RBDmap - Comprehensive Identification of RNA-Binding Domains in Human Cells

The RNA interactomes of HeLa and HEK293 cells jointly comprise 1106 RNA-binding proteins (RBPs), with almost half of these lacking well-defined RNA-binding domains (RBDs), suggesting the existence of numerous unknown RNA-binding architectures. Here, we report on “RBDmap”, a new method built on interactome capture, to comprehensively identify the RBDs of native RBPs in HeLa cells. Making use of in vivo UV-crosslinking of RBPs to polyadenylated RNAs, capture on oligo(dT) magnetic beads, sequential proteolytic digestion and LC-MS/MS combined with a sophisticated scoring algorithm, RBDmap “re-discoveres” many known RNA-binding sites (e.g. RRM, KH) of numerous well characterized RBPs and suggests novel RBDs.