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细胞耐受性被破坏的自身免疫性疾病中，许多ASCs分泌自身抗体，并以主要方式促进器官病理。B细胞向ASCs的分化受一系列关键转录因子控制，其中包括Ets-1，它是这一过程的负调控因子。在缺乏Ets-1的情况下，B细胞向ASCs的分化增强，其中许多ASCs分泌自身抗体，如血清中IgM和IgG自身抗体的高滴度所示。最近，Toll样受体（TLR）信号，特别是通过TLR7和TLR9，被认为与自身反应性B细胞分化为ASCs的激活有关。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，缺乏TLR接头蛋白Myd88的Ets-1敲除小鼠减少了自身抗体的产生。toll样受体信号是Blimp-1表达的有效诱导剂，Blimp-1是驱动ASC分化的关键转录因子。Ets-1与Blimp-1的物理相互作用抑制了Blimp-1结合目标DNA序列的能力。相反，一个密切相关的转录因子Ets-2不能阻断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。在这项应用中，我们提出了多种检测方法来进一步确定Ets-1在调节靶基因表达、Blimp-1活性和ASC分化中的作用。这些研究将有助于比较能够阻断ASC分化的Ets-1蛋白与密切相关的Ets-2蛋白的生物活性，而后者缺乏这种活性。该提案的具体目的是：(1)确定哪种细胞类型需要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)检查TLR7和9配体在激活Ets-1缺陷B细胞中的体内作用，(3)鉴定调节ASC形成的Ets-1依赖基因表达途径，(4)鉴定赋予ASC发展独特能力的Ets-1结构特征。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。

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.