A Systematic Identification of RNA-Binding Proteins (RBPs) Driving Aberrant Splicing in Cancer

Alternative Splicing (AS) is a post-transcriptional process that allows a single RNA to produce different mRNA variants and, in some cases, multiple proteins. Various processes, many yet to be discovered, regulate AS. This study focuses on regulation by RNA-binding proteins (RBPs), which are not only crucial for splicing regulation but also linked to cancer prognosis and are emerging as therapeutic targets for cancer treatment. CLIP-seq experiments help identify where RBPs bind on nascent transcripts, potentially revealing changes in splicing status that suggest causal relationships. Selecting specific RBPs for CLIP-seq experiments is often driven by a priori hypotheses.

We developed an algorithm to detect RBPs likely related to splicing changes between conditions by integrating several CLIP-seq databases and a differential splicing detection algorithm. This work refines a previous study by improving splicing event prediction, testing different enrichment statistics, and performing additional validation experiments. The new method provides more accurate predictions and is included in the Bioconductor package EventPointer 3.14. We tested the algorithm in four experiments involving knockdowns of seven different RBPs. The algorithm accurately assessed the statistical significance of these RBPs using only splicing alterations. Additionally, we applied the algorithm to study sixteen cancer types from The Cancer Genome Atlas (TCGA) and three from TARGET. We identified relationships between RBPs and various cancer types, including alterations in CREBBP and MBNL2 in adenocarcinomas of the lung, liver, prostate, rectum, stomach, and colon. Some of these findings are validated in the literature, while others are novel.

The developed algorithm enhances the ability to predict and understand RBP-related splicing changes, offering more accurate predictions and novel insights into cancer-related splicing alterations. This work highlights the potential of RBPs as therapeutic targets and contributes to the broader understanding of their roles in cancer biology.