A dynamic intron retention program enriched in RNA processing genes regulates gene expression during terminal erythropoiesis

Abstract

Differentiating erythroblasts execute a dynamic alternative splicing program shown here to include extensive and diverse intron retention (IR) events. Cluster analysis revealed hundreds of developmentally-dynamic introns that exhibit increased IR in mature erythroblasts, and are enriched in functions related to RNA processing such as SF3B1 spliceosomal factor. Distinct, developmentally-stable IR clusters are enriched in metal-ion binding functions and include mitoferrin genes SLC25A37 and SLC25A28 that are critical for iron homeostasis. Some IR transcripts are abundant, e.g. comprising ∼50% of highly-expressed SLC25A37 and SF3B1 transcripts in late erythroblasts, and thereby limiting functional mRNA levels. IR transcripts tested were predominantly nuclear-localized. Splice site strength correlated with IR among stable but not dynamic intron clusters, indicating distinct regulation of dynamically-increased IR in late erythroblasts. Retained introns were preferentially associated with alternative exons with premature termination codons (PTCs). High IR was observed in disease-causing genes including SF3B1 and the RNA binding protein FUS. Comparative studies demonstrated that the intron retention program in erythroblasts shares features with other tissues but ultimately is unique to erythropoiesis. We conclude that IR is a multi-dimensional set of processes that post-transcriptionally regulate diverse gene groups during normal erythropoiesis, misregulation of which could be responsible for human disease.

The Y Chromosome Modulates Splicing and Sex-Biased Intron Retention Rates in Drosophila

“…In 9980 of these samples with PTC containing IRs, we observed a higher negative correlation coefficient between change in IR and change in expression than that observed for the in-frame IRs (permutation test P = 0.002; Figure 2F). Together, these results suggest a role for IR in sex-biased expression, and are consistent with previous reports indicating that IR can negatively tune gene expression (BRAUNSCHWEIG et al 2014;DVINGE AND BRADLEY 2015;PIMENTEL et al 2016). Finally, GO analysis showed that genes with testis-biased IR were enriched in oogenesis, neurogenesis, female gamete generation, and others (Table S3, hypergeometric test P < 0.001).…”

“…Intron retention (IR) is a type of AS event in which one or more introns are retained in the mRNA molecule. IR is the dominant AS type in Drosophila (GRAVELEY et al 2011) and affects about half of all introns in mammals (WONG et al 2013;BRAUNSCHWEIG et al 2014;EDWARDS et al 2016;PIMENTEL et al 2016). The "anomalous" mRNA generated by the retention of an intron may contain a premature termination codon (PTC) (JAILLON et al 2008), which can sometimes result in the degradation of the mRNA by the nonsense-mediated mRNA decay (NMD) pathway.…”

“…Because of this, mRNAs generated through IR events could be viewed as failures of the splicing process and void of biological function (JAILLON et al 2008;ROY AND IRIMIA 2008). However, recent studies suggested that IR events are involved in multiple cellular and physiological processes, including hematogenesis (WONG et al 2013;EDWARDS et al 2016;PIMENTEL et al 2016), T cell activation (NI et al 2016), neuronal differentiation (BRAUNSCHWEIG et al 2014), and carcinogenesis (DVINGE AND BRADLEY 2015;JUNG et al 2015). Despite these advances, questions remain regarding the extent to which introns are differentially retained between sexes and how sex-biased IR rates are regulated.…”

“…Introns fulfilling the following two criteria in any sample were regarded as IR events: first, the intron region was mapped with at least 10 reads (each read overlapped at least 6bps with the intron); second, the intron region had 100% coverage. To quantify IR rates, we used the metric Percent Intron Retention (PIR) as calculated in previous studies (BRAUNSCHWEIG et al 2014;PIMENTEL et al 2016) and briefly described here with minor modifications ( Figure 1A). For ease of description, suppose that the length of the target intron and the trimmed reads were L 1 and L 2 , respectively.…”

“…We mapped reads to the reference Drosophila genome ( Figure 1A) and identified 21,664 IR events ( Figure 1B) in 7,546 genes. We used the metric Percent Intron Retention (PIR, see Methods) as previously implemented (BRAUNSCHWEIG et al 2014;PIMENTEL et al 2016) to represent the percentage of transcripts with the focal intron retained in each sample. From these IR events, 1,304 had been annotated in Flybase (i.e., both IR and intron splicing isoforms being annotated), while the other 20,360 were newly identified.…”