Reactivity-based RNA profiling for analyzing transcriptome interactions of small molecules in human cells

Linglan Fang; Eric T Kool

doi:10.1016/j.xpro.2023.102670

Reactivity-based RNA profiling for analyzing transcriptome interactions of small molecules in human cells

STAR Protoc. 2023 Dec 15;4(4):102670. doi: 10.1016/j.xpro.2023.102670. Epub 2023 Oct 31.

Authors

Linglan Fang¹, Eric T Kool²

Affiliations

¹ Department of Chemistry and Sarafan ChEM-H, Stanford University, Stanford, CA 94305, USA.
² Department of Chemistry and Sarafan ChEM-H, Stanford University, Stanford, CA 94305, USA. Electronic address: kool@stanford.edu.

Abstract

Protein-targeted small-molecule drugs may unintentionally bind intracellular RNA, contributing to drug toxicity. Moreover, new drugs are actively sought for intentionally targeting RNA. Here, we present a protocol to globally profile RNA-drug interactions in human cells using acylating probes and next-generation sequencing. We describe steps for cell culture, target acylation, library preparation, and sequencing. Detailed bioinformatic analyses identify drug-binding RNA loci in ∼16,000 poly(A)+ human transcripts. This streamlined workflow identifies RNA-drug interactions at single-nucleotide resolution, revealing widespread transcriptome interactions of drugs. For complete details on the use and execution of this protocol, please refer to Fang et al.¹.

Keywords: Bioinformatics; Chemistry; RNAseq.

MeSH terms

Acylation
Cell Culture Techniques*
Computational Biology
Humans
RNA / genetics
Transcriptome* / genetics

Substances

RNA

Abstract

MeSH terms

Substances

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