Abstract
Lysine 5-hydroxylation (5-Hyl) has been well recognized as an essential protein post-translational modification regulating cellular structural stability, RNA alternative splicing and epigenetic gene expression. System-wide enrichment and quantification of 5-Hyl targets have been challenging due to their chemical inert nature and difficulties in differentiating structural isomers in a complex biological sample. Here, we report the development of an efficient chemical proteomic workflow for affinity enrichment and constitutional isomer specific profiling of endogenous 5-Hyl substrates based on highly selective periodate chemistry. Our study confidently identified over 1600 5-Hyl sites on 630 proteins in human 293T cells, revealing functional significance of the modification in protein structure, transcription and chromatin regulation. Analysis of histone 5-Hyl sites showed that histones H2B and H1 are major targets of the 5-hydroxylysine epigenetic mark. Quantitative proteomic analysis through our chemical enrichment workflow identified specific 5-Hyl substrate proteins mediated by the overexpression of Jumonji-domain containing protein 6 (JMJD6). Our study uncovered two cancer-relevant alternative splice isoforms of JMJD6 that regulate 5-Hyl proteins in distinct cellular pathways, providing unique insights into the functional roles of JMJD6 alternative splicing in transcriptional regulation and cellular development.
Original language | English (US) |
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Pages (from-to) | 18395-18404 |
Number of pages | 10 |
Journal | Chemical Science |
Volume | 15 |
Issue number | 44 |
DOIs | |
State | Published - Oct 8 2024 |
Bibliographical note
Publisher Copyright:© 2024 The Royal Society of Chemistry.
PubMed: MeSH publication types
- Journal Article