Immunoprecipitation and Sequencing of Acetylated RNA

Daniel Arango; David Sturgill; Shalini Oberdoerffer

doi:10.21769/BioProtoc.3278

Immunoprecipitation and Sequencing of Acetylated RNA

Bio Protoc. 2019 Jun 20;9(12):e3278. doi: 10.21769/BioProtoc.3278.

Authors

Daniel Arango¹, David Sturgill¹, Shalini Oberdoerffer¹

Affiliation

¹ Laboratory of Receptor Biology and Gene Expression, National Cancer Institute, NIH, Bethesda, MD 20892, USA.

Abstract

Generation of the epitranscriptome through chemical modifications of protein-coding messenger RNAs (mRNAs) has emerged as a new mechanism of post-transcriptional gene regulation. While most mRNA modifications are methylation events, a single acetylated ribonucleoside has been described in eukaryotes, occurring at the N4-position of cytidine (N4-acetylcytidine or ac4C). Using a combination of antibody-based enrichment of acetylated regions and deep sequencing, we recently reported ac4C as a novel mRNA modification that is catalyzed by the N-acetyltransferase enzyme NAT10. In this protocol, we describe in detail the procedures to identify acetylated mRNA regions transcriptome-wide using acetylated RNA immunoprecipitation and sequencing (acRIP-seq).

Keywords: Epitranscriptome; N4-acetylcytidine; ac4C; acRIP-seq; mRNA modifications.