Analysis and application of the capability of immune system to improve antigen-specificity of antibodies.

免疫系统提高抗体抗原特异性能力的分析与应用。

• 批准号: 12450334
• 负责人: OHMORI Hitoshi
• 金额: $ 8.13万
• 依托单位: Okayama University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (B)
• 财政年份: 2000
• 资助国家: 日本
• 起止时间: 2000 至 2002
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-12450334/
• 关键词: Antibody; antibody genes; affinity maturation; somatic mutation; transgenic mouse; B lymphocytes; germinal center; gene rearrangement; 遺伝子再編成; V(D)J再構成; RAG遺伝子; IL-7; V(D)J再構

Recently, peripheral B cells have been shown to undergo secondary V(D)J rearrangement of Ig genes, but the physiological role of this event has not been fully elucidated. Here, we investigated whether rearrangement of L chain genes in the periphery is involved in the generation of high-affinity Abs. To test this possibility, we used a 17.2.25 rearranged VHDJH gene (VHT)-knockin mouse whose B cell diversity is limited due to the expression of the site-directed transgene. Immunization of the mouse with p-nitrophenylacetyl (pNP)-conjugated chicken γ-globulin (CGG) preferentially led to the production of anti-pNP IgG Abs comprised of non-VHT-encoded H chains and λ chains, which constituted a majority of high-affinity IgG to this hapten. We isolated three independent anti-pNP IgG1 mAbs (two bearing λ1 with high-affinity and one bearing κ with low-affinity), all of which used a common VHDJH containing an identical point mutation, thus suggesting that κ to λ1 replacement contributed to an increase of the Ab affinity. RAG-2 mRNA and the recombination signal sequence break reflecting the λ1 gene rearrangement were detected in the draining lymph node (LN) of immunized mice, but not of non-immunized animals. There was a close correlation between the levels of the λ gene rearrangement and λ^+ high-affinity anti-pNP IgG. Thus, our findings suggest that new rearrangement of λ genes in the periphery contributes to affinity maturation of anti-pNP IgG.

最近，外周B细胞已显示出Ig基因的次级V（D）J重排，但是该事件的物理作用尚未完全阐明。在这里，我们研究了周围L链基因的重排是否参与高亲和力ABS的产生。为了测试这种可能性，我们使用了17.2.25重新排列的VHDJH基因（VHT）-Knockin小鼠，其B细胞多样性由于位置定向转换的表达而受到限制。用P-硝基苯基乙酰基（PNP）偶联的鸡γ-球蛋白（CGG）对小鼠进行免疫接种，优先导致了非VHT指定的H链和λ链的产生抗PNP IgG ABS的产生，该链条构成了高含量IgG的主要性IgG。我们分离了三个独立的抗PNP IgG1 mAB（两个具有高亲和力的脊椎λ1，一个轴承κ具有低亲和力），所有RAG-2 mRNA和重新组合信号序列断裂反映了λ1基因重排的分解，在被抑制的无剂量的淋巴结（LN）中检测到了λ1基因重排（LN），但没有被发现。 λ基因重排的水平与λ^+高亲和力抗PNP IgG之间存在密切相关。这就是我们的发现表明，周围的λ基因的新重排有助于抗PNP IgG的亲和力成熟。

项目成果

大森斉: "末梢リンパ組織における抗体遺伝子の再構成-抗体遺伝子の構造はどこまで柔軟か?"細胞工学. 19巻3号. 482-487 (2000)

Hitoshi Omori：“外周淋巴组织中抗体基因的重组 - 抗体基因的结构有多灵活？”，第 19 卷，第 3 期，482-487 (2000)。

大森斉: "胚中心における抗体遺伝子再構成とその生理的意義"感染・炎症・免疫. 30巻1号. 64-66 (2000)

Hitoshi Omori：“生发中心的抗体基因重排及其生理意义”《感染、炎症和免疫学》，第 30 卷，第 1. 64-66 期（2000 年）。