Identification of RNA-binding proteins involved in splicing regulation of non-canonical splice sites of Fanconi Anemia pathway genes

RNA binding proteins are key regulators of RNA life which assemble with RNA to form ribonucleoprotein complexes. This class of proteins is involved in various processes in cells such as alternative splicing and their impairment can lead to various human diseases. This study inspects introns flanked by GC-AG splice junctions and located in genes of the Fanconi Anemia pathway to find putative RBPs that may regulate the splicing of these non-canonical splice sites. Three sources of RNA binding protein data were used in this study: RBPmap which uses a sequence homology approach to evaluate the presence of the RBPs binding motifs in a given sequence of RNA, RBPsuite, a machine learning-based method for the prediction of the RNA binding sites, and eCLIP data from the ENCODE project containing the experimental data of various RBPs. Moreover, the shRNA RNA-seq data was acquired from the ENCODE database and exploited to confirm the result obtained. All sources were checked for the prediction and presence of RBPs commonly acting on the introns of the FA pathway genes which encompass GC-AG dinucleotides at their splicing junctions. Five RBPs, TRA2A, SRSF7, U2AF2, SUPV3L1, and ZNF800 showed the potential to preferably govern the alternative splicing of the Fanconi Anemia pathway genes that contain non-canonical GC splicing sites in their 5´end. These results indicate the probable presence of trans-acting elements that can commonly support and alter the splicing of the transcripts that contain GC-AG noncanonical splicing site junctions. These results revalidate the importance of these unique cis-elements and their regulatory function which can be tuned by the act of RNA binding proteins.