O-GlcNAc regulates gene expression by controlling detained intron splicing

Abstract

Intron detention in precursor RNAs serves to regulate expression of a substantial fraction of genes in eukaryotic genomes. How detained intron (DI) splicing is controlled is poorly understood. Here, we show that a ubiquitous post-translational modification called O-GlcNAc, which is thought to integrate signaling pathways as nutrient conditions fluctuate, controls detained intron splicing. Using specific inhibitors of the enzyme that installs O-GlcNAc (O-GlcNAc transferase, or OGT) and the enzyme that removes O-GlcNAc (O-GlcNAcase, or OGA), we first show that O-GlcNAc regulates splicing of the highly conserved detained introns in OGT and OGA to control mRNA abundance in order to buffer O-GlcNAc changes. We show that OGT and OGA represent two distinct paradigms for how DI splicing can control gene expression. We also show that when DI splicing of the O-GlcNAc-cycling genes fails to restore O-GlcNAc homeostasis, there is a global change in detained intron levels. Strikingly, almost all detained introns are spliced more efficiently when O-GlcNAc levels are low, yet other alternative splicing pathways change minimally. Our results demonstrate that O-GlcNAc controls detained intron splicing to tune system-wide gene expression, providing a means to couple nutrient conditions to the cell's transcriptional regime.

Mammalian Cell Proliferation Requires Noncatalytic Functions of O-GlcNAc Transferase

“…By quantifying how much each OGT activity impacts the abundance of each protein in the cell, we have identified the biological pathways most affected by each OGT activity. Consistent with known roles for O-GlcNAc, we found that Ser/Thr glycosylation broadly regulates metabolism and gene expression, with enriched pathways including splicing (44), translation (69)(70)(71), and autophagy (72)(73)(74). Among the pathways linked to OGT's noncatalytic activities are oxidative phosphorylation, the electron transport chain, and components of the actin cytoskeleton, consistent with prior data suggesting OGT binds to the actin-regulating Ecadherin/beta-catenin complex (39).…”

“…Expected outliers in the data further validated the regression analysis ( Figure 4E). For example, OGA abundance is known to decrease with low protein O-GlcNAc levels (40,44), and we found that OGA had the most negative β GlcNAc value after two days of dTAG-13 treatment.…”

“…uncleaved HCF-1. OGA synthesis, which is downregulated by low protein O-GlcNAc levels (40,44),greatly decreased without OGT ( Figure S1F). By day 3, we observed a 42 kDa PARP-1 fragment associated with necrosis ( Figure S1F) (45,46).…”

“…Inhibition altered protein levels in the opposite direction from removing OGT's noncatalytic functions, a phenomenon observed at both the protein and biological pathway levels ( Figures 5C, S5B-D). It is well-known that OGT levels rapidly increase upon loss of protein O-GlcNAcylation (44,48,67,68), with the rapid increases driven by splicing and transcriptional controls that we removed in the MEF system. These mechanisms were thought to serve as a means to restore normal O-GlcNAc levels.…”

Arabidopsis ACINUS is O-glycosylated and regulates transcription and alternative splicing of regulators of reproductive transitions

“…While we were getting our revised manuscript ready for submission, evidence was reported for similar function of O-GlcNAc in intron splicing in metazoan and for broad presence of stress-dependent intron retention in plants. Interestingly, inhibition of OGT was found to increase splicing of detained introns in human cells 61 . Detained introns are a novel class of post-transcriptionally spliced (pts) introns, which are one or few introns retained in transcripts where other introns are fully spliced 62 .…”