Treg development and function controlled by cis-regulatory circuits

项目摘要 调节性T细胞(Treg)在维持免疫系统动态平衡和预防 自身免疫学和免疫病理学。Treg功能受损与多种自身免疫性疾病有关 包括1型糖尿病和多发性硬化症。另一方面，人们认为肿瘤内Treg细胞的丰富 成为有效的抗肿瘤免疫反应的屏障。Treg细胞系的发育和维持 依赖于转录因子Foxp3，因为功能丧失突变会导致严重的淋巴增殖性 小鼠和人类的疾病。因此，了解支配Foxp3诱导和稳定性的机制 可能导致自身免疫性疾病和癌症的新疗法的开发。此前，郑博士和 同事们在Foxp3基因座发现了三个进化上保守的顺式调控元件，它们在 在Treg分化或维护中的独特作用。为了扩大这一研究领域，郑博士的团队将使用 一种基于CRISPR/Cas9的方法，称为CRREST-SEQ(通过平铺-删除和删除顺式调节元件扫描 测序)对Foxp3基因座附近的远端顺式元件进行无偏筛选和功能评估。 一项初步研究表明，可能有多达50个顺式调控元件参与Foxp3的控制 表情。对这些新的顺式元件的深入表征将促进对 Foxp3表达的调控电路。大量研究产生了对数百万候选人的注释 Tregs中数千个基因的调控元件，但对于这些元件是如何 控制他们的目标基因，这些基因是千碱基甚至百万碱基的。全基因组技术的初步研究 Treg中的染色体环产生了一组可能调节Treg的远端DNA环 标志性基因。此外，这些数据表明，Foxp3在促进远端DNA环中扮演了一个新的角色。至 更好地了解顺式调节电路如何控制Treg的分化和功能，郑博士的团队将 通过基于平铺缺失的遗传筛选来识别和表征控制Foxp3表达的顺式元件 目的1)并探讨远端增强子如何通过染色体调节调节性T细胞的基因表达 循环(目标2)。此外，他们将调查Foxp3是否促进了染色体环路，以建立和 保持Treg血统。由于Treg细胞与自身免疫性疾病和癌症直接相关，郑博士的 研究将为Treg的发展和功能的分子机制提供洞察力，并提供新的 出于治疗目的操纵Tregs的机会。

Project Summary Regulatory T cell (Treg) plays a critical role in maintaining immune system homeostasis and preventing autoimmunity and immunopathology. Compromised Treg function is linked to multiple autoimmune diseases including type 1 diabetes and multiple sclerosis. On the flip side, enrichment of Treg cells within tumors is thought to be a barrier to effective anti-tumor immune response. The development and maintenance of Treg cell lineage are dependent on transcription factor Foxp3, as loss of function mutations lead to severe lymphoproliferative disease in mice and humans. Thus, understanding the mechanisms that govern Foxp3 induction and stability may lead to the development of novel therapies for autoimmune disease and cancer. Previously, Dr. Zheng and colleagues identified three evolutionarily conserved cis-regulatory elements at the Foxp3 locus, which play unique roles in Treg differentiation or maintenance. To expand this line of research, Dr. Zheng’s group will use a CRISPR/Cas9 based approach named CREST-seq (cis-regulatory element scan by tiling-deletion and sequencing) to perform unbiased screen and functional assessment for distal cis-elements near the Foxp3 locus. A preliminary study showed there are potentially up to 50 cis-regulatory elements involved in the control of Foxp3 expression. In-depth characterization of these new cis-elements will advance the understandings on the regulatory circuitry of Foxp3 expression. Numerous studies generated annotations on millions of candidate regulatory elements for thousands of genes in Tregs, but understanding is incomplete on how these elements control their target genes that are kilo-bases or even mega-bases away. Preliminary studies on genome-wide chromosome looping in Tregs generated a collection of distal DNA loops that can potentially regulate Treg signature genes. Furthermore, these data suggest a novel role for Foxp3 in facilitating distal DNA looping. To better understand how cis-regulatory circuitry control Treg differentiation and function, Dr. Zheng’s group will identify and characterize the cis-elements that control Foxp3 expression by a tiling deletion based genetic screen (Aim 1) and explore how distal enhancers regulate gene expression in regulatory T cell through chromosome looping (Aim 2). Furthermore, they will investigate if Foxp3 facilitates chromosome looping to establish and maintain the Treg lineage. Since Treg cell is directly related to autoimmune diseases and cancer, Dr. Zheng’s study will provide insight into the molecular mechanisms of Treg development and function and offer new opportunities to manipulate Tregs for therapeutic purposes.