Department of Cellular Biology, Institute of Biology, University of Brasilia, Brasilia 70910-900, Brazil.
Instituto Nacional de Ciência e Tecnologia (iii-INCT), São Paulo 05403-000, Brazil.
Int J Mol Sci. 2021 Apr 26;22(9):4541. doi: 10.3390/ijms22094541.
Autoimmunity may have its origins of early repertoire selection in developmental B cells. Such a primary repertoire is probably shaped by selecting B cells that can efficiently perform productive signaling, stimulated by self-antigens in the bone marrow, such as DNA. In support of that idea, we previously found a V segment from V10 family that can form antibodies that bind to DNA independent of CDR3 usage. In this paper we designed four antibody fragments in a novel single-chain pre-BCR (scpre-BCR) format containing germinal V gene segments from families known to bind DNA (V10) or not (V4) connected to a murine surrogate light chain (SLC), lacking the highly charged unique region (UR), by a hydrophilic peptide linker. We also tested the influence of CDR2 on DNA reactivity by shuffling the CDR2 loop. The scpre-BCRs were expressed in bacteria. V10 bearing scpre-BCR could bind DNA, while scpre-BCR carrying the V4 segment did not. The CDR2 loop shuffling hampered V10 reactivity while displaying a gain-of-function in the nonbinding V4 germline. We modeled the binding sites demonstrating the conservation of a positivity charged pocket in the V10 CDR2 as the possible cross-reactive structural element. We presented evidence of DNA reactivity hardwired in a V gene, suggesting a structural mechanism for innate autoreactivity. Therefore, while autoreactivity to DNA can lead to autoimmunity, efficiently signaling for B cell development is likely a trade-off mechanism leading to the selection of potentially autoreactive repertoires.
自身免疫可能起源于发育中的 B 细胞早期库的选择。这样的原始库可能是通过选择能够在骨髓中自身抗原（如 DNA）刺激下有效进行有产信号转导的 B 细胞来塑造的。支持这一观点，我们之前发现了一个来自 V10 家族的 V 段，它可以形成与 CDR3 使用无关的与 DNA 结合的抗体。在本文中，我们设计了四个抗体片段，以新型单链前 B 细胞受体（scpre-BCR）的形式，其中包含已知与 DNA 结合（V10）或不结合（V4）的家族的生发 V 基因片段，连接到缺乏高度带电独特区（UR）的鼠替代轻链（SLC）上，通过亲水肽接头。我们还通过 shuffling CDR2 环测试了 CDR2 对 DNA 反应性的影响。scpre-BCR 在细菌中表达。带有 V10 的 scpre-BCR 可以与 DNA 结合，而带有 V4 片段的 scpre-BCR 则不能。CDR2 环 shuffling 阻碍了 V10 的反应性，同时在非结合的 V4 种系中显示出功能获得。我们对结合位点进行了建模，证明了 V10 CDR2 中带正电荷的口袋的保守性是可能的交叉反应结构元素。我们提出了 DNA 反应性固有地存在于 V 基因中的证据，这表明了先天自身反应性的结构机制。因此，虽然对 DNA 的自身反应性可能导致自身免疫，但有效地进行 B 细胞发育信号转导可能是导致选择潜在自身反应性库的一种权衡机制。
