Specification of Treg cells: learning from FoxP3 deficiencies

Treg 细胞的规范：从 FoxP3 缺陷中学习

• 批准号: 10521755
• 负责人: CHRISTOPHE O. BENOIST
• 金额: $ 55.59万
• 依托单位: HARVARD MEDICAL SCHOOL
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-06-27 至 2027-05-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10521755
• 关键词: 3-Dimensional; ATAC-seq; Affect; Alanine; Appearance; Architecture; Australia; Autoimmune; Autoimmune Diseases; Autoimmunity; Back; Boston; CD4 Positive T Lymphocytes; Cell Nucleus; Cells; Characteristics; Chromatin; Chromatin Structure; Clinical; Clustered Regularly Interspaced Short Palindromic Repeats; Colitis; Collaborations; Complex; Controlled Environment; Cytometry; Data; Defect; Dermatitis; Disease; Disease susceptibility; Engineering; Enhancers; FOXP3 gene; Family; Female; Foundations; Functional disorder; Gene Expression; Gene Expression Profile; Genes; Genetic; Genetic Transcription; Genomics; Grant; Homeostasis; Human; Immune Tolerance; Immunologics; Inflammation; Internal Ribosome Entry Site; Italy; Learning; Lesion; Link; Location; Logistics; Lymphoid Tissue; Maps; Mediator of activation protein; Microbe; Microscopy; Missense Mutation; Modeling; Molecular; Mothers; Mus; Mutant Strains Mice; Mutation; Nuclear; Nuclear Structure; Organ; Organism; Paris, France; Pathology; Patients; Phenotype; Play; Proteins; Recording of previous events; Regulation; Regulatory T-Lymphocyte; Reporter; Resolution; Sampling; Scanning; Severities; Structure; Symptoms; T-Lymphocyte; Testing; United States National Institutes of Health; Variant; Work; base; clinically relevant; cofactor; conditioning; forkhead protein; immunopathology; immunoregulation; in vivo; insight; male; microbial; multiple omics; mutant; novel; pathobiont; programs; promoter; response; single-cell RNA sequencing; symptomatology; transcription factor; transcriptome; transcriptome sequencing; transcriptomics

T regulatory (Treg) cells help maintain immunologic tolerance and control inflammation in many contexts. Their dysfunction leads to multi-organ autoimmunity in FoxP3-deficient mice and human IPEX patients, with variable clinical manifestations. Treg function and homeostasis are dependent on the transcription factor FoxP3, encoded on ChrX, which determines a substantial portion of their specific transcriptome. Our prior work on FoxP3 structure/function explored how FoxP3’s interactions with different cofactors within multimolecular complexes that reside in different nuclear compartments, and condition its transactivating potential on different transcriptional targets. Further, single-cell cytometry and transcriptomics of T cells from FOXP3-deficient patients and mice revealed a narrow cell-intrinsic signature of the deficiency, directly controlled by FoxP3, and a larger disease signature, conferred in a cell-extrinsic manner, that affects both Treg and conventional CD4+ T cells. We hypothesize that these components vary according to the actual FOXP3 lesion, mutations in different interaction facets leading to different pathology, and that environmental and activation triggers promote the unfolding of the full cell-extrinsic IPEX signature. In Aim1, to better understand the components of Treg dysfunction in IPEX patients, we will analyze the impact of missense FOXP3 mutations by single-cell RNAseq in CD4+ T cells from paired IPEX patients and carrier female relatives (typically mothers), in which we distinguish Treg-like cells expressing wild-type or mutant FOXP3 through random ChrX-inactivation and SNP- based cell identification. Effects on chromatin will also be mapped (ATACseq), and related back to symptomatology in these patients. To enable further analysis in a genetically and environmentally controlled environment, where sample access is not limiting, Aim2 uses CRISPR editing to introduce eight selected missense FOXP3 mutations from IPEX patients into B6 mice, where lymphoid and tissue-resident Tregs will be examined and profiled. Super-resolution microscopy and HiChIPseq will assess the effect of the FOXP3 mutations on localization of the mutant proteins in the nucleus, in terms of nuclear structure and enhancer- promoter loops. We will also analyze the timing and determinants of acquisition of the dominant Treg-extrinsic signature. Aim3 will use this FoxP3 mutant mouse panel to ask how mutation-specific transcriptional features relate to the IPEX-relevant clinical characteristics in mice: appearance of spontaneous autoimmunity (scurfy- like disease), susceptibility to induced autoimmunity (colitis, dermatitis). Challenges with defined microbes and microbial molecules will test the hypothesis that clinical variability in IPEX results, at least in part, from environmental and infectious triggers. These interconnected Aims, with complementary explorations on mice and humans, will bring unique information on how variation in a transcription factor modulates its ability to influence gene expression through chromatin and genomic 3D architecture, and how these relate to a monogenic loss of tolerance in human patients.

T调节性细胞（Treg）在许多情况下有助于维持免疫耐受和控制炎症。他们的