Detection and degradation of nonsense-mediated mRNA decay substrates involve two distinct Upf1-bound complexes
Marine Dehecq1,2, Laurence Decourty1, Abdelkader Namane1, Caroline Proux3, Joanne Kanaan4, Hervé Le Hir4, Alain Jacquier1, Cosmin Saveanu1*
1 – Génétique des Interactions Macromoléculaires, Genomes and Genetic Department, Institut Pasteur, 25-28 rue du docteur Roux 75015 Paris, France
2 – Université Pierre et Marie Curie, 4 place Jussieu, 75005 Paris
3 – Transcriptome and Epigenome, CITECH, Institut Pasteur, 25-28 rue du docteur Roux 75015 Paris, France
4 – Expression des ARN messagers eucaryotes, CNRS UMR8197, Inserm U1024, Biology department, Institut de Biologie de l'Ecole Normale Supérieure, 46 rue d'Ulm 75005 Paris, France
Nonsense-mediated mRNA decay (NMD) is a major translation-dependent RNA degradation pathway required for embryo development and telomere maintenance. Core NMD factors Upf1, Upf2 and Upf3 are conserved from yeast to mammals but current NMD molecular models are radically different in these species. We performed the first large-scale quantitative characterization of yeast NMD complexes through affinity purification and mass-spectrometry with 7 different NMD-related factors, with or without RNase, in strains deleted or not for NMD genes. This extensive characterization of NMD complexes identified two distinct complexes associated with Upf1: Detector (Upf1-2-3) and Effector. Effector contained, in addition to Upf1, the mRNA decapping enzyme and two potential equivalents of mammalian Smg6-5-7: Nmd4 and Ebs1. Like the Smg proteins, Nmd4 and Ebs1 were required for efficient NMD. Our results suggest that the core eukaryotic NMD machinery is conserved across species and operates through successive Upf1-bound Detector and Effector complexes.
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Two graphs are shown, one with the enrichment factors in the corresponding purifications, and another one with intensity scores for the proteins. Click -and-drag on the first graph allows the selection of a subset of proteins, visible in the data table at the right. The enrichment graph can be saved as a pdf file. The threshold to label proteins, log2 enrichment values, can be adjusted using the cursor under the graph.
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