Poster abstracts
Poster number 159 submitted by Savanna Schutte
Dissecting UPF3-UPF2 regulation of UPF1 dynamics within the Nonsense-Mediated mRNA Decay pathway
Savanna P. Schutte (Department of Molecular Genetics, The Ohio State University), Rabab Abu Alhasan (Department of Molecular Genetics, The Ohio State University), Rene Arvola (Department of Molecular Genetics, The Ohio State University), Guramrit Singh (Department of Molecular Genetics, The Ohio State University)
Abstract:
Nonsense-Mediated mRNA Decay (NMD) is a post-transcriptional mechanism that targets premature termination codon-containing transcripts, preventing production of truncated proteins. Prematurely terminating ribosomes are distinguished by core NMD factors UPF1, UPF2, and UPF3. While UPF1 is the key NMD activator, the function of UPF3 in this process is poorly understood. Our recent work suggests that human UPF3 paralogs, UPF3A and UPF3B, are non-essential but important enhancers of NMD that can function independently of their association with the exon junction complex (EJC). This challenges the widely accepted model of UPF3 primarily acting to bridge the EJC to the premature termination complex. By investigating NMD complexes in wild-type (WT) and UPF3 double knockout (3DKO) human colorectal cancer cell line (HCT116), we find that loss of UPF3B causes accumulation of UPF1/2 in a large macromolecular complex that is normally short-lived in WT conditions. We also find that the loss of UPF3B’s interaction with UPF2, and not the EJC, is responsible for accumulation of the complex. UPF2’s interaction with UPF1 is known to relieve its autoinhibited state, promoting its ATPase activity and altering RNA binding. Therefore, we have tested UPF1 functional mutants to investigate the possible role of UPF3 in modulating these effects through its UPF2 interaction. Preliminary data show that UPF1 overexpression in 3DKO rescues the enriched UPF2-bound complex, unlike the UPF1 mutants with impaired UPF2 interaction. Ongoing work is focused on characterizing the makeup and function of the accumulating complex as well as further defining the role UPF3 plays in modulating the activity of UPF1. Overall, our work illuminates a novel role of UPF3 in NMD activation and thereby in post-transcriptional gene regulation.
Keywords: Nonsense-mediated mRNA Decay (NMD), UPF3, post-transcriptional regulation