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标题：Widespread hydroxylation of unstructured lysine-rich protein domains by JMJD6
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作者：Matthew E. Cockman ; Yoichiro Sugimoto ; Hamish B. Pegg 等
期刊名称：Proceedings of the National Academy of Sciences
印刷版ISSN：0027-8424
电子版ISSN：1091-6490
出版年度：2022
卷号：119
期号：32
DOI：10.1073/pnas.2201483119
语种：English
出版社：The National Academy of Sciences of the United States of America
摘要：Significance Enzyme-catalyzed posttranslational hydroxylation is increasingly recognized as affecting intracellular proteins and their functions, including in the signaling of hypoxic stress. The current work applies mass spectrometry methods to reveal more than 100 sites of lysine hydroxylation catalyzed by the 2-oxoglutarate–dependent dioxygenase JMJD6 in regions of the proteome that are refractory to conventional trypsin-based analyses. These sites were distributed across proteins involved at multiple levels in the control of gene expression. Targeted regions were strongly enriched for unstructured lysine-rich regions that have been associated with biomolecular condensates. The findings open routes to the understanding of the biological roles of JMJD6, including in subcellular reorganization in response to stress. The Jumonji domain–containing protein JMJD6 is a 2-oxoglutarate–dependent dioxygenase associated with a broad range of biological functions. Cellular studies have implicated the enzyme in chromatin biology, transcription, DNA repair, mRNA splicing, and cotranscriptional processing. Although not all studies agree, JMJD6 has been reported to catalyze both hydroxylation of lysine residues and demethylation of arginine residues. However, despite extensive study and indirect evidence for JMJD6 catalysis in many cellular processes, direct assignment of JMJD6 catalytic substrates has been limited. Examination of a reported site of proline hydroxylation within a lysine-rich region of the tandem bromodomain protein BRD4 led us to conclude that hydroxylation was in fact on lysine and catalyzed by JMJD6. This prompted a wider search for JMJD6-catalyzed protein modifications deploying mass spectrometric methods designed to improve the analysis of such lysine-rich regions. Using lysine derivatization with propionic anhydride to improve the analysis of tryptic peptides and nontryptic proteolysis, we report 150 sites of JMJD6-catalyzed lysine hydroxylation on 48 protein substrates, including 19 sites of hydroxylation on BRD4. Most hydroxylations were within lysine-rich regions that are predicted to be unstructured; in some, multiple modifications were observed on adjacent lysine residues. Almost all of the JMJD6 substrates defined in these studies have been associated with membraneless organelle formation. Given the reported roles of lysine-rich regions in subcellular partitioning by liquid–liquid phase separation, our findings raise the possibility that JMJD6 may play a role in regulating such processes in response to stresses, including hypoxia.
关键词：enJMJD6lysine hydroxylationlysine-rich domain2-oxoglutarate-dependent dioxygenasehypoxia