Structure and Mechanism of Key Nonsense-Mediated mRNA Decay Factor Complexes

Nonsense-mediated mRNA decay (NMD) is an essential eukaryotic surveillance mechanism to eliminate aberrant and potentially harmful mRNAs that contain a premature termination codon (PTC). The discrimination of a PTC from a correct stop codon during active translation is key, but the underlying molecular mechanisms remain elusive. We recently discovered a role for the NMD factor UPF3B in delay of translation termination in vitro, mediated by new interactions of UPF3B with ribosome, release factors and UPF1. This puts UPF3B at centre stage of the events at the terminating ribosome. To validate our discovery and to discover novel interactions of NMD factors, I will establish in vivo cross-linking/mass spectrometry. I will explore the impact of disease-conferring mutations in NMD factors UPF3B and SMG1 kinase on functionally important protein-protein interactions, and characterise novel functions of UPF1 and UPF3B at near-atomic resolution in an integrative approach, combining biochemistry, biophysics, electron cryo-microscopy and crystallography or NMR. This highly interdisciplinary research aims at a step-change in our molecular-level understanding of the role of the individual NMD factors in a paramount step of human translational control; a vital prerequisite for the development of new intervention strategies to treat NMD-related disease.