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Cytokine Gene Regulation by Modification of Arginine Residues

通过精氨酸残基修饰进行细胞因子基因调控

基本信息

批准号：
8075289
负责人：
KERRI A MOWEN
金额：
$ 1.1万
依托单位：
SCRIPPS RESEARCH INSTITUTE, THE
依托单位国家：
美国
项目类别：
财政年份：
2010
资助国家：
美国
起止时间：
2010-06-01 至 2010-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8075289
关键词：
Allergic Amino Acid Sequence Amino Acids Antibodies Arginine Arginine deiminase Autoimmune Diseases Autoimmunity Binding Biological Assay Cell physiology Cells Citrulline Cytokine Gene Data Ectopic Expression Event Exons Gene Expression Regulation Gene Targeting Glutamine Histones Human Hydrolysis Immune Immune response Immunologic Receptors In Vitro Inflammatory Interleukin-4 Ligation Link Lymphocyte Mass Spectrum Analysis Mediating Methylation Modification Molecular Mono-S Mus Mutate Nuclear Protein Nuclear Proteins Pattern Phenotype Positioning Attribute Post-Translational Protein Processing Predisposition Production Protein-arginine deiminase Proteins Receptor Signaling Regulation Reporter Genes Research Personnel Rheumatoid Arthritis Role Signal Pathway Site T-Cell Receptor T-Lymphocyte TRAF2 gene Testing Th2 Cells arginine methyltransferase base cell type cofactor cytokine genetic regulatory protein in vivo methyl group mutant novel overexpression prevent programs response tool

项目摘要

Cytokine production by the Th1 and Th2 subsets have been associated with susceptiblity to infectious, allergic, and autoimmune diseases. Understanding the molecular events which control lineage-specific cytokine expression would provide useful tools to modulate the Th1/Th2 response. IL- 4 production by Th2 cells is regulated by NFAT and the NFAT cofactor, NIP45. We have shown that arginine methylation of NIP45 facilitates its interaction with NFAT and augments IL-4transcription. Our data positioned the arginine methyltransferase PRMT1 downstream of the T cell receptor, suggesting that arginine methylation may be an important modification in immune receptor signaling pathways. Arginine methylation is countered by the actions of peptidylarginine deiminase 4 (PAD4). The five PAD enzymes convert arginine residues within proteins into the atypical amino acid citrulline. PAD4 is expressed mainly in lymphocytes. We have found that NIP45 methylation is negatively regulated by PAD4 via citrullination of arginine residues, which prevents the ability of NIP45 to be methylated by PRMT1. PAD4 expression dramatically reduces NIP45-induced IL-4promoter activity. Therefore, we hypothesize that through actions on NIP45 and other regulatory proteins that PAD4 controls Th cell cytokine expression.The specific aimsare: 1) Determine the mechanism by which PAD4 antagonizes NIP45-mediatedinduction of Th cell cytokine production. We will (i) determine the modified arginine residues in NIP45 by mass spectrometry, (ii) determine how PAD4 influences NIP45 activity by studying the effects of PAD4 on the NIP45/NFAT interaction, (iii) determine whether citrullination within NIP45 serves a function other than preventing methylation. 2) Investigate the regulation of PAD4 activity. We will: (i) determine the PAD4 expression pattern in Th cells, (ii) determine whether PAD4 activity is regulated in Th cells, (iii) determine whether PAD4, itself, is regulated by arginine methylation. 3) Analyzethe phenotype of PAD4 deficient mice. We will create mice in which exons 7-13 of PAD4 are flanked by loxP sites so that we can ablate PAD4 expression in the T lineage using CD4-Cre Tg mice. These mice will allow us to determine the role of PAD4 in Th cell function.

Th1 和 Th2 亚群产生的细胞因子与对以下疾病的易感性相关：传染性、过敏性和自身免疫性疾病。了解控制的分子事件谱系特异性细胞因子表达将为调节 Th1/Th2 反应提供有用的工具。白细胞介素- Th2 细胞产生的 4 蛋白受 NFAT 和 NFAT 辅助因子 NIP45 调节。我们已经证明 NIP45 的精氨酸甲基化促进其与 NFAT 的相互作用并增强 IL-4 转录。我们的数据将精氨酸甲基转移酶 PRMT1 定位在 T 细胞受体的下游，表明精氨酸甲基化可能是免疫受体信号传导的重要修饰途径。精氨酸甲基化可通过肽基精氨酸脱亚胺酶 4 (PAD4) 的作用来抵消。五种 PAD 酶将蛋白质内的精氨酸残基转化为非典型氨基酸瓜氨酸。 PAD4主要在淋巴细胞中表达。我们发现NIP45甲基化呈负向 PAD4 通过精氨酸残基的瓜氨酸化来调节，从而阻止 NIP45 被被 PRMT1 甲基化。 PAD4 表达显着降低 NIP45 诱导的 IL-4 启动子活性。因此，我们假设通过对 NIP45 和其他调节蛋白的作用 PAD4控制Th细胞细胞因子的表达。具体目标是： 1) 确定PAD4拮抗NIP45介导的Th细胞诱导的机制细胞因子的产生。我们将 (i) 按质量确定 NIP45 中修饰的精氨酸残基光谱分析法，(ii) 通过研究 PAD4 的影响来确定 PAD4 如何影响 NIP45 活性关于 NIP45/NFAT 相互作用，(iii) 确定 NIP45 内的瓜氨酸化是否起到除了防止甲基化之外还有其他功能。 2) 研究PAD4活性的调节。我们将： (i) 确定 PAD4 表达模式在 Th 细胞中，(ii) 确定 PAD4 活性是否在 Th 细胞中受到调节，(iii) 确定是否 PAD4 本身受精氨酸甲基化调节。 3)分析PAD4缺陷小鼠的表型。我们将创建小鼠，其中外显子 7-13 PAD4 两侧是 loxP 位点，因此我们可以使用以下方法消除 T 谱系中的 PAD4 表达 CD4-Cre Tg 小鼠。这些小鼠将使我们能够确定 PAD4 在 Th 细胞功能中的作用。