Identification of Specific Roles for Ets-1 in B Cell Tolerance

描述(由申请人提供)：抗体分泌细胞(ASCs)产生的抗体对许多不同病原体的免疫反应至关重要。然而，在B细胞耐受性被打破的自身免疫性疾病中，许多ASCs分泌自身抗体，并在器官病理中起主要作用。B细胞向ASCs的分化受包括Ets-1在内的一系列关键转录因子的控制，Ets-1是这一过程的负调控因子。在缺乏ETS-1的情况下，B细胞经过增强分化为ASCs，其中许多ASCs分泌自身抗体，如血清中高滴度的IgM和Ig G自身抗体所证明的那样。最近，Toll样受体(TLR)信号，特别是通过TLR7和TLR9，参与了自身反应性B细胞分化为ASCs的激活。有趣的是，ETS-1缺陷的B细胞在体外对TLR7和TLR9配体的反应增强，而缺乏TLR适配器蛋白MyD88的ETS-1基因敲除小鼠减少了自身抗体的产生。Toll样受体信号是Blimp-1表达的有效诱导者，Blimp-1是推动ASC分化的关键转录因子。Ets-1与Blimp-1在物理上相互作用，以抑制Blimp-1与靶DNA序列的结合能力。相反，密切相关的转录因子ETS-2无法阻止Blimp-1结合或抑制ASC分化。Ets-1也被认为调节控制ASC形成的关键靶基因的表达，但大多数这些靶基因的身份尚不清楚。在这一应用中，我们提出了各种检测方法，以进一步确定Ets-1在调节靶基因表达、Blimp-1活性和ASC分化中的作用。通过比较Ets-1和Ets-2的生物学活性将有助于这些研究，Ets-1可以阻断ASC的分化，而Ets-2与Ets-2密切相关，而Ets-2缺乏这种活性。该提案的具体目的是(1)确定哪些细胞类型需要ETS-1活性来限制ASC的形成、自身抗体的分泌和自身免疫性疾病，(2)研究TLR7和9个配体在体内激活ETS-1缺陷的B细胞的作用，(3)确定调控ASC形成的ETS-1依赖的基因表达途径，以及(4)确定赋予ASC独特调控能力的ETS-1的结构特征。总之，我们的研究将为B细胞分化的转录调控提供新的见解，特别是在自身抗体分泌的情况下。 公共卫生相关性：作为对病原体的免疫反应的一部分，B细胞分化为抗体分泌细胞(ASCs)，产生高水平的抗体来对抗感染。在自身免疫性疾病中，有大量的ASCs分泌抗体，与自身组织发生反应。从拟议的研究中收集的信息将有助于确定B细胞如何分化为ASCs的分子细节，以及这一过程在自身免疫性疾病中如何存在缺陷，从而潜在地确定在临床上刺激或抑制这一过程的新策略。

DESCRIPTION (provided by applicant): The production of antibodies by antibody-secreting cells (ASCs) is critically important for immune responses to many different pathogens. However, in autoimmune diseases where B cell tolerance is broken, many ASCs secrete autoantibodies and contribute in a major way to organ pathology. The differentiation of B cells into ASCs is controlled by a cohort of key transcription factors including Ets-1, which serves as a negative regulator of this process. In the absence of Ets-1, B cells undergo enhanced differentiation into ASCs many of which secrete autoantibodies as demonstrated by high titers of IgM and IgG autoantibodies in the serum. Recently Toll- like receptor (TLR) signaling, particularly via TLR7 and TLR9, has been implicated in the activation of autoreactive B cells to differentiate into ASCs. Interestingly, Ets-1 deficient B cells exhibit enhanced responses to TLR7 and TLR9 ligands in vitro and Ets-1 knockout mice lacking the TLR adaptor protein Myd88 have reduced autoantibody production. Toll-like receptor signaling is a potent inducer of the expression of Blimp-1, a key transcription factor that drives ASC differentiation. Ets-1 physically interacts with Blimp-1 to inhibit the ability of Blimp-1 to bind target DNA sequences. In contrast, a closely related transcription factor Ets-2 is unable to block Blimp-1 binding or to inhibit ASC differentiation. Ets-1 also is thought to regulate the expression of critical target genes controlling ASC formation, but the identity of most of these targets remains unclear. In this application we propose a variety of assays to further define the role of Ets-1 in regulating target gene expression, Blimp-1 activity and ASC differentiation. These studies will be aided by comparing biological activities of Ets-1, which can block ASC differentiation, with the closely related Ets-2 protein, which lacks this activity. The specific aims of the proposal are (1) to determine which cell types require Ets-1 activity to limit ASC formation, autoantibody secretion and autoimmune disease, (2) to examine the in vivo role of TLR7 and 9 ligands in activating Ets-1 deficient B cells, (3) to identify Ets-1 dependent gene expression pathways regulating ASC formation and (4) to identify structural features of Ets-1 that impart a unique ability to regulate ASC development. Together our studies will provide novel insights into the transcriptional regulation of B cell differentiation particularly in situations of autoantibody secretion. PUBLIC HEALTH RELEVANCE: As part of the immune response to pathogens, B cells differentiate into antibody-secreting cells (ASCs) that produce high levels of antibodies to fight infection. In autoimmune disease, there are large numbers of ASCs that secrete antibodies that react with self-tissues. Information gleaned from the studies proposed will help define the molecular details of how B cells differentiate into ASCs and how that process is defective in autoimmune disease, thereby potentially identifying new strategies to stimulate or inhibit this process clinically.