PXD029955-2

PXD029955 is an original dataset announced via ProteomeXchange.

Title	Reactivity-dependent profiling of RNA 5-methylcytidine dioxygenases
Description	Epitranscriptomic RNA modifications can regulate fundamental biological processes, but we lack approaches to map modification sites and probe writer enzymes. Here we present a chemoproteomic strategy to characterize RNA 5-methylcytidine (m5C) dioxygenase enzymes in their native context based upon metabolic labeling and activity-based crosslinking with 5-ethynylcytidine (5-EC), RNA-protein enrichment, and quantitative proteomics. We profile m5C dioxygenases in human cells including ALKBH1 and TET2 and use quantitative nucleoside LC-MS to show that ALKBH1 is the major hm5C and f5C-forming enzyme in RNA, including upon polyadenylated RNA. Further, we map ALKBH1 modification sites transcriptome-wide using 5-EC-based iCLIP analysis to show that ALKBH1 oxidizes m5C in a variety of tRNA anticodon stem loops (ASL), as well as on mRNA and lncRNA, and analyze its substrate specificity using in vitro enzymatic assays. Finally, we apply pyridine borane-mediated sequencing to identify f5C sites in tRNA ASLs. Our work provides powerful chemical approaches for studying RNA m5C dioxygenases and mapping oxidative m5C modifications and reveals the existence of novel epitranscriptomic pathways for regulating RNA function.
HostingRepository	PRIDE
AnnounceDate	2023-11-14
AnnouncementXML	Submission_2023-11-14_08:54:25.242.xml
DigitalObjectIdentifier
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Unsupported dataset by repository
PrimarySubmitter	Emilia Arguello
SpeciesList	scientific name: Homo sapiens (Human); NCBI TaxID: 9606;
ModificationList	No PTMs are included in the dataset
Instrument	Orbitrap Fusion Lumos

Revision	Datetime	Status	ChangeLog Entry
0	2021-11-26 03:46:07	ID requested
1	2022-07-16 08:48:51	announced
⏵ 2	2023-11-14 08:54:25	announced	2023-11-14: Updated project metadata.

Dataset with its publication pending

submitter keyword: RNA modification, metabolic RNA labeling, m5C,human, LC-MS/MS

Ralph E. Kleiner
contact affiliation	Department of Chemistry, Princeton University, Princeton, NJ 08544, USA
contact email	rkleiner@princeton.edu
lab head
Emilia Arguello
contact affiliation	Princeton University Chemistry Department
contact email	earguello@princeton.edu
dataset submitter

Dataset FTP location NOTE: Most web browsers have now discontinued native support for FTP access within the browser window. But you can usually install another FTP app (we recommend FileZilla) and configure your browser to launch the external application when you click on this FTP link. Or otherwise, launch an app that supports FTP (like FileZilla) and use this address: ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2022/07/PXD029955

PRIDE project URI

• PRIDE
  1. PXD029955
     1. Label: PRIDE project
     2. Name: Reactivity-dependent profiling of RNA 5-methylcytidine dioxygenases