Comprehensive Identification of RNA-Binding Domains in Human Cells

Summary

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.

Graphical Abstract

Highlights

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  Experimental generation of an atlas of RNA-binding sites (RBS) in human cells

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  RBS overlap with enzymatic cores and protein-protein interaction sites

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  About half of the total RBS map to disordered protein regions

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  RBS are enriched for phosphorylation, acetylation, and methylation sites

Abstract

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.