Delineation of a cytokine gene regulatory network and rewiring in disease

细胞因子基因调控网络的描绘和疾病中的重新布线

• 批准号: 9323076
• 负责人: Juan Ignacio Fuxman Bass
• 金额: $ 24.9万
• 依托单位: BOSTON UNIVERSITY (CHARLES RIVER CAMPUS)
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-09-01 至 2019-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9323076
• 关键词: Address; Affect; Alleles; American; Area; Arthritis; Autoimmune Diseases; Autoimmunity; Award; Binding; Binding Sites; Bioinformatics; Biological Assay; Biological Response Modifiers; Cell Line; Cells; Chronic; Collaborations; Complement; Complex; Cues; Cytokine Gene; Cytokine Signaling; DNA-Protein Interaction; Data; Data Analyses; Disease; Enhancers; Environment; Failure; Follow-Up Studies; Gene Expression; Genes; Genetic Transcription; Goals; Health; Hereditary Disease; Human; Human Genetics; Hybrids; Immune; Immune System Diseases; Immune response; Immunology; Infection; Inflammatory Response; Institution; Knock-out; Laboratories; Lead; Leadership; Learning; Link; Malignant Neoplasms; Mentors; Modeling; Mutation; Nucleic Acid Regulatory Sequences; Pathologic; Pathology; Pathway interactions; Phase; Play; Positioning Attribute; Predisposition; Preparation; Process; Production; Public Health; Publications; Regulation; Regulator Genes; Research; Research Personnel; Resources; Role; Secure; Signal Transduction; Signaling Protein; System; Systemic Lupus Erythematosus; Systems Biology; Techniques; Testing; Training; Training Activity; Transcriptional Regulation; United States National Institutes of Health; Validation; Work; Writing; Yeasts; career; career development; cross disciplinary program; cytokine; experience; follow-up; genome editing; genome wide association study; in vivo; innovation; insight; interest; mutant; new therapeutic target; novel; overexpression; programs; promoter; response; skills; therapeutic target; tool; transcription factor

My long-term goal is to become an independent investigator at a major American research institution, developing cutting-edge research at the interface between immunology and systems biology. To achieve this goal my immediate objectives are: (1) to produce innovative, high quality research that will result in high impact publications that are independent of my mentor’s lines of research, (2) to acquire a new set of skills in bioinformatics, complex data analyses, immunology, and genome editing approaches to complement my existing background. Obtaining the prestigious NIH pathway to independence award would bolster my career opportunities and provide me with the means to develop and mature my own research program, plus acquire the additional training I need to succeed as an independent researcher. The proposed research encompasses my major scientific interests: integrating immune regulation with systems biology approaches to study gene expression. Cytokines are cell-to-cell signaling proteins that play fundamental roles in delineating immune responses. Cytokine production is highly regulated at the transcriptional level, and failure to express them appropriately often results in several pathologies such as autoimmune diseases, cancer and susceptibility to infections. After more than three decades of research, many aspects of cytokine expression are still not well understood, and transcriptional regulation of cytokine gene expression has not been explored on a systems level. My research will delineate the first gene-centered cytokine gene regulatory network (GRN) by comprehensively identifying the transcription factors (TFs) that regulate cytokine genes. To do so, I will first identify the TFs that can bind to the promoters and other known regulatory regions for all human cytokines by directly testing for protein-DNA interactions (PDIs) using high-throughput yeast one-hybrid (eY1H) assays, recently developed in the Walhout lab. I will model the PDIs detected into a GRN and will integrate this GRN with publicly available expression data to define which TFs positively or negatively regulate cytokine expression. These analyses, together with functional validation assays in knockout cell lines, will identify novel cytokine regulators linking them to their functional role and relevance in disease. Noncoding mutations are at the core of many human genetic diseases, and constitute 90% of all mutations identified in genome-wide association studies. For instance, mutations in cytokine regulatory regions have been associated with cancer, higher susceptibility to infections, chronic arthritis and systemic lupus erythematosus. These mutations can result in loss or gain of transcription factor binding sites. Assays to test for PDI changes are labor intensive and limited to a few dozen TFs. Thus, I propose to use a novel eY1H pipeline to systematically interrogate for PDI changes that result as a consequence of disease-associated mutations in cytokine regulatory regions. Findings from this experimental approach will be further validated in immune cell lines and will provide mechanistic insights and therapeutic targets for many immune related diseases. The cross-disciplinary program I propose requires that I obtain additional training in multiple areas of research that will complement my background, and will provide me with state-of-the-art tools for my future research as an independent investigator. I have identified Dr. Walhout and Dr. Fitzgerald as ideal mentors to carry out my proposed work, together with Dr. Wolfe and Dr. Myers as collaborators, with highly complementary expertise. Working in collaboration with these world-renowned researchers in a stimulating environment such as that of UMMS, together with the training activities my mentoring committee and I have planned for the mentored phase of the award, will enable me to: (1) become proficient in state-of-the-art immunology techniques and learn up-to-date immunology concepts, (2) acquire bioinformatics and complex data analysis skills, and (3) learn genome editing approaches, all necessary for the interpretation and validation of the proposed work. This scientific expertise will be complemented with career development seminars and activities to acquire the mentoring, leadership, grantsmanship and writing skills I will need to succeed as an independent investigator. Securing this award would place me in an ideal position to capitalize on my experience to date in the Walhout laboratory, while allowing me to define my own research niche. This award will lead to a better understanding of how immune genes are regulated in normal and pathologic conditions, which will have a broad impact on public health.

我的长期目标是成为美国主要研究机构的独立研究员， 发展免疫学和系统生物学之间的前沿研究。为了实现这一目标 目标 我的近期目标是：(1) 进行创新、高质量的研究，产生高影响力 独立于我导师的研究方向的出版物，(2) 获得一套新的技能 生物信息学、复杂数据分析、免疫学和基因组编辑方法来补充我的知识 现有背景。获得著名的 NIH 独立之路奖将促进我的职业生涯 机会并为我提供开发和成熟自己的研究计划的手段，并获得 作为一名成功的独立研究员，我需要额外的培训。 拟议的研究涵盖了我的主要科学兴趣：将免疫调节与 研究基因表达的系统生物学方法。细胞因子是细胞间信号传导蛋白，其作用 描述免疫反应的基本作用。细胞因子的产生受到高度监管 转录水平，并且未能正确表达它们通常会导致多种病理，例如 自身免疫性疾病、癌症和感染易感性。经过三十多年的研究， 细胞因子表达的许多方面仍不清楚，细胞因子的转录调控 尚未在系统水平上探索基因表达。我的研究将描绘第一个以基因为中心的 通过全面识别转录因子 (TF) 来构建细胞因子基因调控网络 (GRN) 调节细胞因子基因。为此，我将首先确定可以与启动子和其他已知的结合的转录因子 通过使用高通量直接测试蛋白质-DNA 相互作用 (PDI)，确定所有人类细胞因子的调控区域 Walhout 实验室最近开发了酵母单杂交 (eY1H) 检测方法。我将对 PDI 进行建模 检测到 GRN 中，并将将此 GRN 与公开可用的表达数据集成以定义哪些 TF 正向或负向调节细胞因子的表达。这些分析以及功能验证 在敲除细胞系中进行检测，将鉴定出新的细胞因子调节因子，将它们与其功能作用联系起来， 与疾病的相关性。 非编码突变是许多人类遗传疾病的核心，占所有遗传疾病的 90% 全基因组关联研究中发现的突变。例如，细胞因子调节区的突变 与癌症、感染、慢性关节炎和系统性狼疮的易感性较高有关 红斑狼疮。这些突变可能导致转录因子结合位点的丢失或增加。待测分析 因为 PDI 更改是劳动密集型的，并且仅限于几十个 TF。因此，我建议使用一种新颖的 eY1H 系统地询问因疾病相关而导致的 PDI 变化的管道 细胞因子调节区的突变。该实验方法的结果将在 免疫细胞系，将为许多免疫相关的疾病提供机制见解和治疗靶点 疾病。 我提出的跨学科计划要求我在多个领域获得额外的培训 研究将补充我的背景，并为我的未来提供最先进的工具 作为独立研究者进行研究。我认为沃尔豪特博士和菲茨杰拉德博士是理想的导师 与沃尔夫博士和迈尔斯博士作为合作者一起开展我提议的工作，并高度重视 互补的专业知识。与这些世界知名的研究人员合作，进行令人兴奋的研究 像 UMMS 那样的环境，以及我和我的指导委员会所进行的培训活动 计划在该奖项的指导阶段，将使我能够：（1）精通最先进的技术 免疫学技术并学习最新的免疫学概念，（2）获得生物信息学和复杂的 数据分析技能，以及（3）学习基因组编辑方法，这些都是解释和解释所必需的。 验证拟议工作。这种科学专业知识将与职业发展相辅相成 研讨会和活动以获得我需要的指导、领导、资助和写作技能 作为一名成功的独立调查员。获得这个奖项将使我处于一个理想的位置来利用 迄今为止我在沃尔豪特实验室的经历，同时让我能够定义自己的研究领域。这 该奖项将有助于更好地理解免疫基因在正常和病理状态下是如何调节的 条件，这将对公众健康产生广泛影响。