Genome Scale Analysis of Alternative Splicing Events in lncRNA Knockdowns

Abstract

Recent developments in our understanding of the interactions between lncRNA and cellular components have improved treatment approaches for various human diseases including cancer, vascular diseases, and brain diseases (1, 2, 3). Although investigation of specific lncRNAs revealed their role in the metabolism of cellular RNA, our understanding of their contribution to post-transcriptional regulation is relatively limited. In this study, we explore the role of lncRNAs in modulating alternative splicing and their impact on downstream protein-RNA interaction networks. Analysis of alternative splicing events across 39 lncRNA wildtype and knockout RNA-sequencing datasets from three human cell lines: Hela (Cervical Cancer), K562 (Myeloid Leukemia), and U87 (Glioblastoma), resulted in high confidence (fdr < 0.01) identification of 4432 skipped exon events and 2474 retained intron events, implicating 759 genes to be impacted at post-transcriptional level due to the loss of lncRNAs. We observed that a majority of the alternatively spliced genes in a lncRNA knockout were specific to the cell type, in agreement with the finding that genes affected by alternative splicing also displayed enriched functions in a cell type specific manner (4, 5). To understand the mechanism behind this cell-type specific alternative splicing patterns, we analyzed RNA binding protein (RBP)-RNA interaction profiles across the spliced regions. Despite limited REP binding data across cell lines, alternatively spliced events detected in lncRNA perturbation experiments were associated with RBPs binding in proximal intron-exon junctions, in a cell type specific manner. Based on the RBP binding profiles in Hela and K562 cells, we hypothesize that several lncRNAs are likely to exhibit a sponge effect in disease contexts, resulting in the functional disruption of RBPs, and their downstream functions. We propose that such lncRNA sponges can extensively rewire the post-transcriptional gene regulatory networks by altering the protein-RNA interaction landscape in a cell-type specific manner.