Transcription-coupled RNA surveillance in human genetic diseases caused by splice site mutations

Abstract

Current estimates indicate that approximately one-third of all disease-causing mutations are expected to disrupt splicing. Abnormal splicing often leads to disruption of the reading frame with introduction of a premature termination codon (PTC) that targets the mRNA for degradation in the cytoplasm by nonsense mediated decay (NMD). In addition to NMD there are RNA surveillance mechanisms that act in the nucleus while transcripts are still associated with the chromatin template. However, the significance of nuclear RNA quality control in the context of human genetic diseases is unknown. Here we used patient-derived lymphoblastoid cell lines as disease models to address how biogenesis of mRNAs is affected by splice site mutations. We observed that most of the mutations analyzed introduce PTCs and trigger mRNA degradation in the cytoplasm. However, for some mutant transcripts, RNA levels associated with chromatin were found down-regulated. Quantification of nascent transcripts further revealed that a subset of genes containing splicing mutations (SM) have reduced transcriptional activity. Following treatment with the translation inhibitor cycloheximide the cytoplasmic levels of mutant RNAs increased, while the levels of chromatin-associated transcripts remained unaltered. These results suggest that transcription-coupled surveillance mechanisms operate independently from NMD to reduce cellular levels of abnormal RNAs caused by SM.

Lessons from non-canonical splicing

“…These exons often introduce premature termination codons (PTCs) into the resulting transcripts, which can target them for nonsense-mediated decay (NMD) in the cytoplasm33–35 (Figure 1A). In some cases, abnormal splicing also leads to transcription-coupled surveillance mechanisms that can decrease the expression of resulting transcripts36. These quality control pathways often decrease the expression of transcripts containing cryptic exons, which makes these exons more difficult to detect and annotate during transcriptome sequencing analysis6–10.…”

“…Therefore, genome-wide sequencing will be required to reveal the full range of intronic variation that can activate cryptic splicing, perturb distal branch points or disturb regulatory regions32. It is also important to bear in mind that abnormally processed pre-mRNAs can interfere with transcription or cause co-transcriptional decay36,180. Dedicated genomic and transcriptomic experiments and computational approaches will therefore be needed to detect the full range of mutations that cause disease via non-canonical splicing.…”

Defective control of pre–messenger RNA splicing in human disease

“…More than 200 human diseases, including progeria and some forms of breast cancer and cystic fibrosis, are caused by point mutations that affect pre-mRNA splicing by destroying or weakening splice sites, or activating cryptic ones ( Wang et al, 2012 ), thereby producing mRNAs that encode defective proteins or that are targets for nonsense-mediated mRNA decay (NMD). Splicing defects can also lead to the cotranscriptional degradation of nascent pre-mRNAs ( Davidson et al, 2012 ; Vaz-Drago et al, 2015 ). A splice site mutation in BRAF is associated with resistance to the anticancer agent vemurafenib, but inhibitors of the generic splicing factor SF3B1 decrease the production of the mutation-induced BRAF variant and inhibit drug-resistant cell proliferation ( Poulikakos et al, 2011 ; Salton et al, 2015 ).…”

The contribution of alternative splicing to genetic risk for psychiatric disorders

“…That study also found that sQTLs were enriched in regions of the histone modification H3K4me3, a marker of gene promoters. Gene transcription and post‐transcriptional processing are coupled and factors that influence transcription can influence exon usage . These factors include the rate of RNA polymerase II elongation and chromatin structure and histone modifications that alter transcription rates .…”