Widespread intron retention in mammals functionally tunes transcriptomes

Most mammalian genes contain introns that have to be spliced out before mRNAs are translated into protein. In a surprisingly high fraction of genes, specific introns are retained predominantly when expression is low in a particular cell type, which potentially acts as a widespread mechanism to reduce inappropriate gene expression.

HFSP Long-Term Fellow Ulrich Braunschweig and colleagues
authored on Thu, 08 January 2015

Retention of unspliced introns in mRNA has been thought to be rare in mammals, in contrast to plants and many protists in which it is the main form of alternative splicing. Translation of transcripts containing unspliced introns would in most cases result in truncated or erroneous polypeptides, and cells have evolved two main quality-control mechanisms for preventing their production. First, transcripts with retained introns are subject to nuclear sequestration and degradation by the exosome, and second, those transcripts that are exported to the cytosol often contain premature termination codons and are subject to turnover by the nonsense-mediated decay machinery. Recently, intron retention has been observed in some mammalian cell types, where it is used to coordinately regulate the expression of functionally coherent sets of genes.

Thanks to an increasing amount of high-quality, deep coverage RNA sequencing data , it was possible to analyze intron retention on a large and systematic scale in more than 100 human and mouse tissues and cell types. Surprisingly, retained introns were found to occur in up to three quarters of all intron-containing human and mouse genes in at least one cell type at robustly detectable levels (Figure A). This is particularly striking given the aforementioned quality control mechanisms that serve to remove such unspliced transcripts. Retention does not randomly affect introns but is more frequent in short introns with elevated C/G content and those with weak splice sites, which have the disadvantage of being recognized by the splicing machinery. It also occurs more frequently in brain tissues than in other organs. Moreover, retention of introns in brain tissues is more often conserved in vertebrates compared to other tissues, suggesting a regulatory role.

Figure: A, Distributions of percentages of total human and mouse introns detected as retained (at a rate of ≥ 10%) in transcripts sorted into ten different expression level bins in each RNA-seq sample (deciles, with deciles 1-4 averaged). B, Average ChIP-seq signal of Ser2-phosphorylated RNA pol II over introns with different percent intron retention (PIR) thresholds in K562 cells. Gray bars, flanking exons; y-axes represent input-subtracted average ChIP fragment density per million reads at each aligned bp.

Strikingly, intron retention is more prevalent in genes with low steady state levels in a given cell type, which are by and large genes whose function is not important or even detrimental to that cell. This was particularly evident in data obtained during differentiation of mouse embryonic stem cells to neurons, when retention of hundreds of introns in genes that are not functionally relevant to neurons increases concomitantly with downregulation of the expression of those genes. Such a mechanism thus affords a way to reduce levels of inappropriately expressed transcripts.

How transcript and intron retention levels are linked is not yet clear, but the authors observed evidence implicating transcription-coupled splicing factor recruitment. Dependent on the degree of retention of introns there is progressive accumulation of RNA polymerase II, indicating slow or stalled transcription (Fig. B). High densities of RNA pol II are similarly detected at introns that are most sensitive to depletion of core spliceosomal components. Considering that the splicing and transcription machineries interact, this suggests that suboptimal splicing of introns that are prone to retention impacts pol II elongation and requires higher transcription rates to be overcome.

Reference

Widespread intron retention in mammals functionally tunes transcriptomes. Braunschweig U, Barbosa-Morais NL, Pan Q, Nachman EN, Alipanahi B, Gonatopoulos-Pournatzis T, Frey B, Irimia M, Blencowe BJ. Genome Res. 2014 Nov;24(11):1774-86. doi: 10.1101/gr.177790.114. Epub 2014 Sep 25.

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