Chromatin Remodeling and Lymphocyte Regulation

染色质重塑和淋巴细胞调节

• 批准号: 8335873
• 负责人: MICHAEL J PAZIN
• 金额: $ 40.25万
• 依托单位: NATIONAL INSTITUTE ON AGING
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/8335873
• 关键词: ATP phosphohydrolase; Ascaridil; Asthma; Autoimmune Diseases; Autoimmunity; Bibliography; Binding; Biochemistry; Biological; CD4 Positive T Lymphocytes; Cell Differentiation process; Cell Line; Cells; Cellular biology; Chromatin; Chromatin Structure; Code; Complex; Cytokine Gene; Data; Deposition; Distal; Elements; Enhancers; Enzymes; Fibroblasts; Future; Gene Cluster; Gene Expression; Gene Expression Regulation; Genes; Genetic Transcription; Granulocyte-Macrophage Colony-Stimulating Factor; Helper-Inducer T-Lymphocyte; Hypersensitivity; ISWI; Interleukin-3; Knock-out; Link; Lymphocyte; Manuscripts; MicroRNAs; Mus; NF-kappa B; Nucleic Acid Regulatory Sequences; Play; Printing; Proteins; Publications; Publishing; Reading; Recruitment Activity; Regulation; Regulatory Element; Reporter; Role; SMARCA4 gene; SMARCA5 gene; STAT6 gene; Signal Transduction; Site; System; T cell differentiation; T cell regulation; T-Lymphocyte; T-Lymphocyte Subsets; Technology; Testing; Th2 Cells; Undifferentiated; Vaccines; Work; atopy; base; chromatin remodeling; fighting; genome-wide; genome-wide analysis; novel; p65; pathogen; programs; promoter; research study; response; therapeutic target; transcription factor

Previously, we found that chromatin remodeling was required for Th2 cytokine gene expression. We found that transcription factors recruited the SWI/SNF remodeling enzyme BRG1 to specific sites in Th2 cells. BRG1 was required before and after Th2 differentiation. We found the ISWI remodeling enzyme SNF2H could activate and repress gene expression in T cells. Progress: T helper cell differentiation and activation require specific transcriptional programs accompanied by changes in chromatin structure. However, little is known about the chromatin remodeling enzymes responsible. We performed genome-wide analysis to determine the general principles of BRG1 binding, followed by analysis of specific genes to determine whether these general rules were typical of key T cell genes. BRG1 binding was to localized, enhancer-sized regions, a finding subsequently confirmed by two other labs. We found that binding of the remodeling protein BRG1 was programmed by both lineage and activation signals. BRG1 binding positively correlated with gene activity at protein-coding and microRNA (miRNA) genes. BRG1 binding was found at promoters and distal regions, including both novel and previously validated distal regulatory elements. Distal BRG1 binding correlated with expression, and novel distal sites in the Gata3 locus possessed enhancer-like activity, suggesting a general role for BRG1 in long-distance gene regulation. One of these regions was tested by an independent lab, and confirmed. BRG1 recruitment to distal sites in Gata3 was impaired in cells lacking STAT6, a transcription factor that regulates lineage-specific genes. Together, these findings suggest that BRG1 interprets both differentiation and activation signals and plays a causal role in gene regulation, chromatin structure, and cell fate. Our findings suggest that BRG1 binding is a useful marker for identifying active cis-regulatory regions in protein-coding and miRNA genes. We investigated gene regulation at the IL-3/GM-CSF gene cluster. We found BRG1, a SWI/SNF remodeling ATPase, bound a novel distal element, CNSa. BRG1 binding was strongest in differentiated, stimulated T helper cells, paralleling IL-3 and GM-CSF expression. Depletion of BRG1 reduced IL-3 and GM-CSF transcription. BAF-specific SWI/SNF subunits bound to this locus and regulated IL-3 expression. CNSa was in closed chromatin in fibroblasts, open chromatin in differentiated T helper cells, and moderately open chromatin in nave (undifferentiated) T helper cells; BRG1 was required for the most open state. CNSa increased transcription of a reporter in an episomal expression system, in a BRG1-dependent manner. The NF-kB subunit RelA/p65 bound CNSa in activated T helper cells. Inhibition of NF-kB blocked BRG1 binding to CNSa, chromatin opening at CNSa, and activation of IL-3 and GM-CSF. Together, these findings suggest CNSa is a distal enhancer that binds BRG1 and NF-kB. This publication is in the NCBI pipeline for PMC, and so may not be listed in the bibliography: NF-kappaB and BRG1 bind a distal regulatory element in the IL-3/GM-CSF locus. Wurster AL, Precht P, Pazin MJ. Mol Immunol. 2011 Aug 8. Epub ahead of print PMID: 21831442 This publication (invited review) is in press (to appear online by October 2011), so it cannot listed in the bibliography: ATP-dependent chromatin remodeling in T cells Andrea L. Wurster and Michael J. Pazin Biochemistry and Cell Biology, Manuscript ID 2011-0042 Genome-wide BRG1 binding data in 6 murine T cell subsets (Raw sequence reads, aligned tags BRG1 binding regions and ChIP-seq stats for all T helper subsests) were deposited in GEO, and are publicly available at accession number GSE23719. Comprehensive gene expression data in 4 murine T helper subsets, WT and BAF180 deficient cells, are at GEO (GSE31676) and will be publicly released upon publication of the BAF180 described work below. Our results provide new information on the regulation of T cell fate and function by chromatin remodeling enzymes. This work could be important in understanding the biological basis of, and identifying therapeutic targets for, autoimmune disorders, allergy and atopy, and vaccine response. Future: We will publish our study of BAF180, a SWI/SNF subunit specific to the PBAF form of the SWI/SNF complex in T helper cells using conditional knockout technology. No further experiments are planned.

以前，我们发现Th2细胞因子基因表达需要染色质重塑。 我们发现，转录因子募集了SWI/SNF重塑酶BRG1到Th2细胞中的特定位点。 Th2分化之前和之后需要BRG1。我们发现ISWI重塑酶SNF2H可以激活和抑制T细胞中的基因表达。 进步： T辅助细胞分化和激活需要特定的转录程序，并伴有染色质结构的变化。但是，对负责的染色质重塑酶知之甚少。我们进行了全基因组分析以确定BRG1结合的一般原理，然后对特定基因进行分析，以确定这些一般规则是否是关键T细胞基因的典型代表。 BRG1结合是与局部增强子大小的区域，这一发现随后由另外两个实验室确认。 我们发现重塑蛋白BRG1的结合均通过谱系和激活信号编程。 BRG1结合与蛋白质编码和microRNA（miRNA）基因的基因活性正相关。在启动子和远端区域发现了BRG1结合，包括新型和先前经过验证的远端调节元件。远端BRG1结合与表达相关，而GATA3基因座中的新远端位点具有增强子样活性，这表明BRG1在长距离基因调节中的一般作用。这些区域之一由独立实验室测试并确认。 在缺乏STAT6的细胞（一种调节谱系特异性基因的转录因子）的细胞中，BRG1募集到GATA3中的远端位点受损。总之，这些发现表明BRG1既解释分化和激活信号，又在基因调节，染色质结构和细胞命运中起因果作用。我们的发现表明，BRG1结合是鉴定蛋白质编码和miRNA基因中活性顺式调节区域的有用标记。 我们研究了IL-3/GM-CSF基因簇的基因调节。我们找到了BRG1，SWI/SNF重塑 ATPase，结合了新型远端元素CNSA。 BRG1结合在分化的，刺激的T辅助器中最强 平行IL-3和GM-CSF表达的细胞。 BRG1的耗竭降低了IL-3和GM-CSF转录。 BAF特异性的SWI/SNF亚基与该基因座结合并调节IL-3表达。 CNSA已关闭 成纤维细胞中的染色质，在分化的T辅助细胞中打开染色质，然后开放染色质 在中殿（未分化的）T辅助细胞中；最开放状态需要BRG1。 CNSA增加了 以BRG1依赖性方式在偶发表达系统中的记者转录。 NF-KB 激活的T辅助细胞中的亚基Rela/p65结合CNSA。 NF-KB抑制BRG1结合到 CNSA，CNSA的染色质开放以及IL-3和GM-CSF的激活。这些发现一起暗示 CNSA是结合BRG1和NF-KB的远端增强子。 该出版物在PMC的NCBI管道中，因此在书目中可能不会列出： NF-KAPPAB和BRG1结合IL-3/GM-CSF基因座中的远端调节元件。 Wurster AL，Precht P，Pazin MJ。 摩尔免疫。 2011年8月8日。 PMID：21831442 该出版物（邀请评论）在媒体上（将在2011年10月在线出现），因此在书目中无法列出： T细胞中依赖ATP的染色质重塑 Andrea L. Wurster和Michael J. Pazin 生物化学和细胞生物学，手稿ID 2011-0042 全基因组BRG1结合数据在6个鼠T细胞子集中（原始序列读取，对齐的标签BRG1结合区域的比对标签和所有t辅助子电池的芯片seq统计数据）存放在GEO中，并且在登录号GSE23719上公开可用。 4个鼠T助手子集，WT和BAF180缺陷细胞中的全面基因表达数据位于GEO（GSE31676），并将在下面描述的BAF180发布后公开发布。 我们的结果提供了有关通过染色质重塑酶调节T细胞命运和功能的新信息。 这项工作对于理解自身免疫性疾病，过敏和特应性以及疫苗反应的治疗靶点的生物学基础和识别的生物学基础可能很重要。 未来： 我们将使用条件基因敲除技术发布对BAF180的研究，这是T辅助细胞中SWI/SNF复合物的PBAF形式的SWI/SNF亚基。 没有计划进一步的实验。