He Chongsheng, Sidoli Simone, Warneford-Thomson Robert, Tatomer Deirdre C, Wilusz Jeremy E, Garcia Benjamin A, Bonasio Roberto
Epigenetics Program, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA; Department of Cell and Developmental Biology, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA.
Epigenetics Program, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA; Department of Biochemistry and Biophysics, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA.
Mol Cell. 2016 Oct 20;64(2):416-430. doi: 10.1016/j.molcel.2016.09.034.
Interactions between noncoding RNAs and chromatin proteins play important roles in gene regulation, but the molecular details of most of these interactions are unknown. Using protein-RNA photocrosslinking and mass spectrometry on embryonic stem cell nuclei, we identified and mapped, at peptide resolution, the RNA-binding regions in ∼800 known and previously unknown RNA-binding proteins, many of which are transcriptional regulators and chromatin modifiers. In addition to known RNA-binding motifs, we detected several protein domains previously unknown to function in RNA recognition, as well as non-annotated and/or disordered regions, suggesting that many functional protein-RNA contacts remain unexplored. We identified RNA-binding regions in several chromatin regulators, including TET2, and validated their ability to bind RNA. Thus, proteomic identification of RNA-binding regions (RBR-ID) is a powerful tool to map protein-RNA interactions and will allow rational design of mutants to dissect their function at a mechanistic level.
非编码RNA与染色质蛋白之间的相互作用在基因调控中发挥着重要作用，但这些相互作用的大多数分子细节尚不清楚。我们利用蛋白质-RNA光交联和质谱技术对胚胎干细胞细胞核进行研究，以肽段分辨率鉴定并绘制了约800种已知和先前未知的RNA结合蛋白中的RNA结合区域，其中许多是转录调节因子和染色质修饰因子。除了已知的RNA结合基序，我们还检测到几个以前未知在RNA识别中起作用的蛋白质结构域，以及未注释和/或无序区域，这表明许多功能性蛋白质-RNA相互作用仍有待探索。我们在包括TET2在内的几种染色质调节因子中鉴定出RNA结合区域，并验证了它们结合RNA的能力。因此，RNA结合区域的蛋白质组学鉴定（RBR-ID）是绘制蛋白质-RNA相互作用的有力工具，将有助于合理设计突变体，从机制层面剖析其功能。
