Mechanism of Bach1-Mediated Transcriptional Regulation and Immune Function

描述(由申请者提供)：为了保护自己免受外来入侵者的侵袭和耐受，需要对免疫系统进行精确的调节。免疫调节功能障碍可导致对感染的免疫反应受损以及自身免疫性疾病的发展。转录因子Bach1在调节免疫过程中的作用已被间接涉及，但尚未被直接研究；因此，该项目的广泛和长期目标是在细胞和分子水平上确定Bach1的功能及其介导保护性免疫和自身免疫性疾病的机制。为了实现这一目标，将采用以下研究设计和方法。1)首先，阐明Bach1调控转录的机制。具体地说，染色质免疫沉淀和微阵列(芯片)将被用来确定Bach1是否与启动子近端和/或远端调控元件结合，直接引发转录激活和抑制。计算基序分析和荧光素酶报告分析将用于确定Bach1对转录调控重要的功能结构域。2)其次，将确定Bach1在病毒和细菌感染中的作用。在感染期间，Bach1将在巨噬细胞和小鼠体内被消融，以确定Bach1调节保护性免疫反应的分子途径。3)第三，研究Bach1在自身免疫中的作用。初步数据表明，Bach1促进了模拟人类多发性硬化症的小鼠实验性自身免疫性脑脊髓炎(EAE)的发展。因此，骨髓重建和过继移植实验将被用来鉴定那些具有Bach1内在功能调节EAE的免疫细胞(S)。微阵列实验将在这些细胞中进行，以确定潜在的靶点和受Bach1调控的下游信号通路。4)第四，将确定并确定负责调解其在EAE发展中的作用的Bach1目标。Bach1介导的EAE对消融这些靶点的影响将通过产生Bach1和感兴趣的靶点(S)的双重敲除动物来检验。此外，还将确定Bach1在体内发挥调节功能所需的功能结构域。总体而言，该项目将确定一种新的免疫系统调节因子(Bach1)，并揭示这些信号级联，通过这些信号级联调节感染期间的保护性免疫和功能障碍时的自身免疫。因此，该项目的成功完成将为免疫相关疾病和自身免疫性疾病的治疗确定新的目标，从而使公众健康受益。 公共卫生相关性：该项目将阐明Bach1在调节免疫系统中的作用，包括Bach1如何在微生物感染期间提供保护，并确定Bach1如何介导多发性硬化症等自身免疫性疾病。该项目还将确定由Bach1调控的基因，这些基因对执行这些免疫过程非常重要。该项目通过确定Bach1和其他潜在的治疗免疫相关和自身免疫性疾病的新目标，与公共卫生相关。

DESCRIPTION (provided by applicant): Precise regulation of the immune system is required for protection from foreign invaders and tolerance of self. Dysfunctional immune regulation can lead to impaired immune responses to infections as well as development of autoimmune diseases. The role of the transcription factor Bach1 in regulating immune processes has been indirectly implicated but has not been directly explored; thus, the broad and long-term objective of this project is to determine the function of Bach1 and the mechanism by which it mediates protective immunity and autoimmune diseases at a cellular and molecular level. The following research design and methods will be employed to achieve this goal. 1) First, the mechanism by which Bach1 regulates transcription will be elucidated. Specifically, chromatin immunoprecipitation followed by microarray (ChIP-chip) will be used to determine whether Bach1 binds promoter proximal and/or distal regulatory elements to directly elicit transcriptional activation and repression. Computational motif analyses and luciferase reporter assays will be used to define those functional domains of Bach1 important for transcriptional regulation. 2) Second, the role of Bach1 during viral and bacterial infections will be determined. Bach1 will be ablated in macrophages and in mice during infection in order to characterize the molecular pathways by which Bach1 regulates protective immune responses. 3) Third, the role of Bach1 in autoimmunity will be investigated. Preliminary data demonstrated that Bach1 promotes the development of murine experimental autoimmune encephalomyelitis (EAE), which mimics human multiple sclerosis. Accordingly, bone marrow reconstitution and adoptive transfer experiments will be used to identify those immune cell(s) in which the intrinsic function of Bach1 modulates EAE. Microarray experiments will be performed in those cells to identify potential targets and downstream signaling pathways regulated by Bach1. 4) Fourth, Bach1 targets responsible for mediating its role in the development of EAE will be identified and characterized. The effect of Bach1-mediated EAE upon ablating these targets will be examined by generating double knockout animals of Bach1 and the target(s) of interest. Furthermore, functional domains of Bach1 necessary for its regulatory function in vivo will be identified. Overall, this project will identify a new regulator of the immune system (Bach1) and reveal those signaling cascades by which it modulates protective immunity during infection and autoimmunity during dysfunction. The successful completion of this project will therefore benefit public health by identifying new targets for therapy against immune-related and autoimmune diseases. PUBLIC HEALTH RELEVANCE: This project will elucidate the role of Bach1 in regulating the immune system, including how Bach1 may offer protection during microbial infections and establish how Bach1 mediates autoimmune diseases, such as multiple sclerosis. This project will also identify genes regulated by Bach1 important for carrying out these immune processes. This project is relevant to public health by identifying Bach1 and other potential new targets for therapy against immune-related and autoimmune diseases.