Investigation into human Tra2 protein-dependent splicing in MDA-MB-231 cells using iCLIP and RNA-seq. Homo sapiens

Alternative splicing—the production of multiple mRNA isoforms from a single gene—is regulated in part by RNA-binding proteins (RBPs). While the RBPs Tra2α and Tra2β have both been implicated in the regulation of alternative splicing, their relative contribution to this process are not well understood. Here we use iCLIP to identify Tra2β target exons in MDA-MB-231 cells. We find that simultaneous—but not individual—depletion of Tra2α and Tra2β induces substantial shifts in the splicing pattern of endogenous Tra2β target exons identified by iCLIP. We next use RNA-seq following joint Tra2 protein depletion to comprehensively identify Tra2 protein-dependent exons in MDA-MB-231 cells. Overall design: Endogenous Tra2β binding sites were mapped across the MDA-MB-231 cell transcriptome in biological triplicate iCLIP experiments. RNA-seq was performed using three biological replicates of negative control siRNA treated MDA-MB-231 cells and three biological replicates of TRA2A and TRA2B siRNA treated MDA-MB-231 cells.