The role of Immunoglobulin CDRH3 in Autoimmune Disease

描述（由申请人提供）：系统性红斑狼疮 (SLE) 是一种常见的多系统自身免疫性疾病，估计影响超过 500,000 名美国人。 我们的目标是更好地了解通常阻止导致疾病的自身反应性抗体表达的机制，并更好地了解这些机制在何处以及如何失效。 由于伦理方面的考虑，对人类疾病的根本原因进行研究通常是不可能的。 数十年的医学研究证明了小鼠模型作为确定根本原因以及治疗和预防新途径的手段的有用性。 C57BL/6 中 sle1、sle2 和/或 sle3/5 NZM2410 狼疮基因座的存在促进了自身抗体驱动性疾病的发展，该疾病具有人类 SLE 的许多特征。 我们的假设是，在易感个体中，未能正确调节抗体 CDR-H3 的一个特定部分会促进抗 DNA 抗体的产生并引发疾病。 CDR-H3 很重要，因为它位于抗原结合位点的正中心。 CDR-H3 是由 VDJ 加入重新创建的。 我们提出DH的序列对CDR-H3含量具有显着影响，并且通常限制CDR-H3的主要机制是通过阅读框保存DH序列。 我们认为，自身免疫是由于未能调节不利阅读框中氨基酸的使用而导致的。这些氨基酸也可以通过N添加或体细胞突变引入。 我们假设 sle1、sle2 和 sle3/5 基因通过增加携带不利 CDR-H3 的 B 细胞的存活率，对 B 细胞和库发育的调节产生不利影响。 为了测试这些假设，我们将在 C57BL/6 基因组中培育三个不同的 DH 等位基因，每个等位基因仅限于单个 DH 基因片段。 第一个 DH 等位基因将用倒置阅读框 1 中的精氨酸和其他带正电荷的氨基酸替换通常由优选阅读框 1 通过删除编码的酪氨酸和甘氨酸。 第二个将包含单个正常 DH，作为其余 DH 基因座丢失的对照。 第三个方法将通过替换疏水阅读框 2 中的密码子来控制酪氨酸和甘氨酸含量的损失。如果我们的假设正确，DH 阅读框 1 中精氨酸的富集将增强不利的 CDR-H3 序列的表达。 这种升高的表达将超过 CDR-H3 含量正常控制的阈值，加速自身反应性抗体（包括抗 DNA）的表达。 在 sle1、sle2 和 sle3/5 存在的情况下，含有改变的 DH 等位基因的小鼠将表现出 B 细胞和库发育模式的改变，证据表明对抗原的免疫反应改变，证明 IgG 抗 dsDNA 抗体的表达增强，并导致自身免疫性疾病的加速表达。

DESCRIPTION (provided by applicant): Systemic lupus erythematosus (SLE) is a common multisystem autoimmune disease that is estimated to affect more than 500,000 Americans. Our goal is to gain a better understanding of the mechanisms that normally prevent expression of the self-reactive antibodies that cause the disease as well as to better understand where and how these mechanisms fail. Research into fundamental causes of disease in humans is often impossible due to ethical considerations. Decades of medical research attest to the usefulness of mouse models as a means to identify root causes as well as new avenues for treatment and prevention. The presence of the sle1, sle2, and/or sle3/5 NZM2410 lupus loci in C57BL/6 promotes development of an autoantibody driven disorder that shares many of the features of human SLE. It is our hypothesis that failure to properly regulate one specific part of the antibody, CDR-H3, in susceptible individuals facilitates production of anti-DNA antibodies and triggers disease. CDR-H3 is important because it lies at the very center of the antigen binding site. CDR-H3 is created de novo by VDJ joining. We propose that the sequence of the DH has a dominant effect on CDR-H3 content and that a primary mechanism normally constraining CDR-H3 is conservation of DH sequence by reading frame. We propose that autoimmunity results from failure to regulate use of the amino acids that can be found in disfavored reading frames. These amino acids can also be introduced by N addition or somatic mutation. We hypothesize that sle1, sle2, and sle3/5 genes adversely affect regulation of B cell and repertoire development by permitting increased survival of B cells bearing disfavored CDR-H3. To test these hypotheses we will breed three different DH alleles, each limited to a single DH gene segment, into the C57BL/6 genome. The first DH allele will substitute arginine and other positively charged amino acids from inverted reading frame 1 for the tyrosine and glycine that are normally encoded by the preferred reading frame 1 by deletion. The second will contain a single, normal DH as a control for the loss of the rest of the DH locus. The third will control for the loss of tyrosine and glycine content by substituting codons from hydrophobic reading frame 2. If our hypotheses are correct, enrichment for arginine in DH reading frame 1 will enhance expression of disfavored CDR-H3 sequence. This heightened expression will exceed the threshold for normal control of CDR-H3 content, accelerating expression of self-reactive antibodies, including anti-DNA. In the presence of sle1, sle2, and sle3/5, mice containing altered DH alleles will display altered patterns of B cell and repertoire development, evidence altered immune responses to antigen, demonstrate enhanced expression of IgG anti-dsDNA antibodies, and lead to accelerated expression of autoimmune disease.