GABP-controlled gene regulatory network in T lymphocytes

描述(由申请人提供)：针对外来抗原的保护性细胞免疫反应依赖于功能性T淋巴细胞。T细胞的发育、激活和分化受不同组合的转录因子调控。GA结合蛋白(GABP)是一种普遍表达的Ets家族转录因子，其功能由两个不同的亚基GABP1和GABP2组成，分别介导DNA结合和反式激活。在此之前，我们已经证明GABP对T细胞中IL-7R1链的表达具有重要的调节作用，提示GABP在免疫系统中起着关键作用。虽然GABP1基因的完全失活会导致植入前的死亡，但我们已经产生了一种GABP1突变小鼠品系，它是亚型的，在胚胎发生期间存活时间更长。在我们利用GABP1突变胚胎中的胎肝细胞重建RAG2基因缺陷小鼠的淋巴室的初步研究中，我们发现GABP在胸腺细胞发育和T细胞介导的免疫反应中具有重要作用。 本研究旨在构建一个在T细胞内GABP调控下的层次化、互动式基因调控网络。我们的长期目标是阐明GABP如何通过调节其靶基因和与其他关键因素相互作用来规划T淋巴细胞生成和T细胞免疫反应。这一信息将为在生理和病理条件下调节造血干细胞分化和免疫反应的结果提供科学依据。 明确目标。鉴定T淋巴细胞中直接的GABP靶基因，建立由GABP调控的T细胞生物学基因调控网络。利用我们的基因芯片微阵列分析数据，我们将扩展我们的研究，使用比较基因组学和染色质免疫沉淀分析(ChIP)与小鼠启动子阵列(芯片上的芯片)相结合的方法来识别GABP直接靶基因。然后，我们将绘制GABP结合元件的精确位置图，并研究它们在激活/抑制各自基因方面的功能重要性。有了这些知识，可以构建T细胞的基因调控网络，这将为更广泛地研究胸腺细胞发育和T细胞功能的交互转录调控提供平台。遗传性和获得性免疫缺陷患者的免疫系统重建仍然是一个重大挑战，更好地了解造血干细胞分化为淋巴系的分子机制是改进免疫系统重建策略的基础。该项目将深入了解GABP作为一种转录因子如何有助于将造血干细胞指定为T细胞，以及GABP如何调节细胞免疫反应。这一认识将为在生理和病理条件下调控造血干细胞分化和免疫反应的结果提供科学依据。

DESCRIPTION (provided by applicant): Protective cellular immune responses against foreign antigens rely on functional T lymphocytes. The development, activation and differentiation of T cells are controlled by transcription factors in different combinations. GA binding protein (GABP) is a ubiquitously expressed Ets family transcription factor, and functional GABP consists of two different subunits, GABP1 and GABP2, which mediate DNA binding and transactivation, respectively. Previously we have demonstrated that GABP critically regulates the expression of interleukin-7 receptor 1 (IL-7R1) chain in T cells, suggesting a key role of GABP in the immune system. Whereas complete inactivation of the GABP1 gene caused pre- implantation lethality, we have generated a GABP1 mutant mouse strain that is hypomorphic and has prolonged survival during embryogenesis. In our preliminary studies using fetal liver cells from the GABP1 mutant embryos to reconstitute lymphoid compartment in Rag2-deficient mice, we found that GABP has essential roles in thymocyte development and T cell-mediated immune responses. This research project aims to construct a hierarchical and interactive gene regulatory network under the control of GABP in T cells. Our long-term goal is to elucidate how GABP programs T lymphopoiesis and T cell immune responses through regulating its target genes and interacting with other key factors. This information will provide a scientific basis for modulating outcomes of hematopoietic stem cell differentiation and immune responses under physiological and pathological conditions. Specific Aim. To identify direct GABP target genes in T lymphocytes and to establish a GABP- controlled gene regulatory network programming T cell biology. Taking advantage of our GeneChip microarray analysis data, we will extend our studies to identify GABP direct target genes using approaches of comparative genomics and chromatin immunoprecipitation assay (ChIP) coupled with a mouse promoter array (ChIP-on-chip). We will then map the precise locations of GABP-binding elements and investigate their functional importance in activating/repressing respective genes. With this knowledge, a gene regulatory network in T cells can be constructed, and this will provide a platform for a broader investigation of the interactive transcriptional control of thymocyte development and T cell functions. Reconstitution of the immune system in individuals with inherited and acquired immunodeficiency remains a major challenge, and a better understanding of the molecular mechanism by which hematopoietic stem cells differentiate to lymphoid lineages is fundamental to improve strategies for immune system reconstitution. This project will generate insights into how GABP, a transcription factor, contributes to specifying hematopoietic stem cells to the T-lineage cells, and how GABP modulates the cellular immune response. This knowledge will provide a scientific basis for manipulating outcomes of hematopoietic stem cell differentiation and immune response under physiological and pathological conditions.