q000235:genetics*; D004612:Elliptocytosis, Hereditary / Q000235:genetics*; D006801:Humans; D007438:Introns; D008565:Membrane Proteins / Q000172:deficiency / Q000235:genetics*; D020033:Protein Isoforms; D020871:RNA Stability; D012333:RNA, Messenger / Q000235:genetics
D018389:Codon, Nonsense; D003598:Cytoskeletal Proteins; D008565:Membrane Proteins; D020033:Protein Isoforms; D012333:RNA, Messenger; C028841:erythrocyte membrane band 4.1 protein