Abstract
Non-sense-mediated mRNA decay (NMD) is a mechanism of translation-dependent mRNA surveillance in eukaryotes: it degrades mRNAs with premature termination codons (PTCs) and contributes to cellular homeostasis by downregulating a number of physiologically important mRNAs. In the NMD pathway, Upf proteins, a set of conserved factors of which Upf1 is the central regulator, recruit decay enzymes to promote RNA cleavage. In mammals, the degradation of PTC-containing mRNAs is triggered by the exon–junction complex (EJC) through binding of its constituents Upf2 and Upf3 to Upf1. The complex formed eventually induces translational repression and recruitment of decay enzymes. Mechanisms by which physiological mRNAs are targeted by the NMD machinery in the absence of an EJC have been described but still are discussed controversially. Here, we report that the DEAD box proteins Ddx5/p68 and its paralog Ddx17/p72 also bind the Upf complex by physical interaction with Upf3, thereby interfering with the binding of EJC. By activating the NMD machinery, Ddx5 is shown to regulate the expression of its own, Ddx17 and Smg5 mRNAs. For NMD triggering, the adenosine triphosphate-binding activity of Ddx5 and the 3′-untranslated region of substrate mRNAs are essential.
Highlights
Non-sense-mediated mRNA decay (NMD) is an mRNA quality control mechanism that protects eukaryotic cells from incomplete and potentially toxic proteins [1,2,3,4] and regulates protein expression from a number of physiologically important mRNAs (5–10%) [5,6,7,8,9,10,11].Aberrant mRNAs with a premature translation termination codon (PTC) result from mutation or rearrangement of genomic DNA or defects in mRNA biogenesis
Functional Upf1-Upf2-Ufp3 complexes are integral components of messenger ribonucleoproteins, whereby the overall protein composition of an mRNP depends on its function, apparent, e.g. through the replacement of nuclear cap-binding protein CBP80 by the cytoplasmic cap-binding protein eIF4E after the first round of translation
Detailed protein analysis of the Ddx5-specific immunoprecipitates revealed some of these typical mRNP components, like CBP80, eIF4E, eukaryotic initiation factor 4G (eIF4G), MAGOH, Poly(A)-binding protein C1 (PABPC1) and PABPN1, bound to Ddx5 in an RNase-sensitive manner, suggesting an indirect, RNA-mediated interaction with Ddx5
Summary
Non-sense-mediated mRNA decay (NMD) is an mRNA quality control mechanism that protects eukaryotic cells from incomplete and potentially toxic proteins [1,2,3,4] and regulates protein expression from a number of physiologically important mRNAs (5–10%) [5,6,7,8,9,10,11].Aberrant mRNAs with a premature translation termination codon (PTC) result from mutation or rearrangement of genomic DNA or defects in mRNA biogenesis. The signal for their degradation is a translation-termination codon located at least 50–55 nt upstream of an exon–exon junction [1]. According to the exon junction complex (EJC) model, EJC proteins Upf (upstream frame shifting) 2 and Upf (bound by MAGOH, Y14, and eIF4AIII) signal degradation of these mRNAs by binding to the SURF complex (consisting of Smg, Smg, Smg, Upf, eRF3 and eRF1) formed at the stalling ribosome [12,13,14,15,16]. As with aberrant mRNAs, direct or indirect binding of Upf to the 30-UTR might be envisaged as to result in a competition between Upf and cytoplasmic poly(A)-binding protein (PABP) for binding to the translation release factors eRF1 and eRF3 [19,22]. Binding of PABP to release factors is thought to preserve translational competence and transcript stability
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