MassIVE MSV000096084

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A sequence-specific RNA acetylation catalyst

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Description

N4-acetylcytidine (ac4C) is a ubiquitous RNA modification incorporated by cytidine acetyltransferase enzymes. Here, we report the biochemical characterization of Thermococcus kodakarensis Nat10 (TkNat10), an RNA acetyltransferase involved in archaeal thermotolerance. We demonstrate that TkNat10's catalytic activity is critical for T. kodakarensis fitness at elevated temperatures. Unlike eukaryotic homologs, TkNat10 exhibits robust stand-alone activity, modifying diverse RNA substrates in a temperature, ATP, and acetyl-CoA-dependent manner. Transcriptome-wide analysis reveals TkNat10 preferentially modifies unstructured RNAs containing a 5'-CCG-3' consensus sequence. Using a high-throughput mutagenesis approach, we define sequence and structural determinants of TkNat10 substrate recognition. We find TkNat10 can be engineered to use non-native acyl-CoA donors, providing insight into its cofactor specificity. Finally, we demonstrate TkNat10's utility for site-specific acetylation of RNA oligonucleotides, enabling analysis of ac4C-dependent RNA-protein interactions. Our findings establish a framework for understanding archaeal RNA acetylation and a new tool for studying the functional consequences of ac4C in diverse RNA contexts. [doi:10.25345/C5P26QF9J] [dataset license: CC0 1.0 Universal (CC0 1.0)]

Keywords: ac4C, TkNat10, acetylation, RNA ; DatasetType:Proteomics

Contact

Principal Investigators:
(in alphabetical order) 		Jordan Meier, National Cancer Institute, USA
Submitting User: ronholes7059

Publications

Thalalla Gamage S, Howpay Manage S, Sas-Chen A, Nir R, Burkhart BW, Jhulki I, Link CN, Penikalapati MS, Jones JE, Iyer LM, Aravind L, Santangelo TJ, Schwartz S, Meier JL.
A sequence-specific RNA acetylation catalyst.
bioRxiv. Epub 2024 Oct 26.

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