Comprehensive Identification of RNA-Binding Domains in Human Cells

• Published in: Molecular cell 2016-08, Vol.63 (4), p.696-710
• Main Authors: Castello, Alfredo, Fischer, Bernd, Frese, Christian K., Horos, Rastislav, Alleaume, Anne-Marie, Foehr, Sophia, Curk, Tomaz, Krijgsveld, Jeroen, Hentze, Matthias W.
• Format: Article
• Language: English
• Subjects: Acetylation; binding sites; Computational Biology; Databases, Protein; Evolution, Molecular; HeLa Cells; Humans; lysine; Methylation; Models, Molecular; Mutation; Nucleic Acid Conformation; Phosphorylation; post-translational modification; Protein Binding; Protein Interaction Domains and Motifs; Protein Processing, Post-Translational; protein-protein interactions; Proteomics - methods; Resource; RNA; RNA - chemistry; RNA - genetics; RNA - metabolism; RNA-Binding Motifs; RNA-binding proteins; RNA-Binding Proteins - chemistry; RNA-Binding Proteins - genetics; RNA-Binding Proteins - metabolism; Structure-Activity Relationship; tyrosine
• ISSN: 1097-2765; 1097-4164
• DOI: 10.1016/j.molcel.2016.06.029

Mammalian cells harbor more than a thousand RNA-binding proteins (RBPs), with half of these employing unknown modes of RNA binding. We developed RBDmap to determine the RNA-binding sites of native RBPs on a proteome-wide scale. We identified 1,174 binding sites within 529 HeLa cell RBPs, discovering numerous RNA-binding domains (RBDs). Catalytic centers or protein-protein interaction domains are in close relationship with RNA-binding sites, invoking possible effector roles of RNA in the control of protein function. Nearly half of the RNA-binding sites map to intrinsically disordered regions, uncovering unstructured domains as prevalent partners in protein-RNA interactions. RNA-binding sites represent hot spots for defined posttranslational modifications such as lysine acetylation and tyrosine phosphorylation, suggesting metabolic and signal-dependent regulation of RBP function. RBDs display a high degree of evolutionary conservation and incidence of Mendelian mutations, suggestive of important functional roles. RBDmap thus yields profound insights into native protein-RNA interactions in living cells. [Display omitted] •Experimental generation of an atlas of RNA-binding sites (RBS) in human cells•RBS overlap with enzymatic cores and protein-protein interaction sites•About half of the total RBS map to disordered protein regions•RBS are enriched for phosphorylation, acetylation, and methylation sites Many recently discovered RNA-binding proteins (RBPs) do not show architectural similarities with classical RBPs, and their modes of interaction with RNA were unclear. We developed and employed RBDmap as a method for the comprehensive determination of the RNA-interacting sites of RBPs, identifying more than a thousand such sites. These data yield unprecedented insight into RNA-protein interactions in cells with implications for numerous biological contexts.