The presence of an exon junction complex (EJC) positioned downstream of a termination codon is a key indicator for triggering nonsense-mediated mRNA decay (NMD), provided that the distance between the termination codon and the downstream exon-exon junction exceeds 50-55 nucleotides. As a key factor in this pathway, UPF3B bridges CASC3-EJC core complex to UPF2 via its C-terminal and N-terminal regions, respectively. The middle region of UPF3B remains poorly characterized, despite being a hotspot for mutations associated with neurodevelopmental disorders. Here, an EJC complex containing UPF2, UPF3B, CASC3, the EJC core, RNA (U55), and the ATP analogue AMPPNP was reconstituted in vitro. Crosslinking mass spectrometry (CLMS) analysis revealed a previously unrecognized interaction between the middle region of UPF3B and UPF2. Cryo-EM analysis and binding affinity assays using UPF3B mutants are currently underway. Collectively, this work advances our understanding of EJC-mediated NMD and will provide deeper insights into the molecular basis of neurodevelopmental diseases associated with UPF3B mutations.
Reconstitution and Biochemical Characterization of Exon Junction Complexes Involved in Nonsense-mediated mRNA Decay
Sun, L. (Author). 20 Jan 2026
Student thesis: Doctoral Thesis › Doctor of Philosophy (PhD)