Roles of DDX5 and its associated long non-coding RNAs in RORgt-mediated host immune system functions

DDX5及其相关长非编码RNA在RORgt介导的宿主免疫系统功能中的作用

基本信息

批准号：
9179587
负责人：
Dan Littman
金额：
$ 53.63万
依托单位：
NEW YORK UNIVERSITY SCHOOL OF MEDICINE
依托单位国家：
美国
项目类别：
财政年份：
2015
资助国家：
美国
起止时间：
2015-12-01 至 2020-11-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9179587
关键词：
Adipocytes Adipose tissue Animals Attenuated Autoimmune Diseases Autoimmune Process Autoimmunity Binding Biochemical Bone Growth Bone Marrow CD8-Positive T-Lymphocytes Cartilage Cartilage Diseases Cell Differentiation process Cell Lineage Cells Cellular Immunity Chimera organism Chromatin Circadian Rhythms Citrobacter rodentium Colitis Complex Crohn&apos s disease Defect Dependence Dependency Development Disadvantaged Disease Dysplasia Epithelial Evaluation Experimental Autoimmune Encephalomyelitis Gene Expression Gene Targeting Genes Genetic Transcription Genome Hair Helper-Inducer T-Lymphocyte Hematopoietic Hepatocyte Hereditary Disease Human Human Genetics Immune Immune system In Vitro Infection Inflammation Inflammatory Inflammatory Bowel Diseases Interleukin-17 Intestines Knock-in Liver Lymphoid Cell Lymphoid Tissue Macromolecular Complexes Mediating Metabolic Modeling Multiple Sclerosis Mus Mutant Strains Mice Mutation Neurons Nuclear Nuclear Receptors Nucleotides Pathogenesis Pathway interactions Patients Pharmacology Protein Isoforms Proteins Proteomics Psoriasis RNA Helicase Recombinant Proteins Regulation Resistance Rheumatoid Arthritis Role Site T cell differentiation T-Lymphocyte Therapeutic Thymocyte Development Tissues Transcription Coactivator Transgenic Organisms Untranslated RNA base cell type conditional mutant cytokine design energy balance genome-wide immune system function improved in vitro Assay in vivo insight interest microbiota mutant orphan nuclear receptor ROR-gamma polarized cell programs targeted treatment thymocyte transcription factor transcriptomics γδ T cells

项目摘要

PROJECT SUMMARY/ABSTRACT The nuclear receptor RORγt directs the differentiation of T cells that have critical roles in multiple autoimmune and inflammatory diseases. Pharmacologic targeting of RORγt is hence being developed to treat diseases such as psoriasis, rheumatoid arthritis, and inflammatory bowel disease that are thought to be mediated in large part by Th17 cells and related cells that produce IL-17 cytokines. However, RORγt has multiple other functions in diverse cell types, including mediating survival of thymocytes and regulating the development of lymphoid tissue inducer (LTi) cells, that guide the formation of secondary and tertiary lymphoid tissues, and of type 3 innate lymphoid cells (ILC3) that protect epithelial barriers. In addition, RORγ, a closely related isoform encoded by the same gene, has metabolic functions in multiple tissues, including liver and adipocytes. How RORγ and RORγt function in different cell types to direct distinct transcriptional and functional programs is not understood. The transcriptional networks regulated by RORγt and multiple other transcription factors have been studied in Th17 cells polarized in vitro with combinations of cytokines, but how RORγ/γt functions in Th17 cells and other cell types in vivo is not yet known. We have used a proteomics approach to characterize macromolecular complexes that govern RORγ/γt functions in different cell types and identified DDX5, a DEAD- box RNA helicase, as associated with RORγt in Th17 cells. In mice deficient for DDX5 in T cells, expression of multiple RORγt target genes is attenuated and the animals are resistant to Th17 cell-mediated inflammatory disease. We also found the long noncoding (lnc) RNA Rmrp associated with DDX5 and with RORγt, both in Th17 cells and when all three components were synthesized in vitro. Mutations in Rmrp result in cartilage-hair hypoplasia (CHH), a recessive human genetic disease characterized by abnormal bone growth, immune deficiency and neuronal dysplasia in the intestine. Expression of wild-type, but not a CHH mutant, Rmrp promoted Th17 cell differentiation in a DDX5-dependent manner, indicating that this is a rare lncRNA that functions in trans. Our results suggest that Rmrp binding to DDX5 promotes the interaction of DDX5 with RORγt and coactivation of RORγt target genes. We propose in Specific Aim 1 to identify gene targets whose expression is dependent on DDX5, Rmrp, and RORγt, using genome-wide transcriptomics, chromatin accessibility, and chromatin occupancy studies with in vitro polarized or in vivo generated Th17 cells from wild type and mutant animals. In Specific Aim 2, we will determine, using biochemical approaches, how these molecules interact with each other and whether this interaction is restricted to Th17 cells or is present in other RORγ/γt-dependent cells. In Specific Aim 3, we will characterize the influence of the interactions in Th17 cell- mediated inflammation, ILC3-dependent barrier protection, and development of thymocytes and lymphoid tissues. Our results will provide a better understanding of the functions of RORγt, which may facilitate design of more specific therapeutics for autoimmune disease and for immune deficiency in CHH patients.

项目概要/摘要核受体 RORγt 指导 T 细胞的分化，在多种自身免疫性疾病中发挥关键作用因此，正在开发 RORγt 的药理学靶向来治疗疾病。例如牛皮癣、类风湿性关节炎和炎症性肠病，这些疾病被认为是由大部分由 Th17 细胞和产生 IL-17 细胞因子的相关细胞产生，但是，RORγt 还有多种其他细胞因子。在多种细胞类型中发挥作用，包括介导胸腺细胞的存活和调节胸腺细胞的发育淋巴组织诱导（LTi）细胞，引导二级和三级淋巴组织的形成，以及保护上皮屏障的 3 型先天淋巴细胞 (ILC3) 此外，RORγ 是一种密切相关的亚型。由同一基因编码，在多种组织中具有代谢功能，包括肝脏和脂肪细胞。 RORγ 和 RORγt 在不同的细胞类型中发挥作用，以指导不同的转录和功能程序。 RORγt 和多种其他转录因子调控的转录网络已被了解。已在体外用细胞因子组合极化的 Th17 细胞中进行了研究，但 RORγ/γt 在 Th17 中如何发挥作用细胞和其他细胞类型在体内的情况尚不清楚，我们已经使用蛋白质组学方法来表征。控制不同细胞类型中 RORγ/γt 功能的大分子复合物，并鉴定出 DDX5（一种 DEAD- box RNA 解旋酶，与 Th17 细胞中的 RORγt 相关。在 T 细胞中缺乏 DDX5 的小鼠中，表达多个 RORγt 靶基因减弱，动物对 Th17 细胞介导的炎症具有抵抗力我们还在疾病中发现了与 DDX5 和 RORγt 相关的长非编码 (lnc) RNA Rmrp。 Th17 细胞以及当所有三种成分均在体外合成时，Rmrp 突变会导致软骨毛。发育不全（CHH），一种隐性人类遗传病，其特征是骨骼生长异常、免疫肠道中野生型 Rmrp 的表达缺乏和神经元发育不良，但 CHH 突变体不表达。以 DDX5 依赖性方式促进 Th17 细胞分化，表明这是一种罕见的 lncRNA 我们的结果表明，Rmrp 与 DDX5 的结合促进了 DDX5 与我们在具体目标 1 中建议确定 RORγt 和 RORγt 靶基因的共激活。表达依赖于 DDX5、Rmrp 和 RORγt，使用全基因组转录组学、染色质使用体外极化或体内产生的野生 Th17 细胞进行可及性和染色质占据研究在具体目标 2 中，我们将使用生化方法确定这些动物的类型和突变体。分子之间相互作用，以及这种相互作用是否仅限于 Th17 细胞或存在于其他细胞中在特定目标 3 中，我们将描述 Th17 细胞中相互作用的影响。介导的炎症、ILC3 依赖性屏障保护以及胸腺细胞和淋巴细胞的发育我们的结果将有助于更好地理解 RORγt 的功能，这可能有助于设计针对自身免疫性疾病和 CHH 患者免疫缺陷的更具体的治疗方法。