Intron retention and its impact on gene expression and protein diversity: A review and a practical guide

Abstract

Intron retention (IR) occurs when a complete and unspliced intron remains in mature mRNA. An increasing body of literature has demonstrated a major role for IR in numerous biological functions, including several that impact human health and disease. Although experimental technologies used to study other forms of mRNA splicing can also be used to investigate IR, a specialized downstream computational analysis is optimal for IR discovery and analysis. Here we provide a review of IR and its biological implications, as well as a practical guide for how to detect and analyze it. Several methods, including long read third generation direct RNA sequencing, are described. We have developed an R package, FakIR, to facilitate the execution of the bioinformatic tasks recommended in this review and a tutorial on how to fit them to users aims. Additionally, we provide guidelines and experimental protocols to validate IR discovery and to evaluate the potential impact of IR on gene expression and protein output. This article is categorized under: RNA Evolution and Genomics > Computational Analyses of RNA RNA Processing > Splicing Regulation/Alternative Splicing RNA Methods > RNA Analyses in vitro and In Silico K E Y W O R D S intron retention, nonsense mediated decay, post-transcriptional gene regulation, RNA export, RNA processing 1 | INTRODUCTION Alternative splicing of messenger RNA (mRNA) is responsible for much of the proteome complexity in mammals (Berget, Moore, & Sharp, 2000; Nilsen & Graveley, 2010). We now know that at least 90% of all mammalian genes can undergo some form of alternative splicing, often generating multiple protein isoforms with sometimes disparate biological functions (Wang et al., 2008). Intron retention (IR) is a type of alternative splicing that is gaining increased interest in human health and disease research. Originally described in plants and viruses, IR has now been shown to be a common form of alternative David F. Grabski, Lucile Broseus, and Bandana Kumari contributed equally to this study.

To “Z” or not to “Z”: Z-RNA, self-recognition, and the MDA5 helicase

“…Intron retention also can occur in linear sequences. It acts to increase protein isoform diversity [ 64 ] and enables rapid responses by delaying splicing until the protein product is required [ 65 ]. As discussed below, introns also can be processed to produce cytoplasmic dsRNA [ 66 ].…”

“…Only recently have the technical challenge to fully documenting the extent and consequences of these noncanonical RNA processing pathways been solved. How they alter the dsRNAome and MDA5 activation is currently unresolved [ 64 , 67 ].…”

Specific transcriptomic signatures and dual regulation of steroidogenesis between fetal and adult mouse Leydig cells

“…Intron retention is characterized by the inclusion of one or more introns in mature mRNA transcripts and has been previously considered to be an artefact of a dysfunctional spliceosome. It is known that alternative splicing, including intron retention, is frequent during embryonic development and contribute not only to the plasticity of the transcriptome but also the regulation of gene expression and protein diversity (Revil et al, 2010;Kalsotra and Cooper, 2011;Grabski et al, 2021). Here, we showed that intron retention is a landmark of the FLC transcriptome.…”

Specific Transcriptomic Signatures and Dual Regulation of Steroidogenesis Between Fetal and Adult Mouse Leydig Cells

“…Overall, we showed that intron retention is a landmark of the FLC transcriptome. It is known that alternative splicing is frequent during embryonic development, usually cell/organ specific and plays a role in gene expression regulation and protein diversity ( Revil et al, 2010 ; Kalsotra and Cooper, 2011 ; Grabski et al, 2021 ).…”

“…Intron retention is characterized by the inclusion of one or more introns in mature mRNA transcripts and has been previously considered to be an artifact of a dysfunctional spliceosome. It is known that alternative splicing, including intron retention, is frequent during embryonic development and contribute not only to the plasticity of the transcriptome but also the regulation of gene expression and protein diversity ( Revil et al, 2010 ; Kalsotra and Cooper, 2011 ; Grabski et al, 2021 ). Here, we showed that intron retention is a landmark of the FLC transcriptome.…”