Role of the immunoglobulin DQ52 DH gene segment in fetal immunosuppression

免疫球蛋白 DQ52 DH 基因片段在胎儿免疫抑制中的作用

• 批准号: 10451016
• 负责人: Harry William Schroeder
• 金额: $ 22.28万
• 依托单位: UNIVERSITY OF ALABAMA AT BIRMINGHAM
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-04-01 至 2024-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10451016
• 关键词: Address; Adult; Affinity; Age; Amino Acid Sequence; Amino Acids; Antibodies; Antibody Formation; Antigens; Autoantibodies; Autoantigens; Autoimmune Diseases; Autoimmunity; B-Cell Development; B-Lymphocytes; Birth; CAR T cell therapy; CRISPR/Cas technology; Complementary RNA; Complex; Consensus; Development; Disease; Elements; Embryo; Embryonic and Fetal Development; Enterobacter cloacae; Epitopes; Equilibrium; Exhibits; Fetal Development; Fetal Liver; Flow Cytometry; Generations; Genes; Genetic Recombination; Glycine; Goals; Growth; Growth and Development function; Guide RNA; Heavy-Chain Immunoglobulins; Hen Egg Lysozyme; Host resistance; Human; Immune; Immune response; Immune system; Immunoglobulin Variable Region; Immunoglobulins; Immunosuppression; Inbred BALB C Mice; Infection; Infectious Agent; Knowledge; Leucine; Life; Light; Lymphoid Tissue; Mature B-Lymphocyte; Modeling; Mus; Neonatal; Nucleotides; Oocytes; Organ; Pathogenicity; Pattern; Peptide/MHC Complex; Population; Pregnancy; Premature Infant; Production; Reading Frames; Regulation; Reporting; Research; Resistance to infection; Rest; Risk; Role; Site; T-Cell Receptor; T-Lymphocyte; T-cell receptor repertoire; Terminator Codon; Testing; Tissues; Tyrosine; V(D)J Recombination; autoreactive B cell; autoreactivity; base; cell type; combinatorial; complementarity-determining region 3; design; embryo/fetus; fetal; immunoglobulin receptor; in utero; infection risk; lymphoid organ; mouse model; neoantigens; preference; prenatal; progenitor; receptor function; response; secondary lymphoid organ; sensor; stem cells; theories; vaccination strategy; virtual

Project Summary Our goal is to address the mechanisms that control the humoral immune response during embryonic and fetal development. During ontogeny, the B cells that are needed to populate the various lymphoid tissues and organs sequentially encounter an increasingly complex array of self-antigens as new cell types and non-lymphoid tissues and organs appear. There is a significant risk that embryonic B cells producing potentially pathogenic autoreactive immunoglobulins (Igs) could pass essential developmental checkpoints and thus survive to be activated after these `neo' antigens are ultimately expressed. Unsurprisingly, there is strong evidence that the humoral immune response is selectively suppressed in mammalian embryos and fetuses. Thus, although in theory Ig and T cell receptor (TCR) repertoires appear to be generated in a strictly stochastic fashion through random VDJ rearrangement and N addition; in practice both VDJ rearrangement and N addition are regulated in utero to restrict Ig and TCR diversity. There are 27 functional DH gene segments in humans and 13 in BALB/c mice. We previously showed that VDJ joins from both human and mouse embryonic B cell progenitors were enriched for use of the DQ52 gene segment (D7-27 in human and D4-01 in mouse). After birth, however, both species use DQ52 sparingly. Consequently, regulation of complementarity-determining region 3 of the Ig heavy (H) chain (CDR-H3), which contains the DQ52 gene segment, does not occur by chance and must be a major element of embryonic and early fetal repertoire control. Our prior research has shown that the sequence of the diversity (D) gene segment both delimits the range of CDR diversity and directs, or even dictates, patterns of epitope recognition and antibody production, thereby influencing host resistance to infection and autoimmune disease. DQ52 is the most highly conserved DH between human in mouse. Interestingly, DQ52 differs dramatically in amino acid content from the rest of the Ds, the most striking difference being the absence of tyrosine and enrichment for glycine. In this regard, DQ52 is more similar to TCR Dβ than the other Ig DH, making the in utero DQ52 CDR-H3 repertoire more like TCRβ CDR-B3 than adult Ig CDR-H3. TCRs function more like polyreactive sensors than monospecific effectors. Thus, using DQ52 in CDR-H3 could have the effect of reducing monospecificity and affinity for antigen, and reducing antibody production. These fundamental observations led us to our hypothesis that the role of Ig DQ52 during ontogeny is to promote the production of B cells while simultaneously serving as an immune suppressant DH. To test this hypothesis and to provide proof-of-concept, we propose to (a) use CRISPR/Cas9 technology to create a mouse whose DH locus contains only DQ52 (ΔD-DQ52), (b) test whether use of DQ52 facilitates B cell production, and (c) test whether use of DQ52 results in reduced antibody production. These studies will fill a major gap in our knowledge about the mechanisms that underlie immune suppression of the fetal humoral immune response, which is a major factor in the increased risk of infection in premature infants and in developing in utero vaccination strategies.

项目概要 我们的目标是解决胚胎和胎儿时期控制体液免疫反应的机制 发展。在个体发育过程中，B 细胞需要填充各种淋巴组织和器官 依次遇到一系列日益复杂的自身抗原作为新的细胞类型和非淋巴细胞 组织和器官出现。胚胎 B 细胞产生潜在致病性的风险很大 自身反应性免疫球蛋白（Igs）可以通过重要的发育检查点，从而存活下来 这些“新”抗原最终表达后被激活。不出所料，有强有力的证据表明 体液免疫反应在哺乳动物胚胎和胎儿中被选择性抑制。因此，虽然在 理论上 Ig 和 T 细胞受体 (TCR) 库似乎是通过严格随机的方式生成的 随机VDJ重排和N加法；实际上，VDJ 重排和 N 添加均在 子宫内限制 Ig 和 TCR 多样性。人类有 27 个功能性 DH 基因片段，BALB/c 中有 13 个 老鼠。我们之前表明，来自人和小鼠胚胎 B 细胞祖细胞的 VDJ 连接是 富集以使用 DQ52 基因片段（人类中的 D7-27 和小鼠中的 D4-01）。然而出生后，两人 物种很少使用 DQ52。因此，Ig 重链互补决定区 3 的调节 (H)链(CDR-H3)，含有DQ52基因片段，不是偶然出现的，一定是一个主要的 胚胎和早期胎儿曲目控制的要素。我们之前的研究表明， 多样性 (D) 基因片段既界定了 CDR 多样性的范围，又指导甚至决定了 CDR 多样性的模式。 表位识别和抗体产生，从而影响宿主对感染和自身免疫的抵抗力 疾病。 DQ52 是人类与小鼠之间最高度保守的 DH。有趣的是，DQ52 有所不同 与其他 D 相比，氨基酸含量有显着差异，最显着的差异是缺乏 酪氨酸和甘氨酸的富集。在这方面，DQ52 与 TCR Dβ 比其他 Ig DH 更相似，使得 与成人 Ig CDR-H3 相比，子宫内 DQ52 CDR-H3 库更像 TCRβ CDR-B3。 TCR 的功能更像 多反应传感器优于单特异性效应器。因此，在 CDR-H3 中使用 DQ52 可能会产生以下效果： 降低对抗原的单特异性和亲和力，并减少抗体的产生。这些基本的 观察结果使我们得出这样的假设：Ig DQ52 在个体发育过程中的作用是促进生产 B 细胞，同时充当免疫抑制剂 DH。为了检验这个假设并 提供概念验证，我们建议 (a) 使用 CRISPR/Cas9 技术创建 DH 基因座的小鼠 仅包含 DQ52 (ΔD-DQ52)，(b) 测试使用 DQ52 是否促进 B 细胞产生，以及 (c) 测试是否 使用 DQ52 会导致抗体产生减少。这些研究将填补我们在以下方面的知识的重大空白： 胎儿体液免疫反应的免疫抑制机制，这是一个主要的 早产儿感染风险增加和制定子宫内疫苗接种策略的因素。