The profile and dynamics of RNA modifications in animals

Abstract

More than a hundred distinct modified nucleosides have been identified in RNA, but little is known about their distribution across different organisms, dynamic nature and their response to cellular and environmental stress. Mass spectrometry based methods have been at the forefront of identifying and quantifying modified nucleosides. However, they often require synthetic reference standards, which do not exist for many modified nucleosides and therefore impedes their analysis. Here, we use a metabolic labeling approach to rapidly generate bio-isotopologues of the complete C. elegans transcriptome and its modifications, and use them as reference standards to characterize the RNA modification profile in this multicellular organism via an untargeted liquid-chromatography tandem high-resolution mass spectrometry (LC-MS/HRMS) approach. We furthermore show that several of these RNA modifications have a dynamic response to environmental stress and that in particular changes in the tRNA wobble base modification 5-methoxycarbonylmethyl-2-thiouridine (mcm5s2U) lead to codon biased gene expression changes in starved animals.CRUK Wellcome Trus