Effect of nonsense-mediated decay inhibition on expression of 3' UTR spliced transcripts in colorectal carcinoma cell line HCT116

The presence of an exon junction more than 55nt downstream of a stop codon is generally considered to be a trigger for nonsense-mediated decay (NMD). Thus, transcripts containing 3’UTR introns (3UIs) are often considered as transcriptional noise. Interestingly, upon knockdown of NMD factor UPF1 we see examples of both increased and decreased expression of the 3UI-spliced isoforms. In the oncogene HRAS, upon UPF1 knockdown, we see the 3UI-retaining isoform increases in expression where the spliced isoform decreases, suggesting that 3UI splicing protects the transcript against UPF1-mediated degradation. To investigate the role of nonsense-mediated decay on expression of 3'UTR spliced transcripts, HCT116 cells were treated with either siUPF1 or siDsRed (control) in the presence or absence of an NMD small molecule inhibitor. Following RNAseq, we conducted differential transcript usage and differential splicing analyses.