The RNA-binding protein repertoire of embryonic stem cells

Abstract

RNA-binding proteins (RBPs) have essential roles in RNA-mediated gene regulation, and yet annotation of RBPs is limited mainly to those with known RNA-binding domains. To systematically identify the RBPs of embryonic stem cells (ESCs), we here employ interactome capture, which combines UV cross-linking of RBP to RNA in living cells, oligo(dT) capture and MS. From mouse ESCs (mESCs), we have defined 555 proteins constituting the mESC mRNA interactome, including 283 proteins not previously annotated as RBPs. Of these, 68 new RBP candidates are highly expressed in ESCs compared to differentiated cells, implicating a role in stem-cell physiology. Two well-known E3 ubiquitin ligases, Trim25 (also called Efp) and Trim71 (also called Lin41), are validated as RBPs, revealing a potential link between RNA biology and protein-modification pathways. Our study confirms and expands the atlas of RBPs, providing a useful resource for the study of the RNA-RBP network in stem cells.

Specific RNP capture with antisense LNA/DNA mixmers

“…Applied to HeLa and HEK293 cells, RNA interactome capture identified more than a thousand RBPs, hundreds of which were previously unknown to bind RNA. This method was successfully used to determine the poly(A)-RNA-binding proteome of different human cell types, including human (Baltz et al 2012;Castello et al 2012;Beckmann et al 2015) and murine (Kwon et al 2013;Liao et al 2016;Liepelt et al 2016) cells, as well as from organisms such as S. cerevisiae (Mitchell et al 2013;Beckmann et al 2015;Matia-González et al 2015), C. elegans (Matia-González et al 2015), D. melanogaster (Sysoev et al 2016;Wessels et al 2016), A. thaliana (Marondedze et al 2016;Reichel et al 2016), and the parasites plasmodium and trypanosoma (Bunnik et al 2016;Lueong et al 2016). However, the identification of the RBPs bound to specific RNAs remains challenging.…”

The human RBPome: From genes and proteins to human disease

“…al [17], human orthologs of RBPs identified in mouse embryonic stem cells by Kwon et. al [18] and RBPs reported in RBPDB [19] (Supplementary Table 1) for analysis in this study. Proteins annotated in ENSEMBL's human genome build which were not identified as RBPs were considered as Non-RBPs.…”

“…In this study, we construct a catalogue of 1344 genes encoding for RBPs in the human genome (Supplementary Table 1): identified from recent high-throughput screens including the mRNA interactome of proliferating HeLa cells through interactome capture [1], mRNA -bound proteome in the human embryonic kidney cells identified using the photoreactive nucleotide-enhanced UV crosslinking and oligo(dT) purification approach , [16] , proteins with the ability to bind RNA from the RNA compete experiments [17], human orthologs of RBPs identified in the mouse embryonic stem cells through interactome capture [18] and RBPs with known binding specificities manually curated and reported in RBPDB [19] to perform a systematic survey of their domain composition, structural disorder, expression across 23 tissues, evolutionary conservation across 62 species and associated diseases by integrating diverse datasets in the public domain. This allowed us to not only uncover the domain architecture, expression and evolutionary dynamics of RNA-binding proteins but also provide novel insights into their roles in diverse human tissues and disease phenotypes.…”

Peroxiredoxin 1 Contributes to Host Defenses against Mycobacterium tuberculosis

“…PRDX1 can bind RNA and acts as a RNA chaperone (47,48). UV crosslinking studies suggest that PRDX1 binds RNA in cells (49)(50)(51). These reports indicate that PRDX1 binds the 59 untranslated region of c-Rel mRNA and helps translation.…”