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.

项目摘要 我们的目标是解决机制，控制体液免疫反应，在胚胎和胎儿 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 作为新的细胞类型和非淋巴细胞， 组织和器官出现。胚胎B细胞产生潜在的致病性 自身反应性免疫球蛋白（Ig）可以通过必要的发育检查点，从而存活下来， 在这些“neo”抗原最终表达后激活。毫不奇怪，有强有力的证据表明， 体液免疫应答在哺乳动物胚胎和胎儿中被选择性抑制。因此，虽然在 理论IG和T细胞受体（TCR）库似乎是以严格随机的方式产生的， 随机VDJ重排和N加成;在实践中，VDJ重排和N加成都受到调节， 限制IG和TCR多样性。人DH基因有27个功能片段，BALB/c有13个 小鼠我们以前的研究表明，人和小鼠胚胎B细胞祖细胞的VDJ连接是 富集用于使用DQ 52基因区段（人中的D 7 -27和小鼠中的D4-01）。出生后，两人 物种谨慎地使用DQ 52。因此，调节IG重链的互补决定区3， (H)含有DQ 52基因片段的CDR-H3链不是偶然出现的，必须是一个主要的 胚胎和早期胎儿剧目控制的元素。我们先前的研究表明， 多样性（D）基因区段既界定了CDR多样性的范围，又指导或甚至决定了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抗性， 疾病DQ 52是人与小鼠之间最高度保守的DH。有趣的是，DQ 52不同 在氨基酸含量方面与其余的D显著不同，最显著的差异是缺乏 酪氨酸和富集甘氨酸。在这方面，DQ 52比其他IG DH更类似于TCR Dβ，使得 子宫内DQ 52 CDR-H3库比成人IG CDR-H3更像TCRβ CDR-B3。TCR的功能更像是 多反应传感器比单特异性效应器。因此，在CDR-H3中使用DQ 52可以具有以下效果： 降低对抗原的单特异性和亲和力，并降低抗体产生。这些基本 观察结果使我们假设IG DQ 52在个体发育过程中的作用是促进 B细胞，同时作为免疫抑制剂DH。为了验证这一假设， 为了提供概念验证，我们提出（a）使用CRISPR/Cas9技术来创建其DH基因座 仅含有DQ 52（Δ D-DQ 52），（B）测试使用DQ 52是否促进B细胞生产，和（c）测试是否 使用DQ 52导致抗体产生减少。这些研究将填补我们对以下方面知识的重大空白 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。