Unusual bipartite mode of interaction between the nonsense-mediated decay factors, UPF1 and UPF2.
Nonsense-mediated decay (NMD) is a eukaryotic quality control mechanism that degrades mRNAs carrying premature stop codons. In mammalian cells, NMD is triggered when UPF2 bound to UPF3 on a downstream exon junction complex interacts with UPF1 bound to a stalled ribosome. We report structural studies on the interaction between the C-terminal region of UPF2 and intact UPF1. Crystal structures, confirmed by EM and SAXS, show that the UPF1 CH-domain is docked onto its helicase domain in a fixed configuration. The C-terminal region of UPF2 is natively unfolded but binds through separated alpha-helical and beta-hairpin elements to the UPF1 CH-domain. The alpha-helical region binds sixfold more weakly than the beta-hairpin, whereas the combined elements bind 80-fold more tightly. Cellular assays show that NMD is severely affected by mutations disrupting the beta-hairpin binding, but not by those only affecting alpha-helix binding. We propose that the bipartite mode of UPF2 binding to UPF1 brings the ribosome and the EJC in close proximity by forming a tight complex after an initial weak encounter with either element.
Clerici
Marcello
M
European Molecular Biology Laboratory, Grenoble Outstation, Grenoble Cedex 9, France.
Mourão
André
A
Gutsche
Irina
I
Gehring
Niels H
NH
Hentze
Matthias W
MW
Kulozik
Andreas
A
Kadlec
Jan
J
Sattler
Michael
M
Cusack
Stephen
S
D000595:Amino Acid Sequence; D018360:Crystallography, X-Ray; D004252:DNA Mutational Analysis; D008854:Microscopy, Electron; D008969:Molecular Sequence Data; D020125:Mutation, Missense; D019906:Nuclear Magnetic Resonance, Biomolecular; D011485:Protein Binding; D054730:Protein Interaction Domains and Motifs; D025941:Protein Interaction Mapping; D020836:Protein Structure, Quaternary; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D020365:RNA Helicases; D020871:RNA Stability; D012333:RNA, Messenger / Q000378:metabolism*; D016601:RNA-Binding Proteins; D053838:Scattering, Small Angle; D016415:Sequence Alignment; D015534:Trans-Activators / Q000737:chemistry / Q000378:metabolism*; D014157:Transcription Factors / Q000737:chemistry / Q000378:metabolism*
D012333:RNA, Messenger; D016601:RNA-Binding Proteins; D015534:Trans-Activators; D014157:Transcription Factors; C468286:UPF2 protein, human; D020365:RNA Helicases; C101828:UPF1 protein, human