Direct and indirect effects of spliceosome disruption compromise gene regulation by nonsense-mediated mRNA decay

Pre-mRNA splicing, carried out in the nucleus by a large ribonucleoprotein machine known as the spliceosome, is functionally and physically coupled to the mRNA surveillance pathway in the cytoplasm called nonsense-mediated mRNA decay (NMD). The NMD pathway monitors for premature translation termination, which can result from alternative splicing, by relying on the exon junction complex (EJC) deposited on exon-exon junctions by the spliceosome. Recently, multiple genetic screens in human cell lines have identified numerous spliceosome components as putative NMD factors. Using publicly available RNA-seq datasets from K562 and HepG2 cells depleted of 18 different spliceosome components, we found that natural NMD-targeted mRNA isoforms were upregulated when catalytic spliceosome members were reduced. While some of this increase could be due to widespread pleiotropic effects of spliceosome dysfunction (e.g. reduced expression of NMD factors due to missplicing of their mRNAs), we identified that AQR, SF3B1, SF3B4, and CDC40 may have a more direct role in NMD. We also tested the hypothesis that increased production of novel NMD substrates may overwhelm the pathway to find a direct correlation between the amount of novel NMD substrates detected and the degree of NMD inhibition observed. Finally, similar transcriptome alterations and NMD substrate upregulation were observed in cells treated with spliceosome inhibitors and in cells derived from retinitis pigmentosa patients with mutations in <i>PRPF8</i> and <i>PRPF31</i>. Overall, our results show that regardless of the cause, spliceosome disruption upregulates a broad set of NMD targets, which could contribute to cellular dysfunction in spliceosomopathies.