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PRMT5 dimethylates R30 of the p65 subunit to activate NF-?B.

ABSTRACT: The ubiquitous inducible transcription factor NF-?B plays central roles in immune and inflammatory responses and in tumorigenesis. Complex posttranslational modifications of the p65 subunit (RelA) are a major aspect of the extremely flexible regulation of NF-?B activity. Although phosphorylation, acetylation, ubiquitination, and lysine methylation of NF-?B have been well described, arginine methylation has not yet been found. We now report that, in response to IL-1?, the p65 subunit of NF-?B is dimethylated on arginine 30 (R30) by protein-arginine methyltransferase 5 (PRMT5). Expression of the R30A and R30K mutants of p65 substantially decreased the ability of NF-?B to bind to ?B elements and to drive gene expression. A model in which dimethyl R30 is placed into the crystal structure of p65 predicts new van der Waals contacts that stabilize intraprotein interactions and indirectly increase the affinity of p65 for DNA. PRMT5 was the only arginine methyltransferase that coprecipitated with p65, and its overexpression increased NF-?B activity, whereas PRMT5 knockdown had the opposite effect. Microarray analysis revealed that ?85% of the NF-?B-inducible genes that are down-regulated by the R30A mutation are similarly down-regulated by knocking PRMT5 down. Many cytokine and chemokine genes are among these, and conditioned media from cells expressing the R30A mutant of p65 had much less NF-?B-inducing activity than media from cells expressing the wild-type protein. PRMT5 is overexpressed in many types of cancer, often to a striking degree, indicating that high levels of this enzyme may promote tumorigenesis, at least in part by facilitating NF-?B-induced gene expression.

SUBMITTER: Wei H

PROVIDER: S-EPMC3746871 | biostudies-literature | 2013 Aug

REPOSITORIES: biostudies-literature

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PRMT5 dimethylates R30 of the p65 subunit to activate NF-κB.

Wei Han H Wang Benlian B Miyagi Masaru M She Yun Y Gopalan Banu B Huang De-Bin DB Ghosh Gourisankar G Stark George R GR Lu Tao T

Proceedings of the National Academy of Sciences of the United States of America 20130731 33

The ubiquitous inducible transcription factor NF-κB plays central roles in immune and inflammatory responses and in tumorigenesis. Complex posttranslational modifications of the p65 subunit (RelA) are a major aspect of the extremely flexible regulation of NF-κB activity. Although phosphorylation, acetylation, ubiquitination, and lysine methylation of NF-κB have been well described, arginine methylation has not yet been found. We now report that, in response to IL-1β, the p65 subunit of NF-κB is ...[more]

PMID: 23904475

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Project description:Background: Atherosclerosis leads high mortality, highlighting an urgent need for new therapeutic strategies. Protein kinases orchestrate multiple cellular events in atherosclerosis and may provide new therapeutic targets for atherosclerosis. Here, we identified a protein kinase, WEE1 G2 checkpoint kinase (WEE1), promoting inflammatory response in atherosclerosis. Methods: The ApoE-/- mice without macrophage-specific WEE1 deletion (ApoE-/-WEE1f/f) and ApoE-/- mice with macrophage-specific WEE1 deletion (ApoE-/-WEE1MCKO) were generated using bone marrow transplantation. These mice and WEE1 inhibitor MK1775 were used in a high-fat diet (HFD)-induced atherosclerotic model. Human atherosclerotic tissues, mouse primary peritoneal macrophages (MPMs), 293T cells, and recombinant proteins were utilized to investigate the pathogenic role and underlying mechanisms of WEE1. Results: We identified up-regulated p-WEE1 in macrophages in atherosclerotic lesions of HFD-fed ApoE-/- mice. Transcriptome sequencing analysis indicated that WEE1 promotes oxLDL-induced inflammation in macrophages. We then demonstrated that macrophage-specific deletion or pharmacological inhibition of WEE1 attenuates atherosclerosis by reducing inflammation both in vivo and in vitro. The overexpression of wild-type WEE1 or activating-mutant WEE1, but not inactivating-mutant WEE1, exacerbates inflammation in macrophages. Mechanistically, transcriptome sequencing analysis and co-immunoprecipitation followed by quantitative proteomics analysis identified p65 as a binding protein of WEE1. We confirmed that WEE1 directly interacts with the RHD domain of p65 via kinase domain and phosphorylates p65 at S536, thereby facilitating subsequent NF-κB activation and inflammatory response in macrophages. Conclusions: Our findings demonstrated that macrophage WEE1 promotes inflammatory atherosclerosis by directly binding to p65 and phosphorylate it at S536. This study provides WEE1 as a new p65 regulator in inflammation and a potential therapeutic target for atherosclerosis.

Dataset Information

PRMT5 dimethylates R30 of the p65 subunit to activate NF-?B.

Publications

PRMT5 dimethylates R30 of the p65 subunit to activate NF-κB.

Similar Datasets

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