The RNA-binding protein SFPQ preserves long-intron splicing and regulates circRNA biogenesis in mammals

Abstract

Circular RNAs (circRNAs) represent an abundant and conserved entity of non-coding RNAs; however, the principles of biogenesis are currently not fully understood. Here, we identify two factors, splicing factor proline/glutamine rich (SFPQ) and non-POU domain-containing octamer-binding protein (NONO), to be enriched around circRNA loci. We observe a subclass of circRNAs, coined DALI circRNAs, with distal inverted Alu elements and long flanking introns to be highly deregulated upon SFPQ knockdown. Moreover, SFPQ depletion leads to increased intron retention with concomitant induction of cryptic splicing, premature transcription termination, and polyadenylation, particularly prevalent for long introns. Aberrant splicing in the upstream and downstream regions of circRNA producing exons are critical for shaping the circRNAome, and specifically, we identify missplicing in the immediate upstream region to be a conserved driver of circRNA biogenesis. Collectively, our data show that SFPQ plays an important role in maintaining intron integrity by ensuring accurate splicing of long introns, and disclose novel features governing Alu-independent circRNA production.

Identification of Novel RNA Binding Proteins Influencing Circular RNA Expression in Hepatocellular Carcinoma

“…Another discordance was in larger and more diverse enrichment in the upstream regions compared to the downstream regions of splice sites of upregulated circRNAs. This finding was supported by a recent study that identified a class of circRNAs with aberrant splicing that differs between upstream and downstream regions [ 6 ]. Another example of this phenomenon is ESRP2, for which it has been shown that the position of the binding site defines the fate of the exon [ 52 , 53 ].…”

“…Furthermore, the epitranscriptomic nucleotide modification m6A is important for YTHDC1-mediated circRNA biogenesis [ 34 ]. Finally, a study using ENCODE datasets identified NONO as one of the most enriched RBPs in the flanking intron regions for a group of most highly expressed circRNAs in the K562 cell line [ 6 ].…”

“…The accumulation of short interspersed nuclear repetitive DNA elements (SINEs) also positively correlates with increased circRNA variety and number in more complex organisms [ 5 ]. Other mechanisms, such as the presence of long flanking introns, have been shown to drive the biogenesis of a class of circRNAs [ 6 ]. Different studies have shown that circRNAs are not just passive by-products of canonical splicing and that their expression is actively regulated [ 7 , 8 , 9 , 10 ].…”

“…The circRNA/mRNA expression ratio from the same gene locus can shift in different biological conditions since linear and circular transcripts can change their expression in different directions [ 8 , 9 , 10 ]. Core spliceosomal components have been shown to regulate circRNA expression [ 11 ], as well as different RBPs and splicing factors [ 6 , 12 , 13 , 14 , 15 , 16 , 17 , 18 , 19 , 20 ]. Quaking (QKI) is an example of a splicing factor that actively regulates circRNA biogenesis during the epithelial–mesenchymal transition [ 13 ].…”

The Regulation Network and Clinical Significance of Circular RNAs in Breast Cancer

“…Similarly, Pagliarini et al found that SAM68 can bind to the distal intron inverted repeat Alu sequence of the spinal muscular atrophy gene (SMN), thereby supporting circRNA biosynthesis in vivo and in vitro (21). Splicing factor proline/glutamine-rich (SFPQ) is specifically enriched in introns flanking a type of Distal-Alu-Long-Intron (DALI) circRNA that is characterized by distal inverted Alu elements and long flanking introns, and depletion of SFPQ has been shown to significantly repress DALI circRNA production (22).…”