CD180(RP105) Regulation of TLR and BCR Responses in B Cells

描述（由申请人提供）：本提案将研究CD180刺激对B细胞免疫调节的潜力，并研究CD180与BCR和TLR信号相互作用的机制。CD180 （RP105）的表达是TLR4和tlr2介导的B细胞增殖和T细胞非依赖性抗体反应（TI-1反应）所必需的。我们建议对CD180受体进行全面研究，包括研究CD180与BCR和TLR信号相互作用的机制，并结合小鼠CD180激活分析，以确定其对表达NP特异性高亲和力BCR （B1-8hi）的抗原特异性B细胞的影响，以及对NP- cgg抗体应答的影响。这些实验将确定CD180激活是否可以作为致敏信号，在抗原结合时导致选择性B细胞缺失。CD180被认为在B细胞中协调或整合TLR4和TLR2信号与BCR信号，以驱动所有同型的抗原特异性抗体反应。我们发现CD180刺激与TLR激动剂一起导致协同B细胞增殖，并增加B细胞炎症因子IL-6、IL-10和TNF1的产生。相比之下，cd180刺激的B细胞增殖受到BCR同时刺激的抑制，cd180激活的B细胞对抗BCR凋亡敏感。该基金的第二个主要组成部分将使用在哺乳动物细胞中表达的CD180/MD-1可溶性形式的分子工程来帮助识别和潜在地阻断天然CD180配体。我们还将表达抗人和抗小鼠CD180 scFv-Ig融合蛋白。为了研究FcR结合对CD180信号的调控，scFvs将使用wt或突变的人类和小鼠Ig尾构建。这些结果将有助于阐明CD180作为TLR2和TLR4的主要调节因子的作用，并将有助于确定CD180作为基于抗体的治疗靶点的潜力。这项工作还将导致重组分子的构建和表达，这将有助于未来研究CD180在感染性疾病和癌症模型中的治疗。

DESCRIPTION (provided by applicant): This proposal will investigate the potential for B cell immunoregulation by CD180 stimulation and study the mechanisms of CD180 interactions with BCR and TLR signals. CD180 (RP105) expression is required for TLR4 and TLR2-mediated B cell proliferation and T cell- independent antibody responses (TI-1 response). We propose a comprehensive study of the CD180 receptor, including studies of the mechanisms of CD180 interaction with BCR and TLR signals, combined with analysis of CD180 activation in mice to determine the effect on antigen-specific B cells expressing transgenic high affinity BCR (B1-8hi) specific for NP, and on the antibody response to NP-CGG. These experiments will determine whether CD180 activation may serve as a sensitization signal that could result in selective B cell deletion upon antigen binding. CD180 is thought to coordinate or integrate the TLR4 and TLR2 signals with BCR signals in B cells to drive an antigen specific antibody response of all isotypes. We have found that CD180 stimulation together with TLR agonists results in synergistic B cell proliferation, and augments B cell production of inflammatory cytokines IL-6, IL-10, and TNF1. In contrast, proliferation of CD180-stimulated B cells is inhibited by simultaneous BCR stimulation, and CD180-activated B cells are sensitized to anti-BCR apoptosis. The second major component of this grant will use molecular engineering of soluble forms of CD180/MD-1 that are expressed in mammalian cells to help identify and potentially block natural CD180 ligands. We will also express anti-human and anti-mouse CD180 scFv-Ig fusion proteins. The scFvs will be constructed with wt or mutated human and mouse Ig tails in order to study the regulation of CD180 signals by FcR binding. The results will help clarify the role of CD180 as a master regulator of TLR2 and TLR4 and will help determine the potential for CD180 as a target for antibody-based therapy. The work will also result in construction and expression of recombinant molecules that will be useful in future studies of CD180 therapy in infectious disease and cancer models. PUBLIC HEALTH RELEVANCE: CD180 is a critical receptor for regulation of TLR4 and TLR2 function in B cells and could be an important target for immunoregulation. This grant will express recombinant molecules for CD180 stimulation and will test the potential for CD180 as a therapeutic target for deletion of autoreactive B cells.