A weird way to recognize phosphoantigens In contrast to the well-studied αβ T cells, which recognize peptide antigens presented by major histocompatibility complex (MHC) and MHC-like molecules, how γδ T cells recognize antigens remains largely a mystery. One major class of γδ T cells, designated Vγ9Vδ2+, is activated by small, phosphorylated nonpeptide antigens, or phosphoantigens, produced by microbes and cancer cells. Rigau et al. found that these cells needed the combination of two immunoglobulin superfamily members, butyrophilin 2A1 (BTN2A1) and BTN3A1, on their cell surface to recognize these phosphoantigens. BTN2A1 directly binds the Vγ9+ domain of the T cell receptor (TCR), whereas a second ligand, potentially BTN3A1, binds the Vδ2 and γ-chain regions on the opposite side of the TCR. A better understanding of this unexpected form of T cell antigen recognition should inform and enhance future γδ T cell–mediated immunotherapies. Science, this issue p. eaay5516 A key ligand is involved in the recognition of pathogen- or cancer-associated phosphoantigens by γδ immunological T cells. INTRODUCTION T cells represent a key component of the immune system that can recognize foreign molecules (antigens) via cognate cell surface receptors termed T cell receptors (TCRs). The two main families of T cells, known as αβ and γδ T cells, are defined by the different gene loci that they use to generate their respective TCRs. αβ T cells typically recognize antigens displayed on the surface of target cells in association with antigen-presenting molecules known as major histocompatibility complex (MHC) molecules. Much less is known about how γδ T cells recognize antigen, although it is clear they are also essential to protective immunity. In humans, many γδ T cells (classified as Vγ9Vδ2 T cells) respond to small phosphorylated nonpeptide antigens, called phosphoantigens (pAgs), which are produced by cellular pathogens and cancers. In turn, γδ T cells become activated, proliferate, rapidly produce proinflammatory cytokines such as IFN-γ, and exert cytotoxic activity. pAg recognition appears to involve a cell surface molecule, butyrophilin 3A1 (BTN3A1), which plays a necessary, but not sufficient, role in this process. Therefore, the molecular basis that underpins pAg recognition by Vγ9Vδ2 T cells remains unclear and represents a long-standing conundrum, which has impeded the study of these important immune cells. RATIONALE In contrast to αβ T cells, Vγ9Vδ2 T cells are not MHC-restricted and can recognize pAg expressed by multiple cancers and infectious diseases. Thus, they represent an attractive target for the development of new immunotherapy treatments. A much clearer understanding of the molecular basis for pAg recognition is required to optimally harness these cells for immunotherapy. We undertook a multipronged approach to investigate which molecules are necessary for pAg detection by γδ T cells. We used a genome-wide screen to identify molecules that mediate pAg-driven γδ T cell activation. Furthermore, we asked if these molecules directly bind to the Vγ9Vδ2 TCR and how they work in conjunction with BTN3A1. RESULTS The top candidate molecule identified in our genome-wide screen was butyrophilin 2A1 (BTN2A1), a molecule distinct from, but related to, BTN3A1. We show that without BTN2A1, Vγ9Vδ2 T cells cannot be activated by either bacterial or mammalian pAgs, and that BTN2A1 expression was required for Vγ9Vδ2 T cell–mediated tumor cell killing. Neither BTN3A1 nor the other butyrophilin family members tested can compensate for loss of BTN2A1. BTN2A1 can bind directly to the Vγ9Vδ2 TCR and associates closely with BTN3A1 on the surface of target cells. We also identify an important role for the transmembrane and/or intracellular domain of BTN2A1. Furthermore, pAg-mediated activation of γδ T cells requires coexpression of both BTN2A1 and BTN3A1, which together appear to convey pAg recognition and responsiveness by Vγ9Vδ2 T cells. Lastly, we show that BTN2A1 binds to the side of the Vγ9 domain of the TCR, and also reveal the existence of a critical putative second ligand-binding domain on a separate region of the TCR that incorporates Vδ2. Disruption of either of these binding sites abrogated the ability of Vγ9Vδ2 T cells to respond to pAg. CONCLUSION Our findings suggest that γδ T cells recognize pAg in an entirely different way to how any other immune cell recognizes antigen. We propose a model whereby BTN2A1 and BTN3A1 co-bind the Vγ9Vδ2 TCR in response to pAg. This pAg likely modifies the BTN2A1–BTN3A1 complex to make it stimulatory, which may occur through BTN molecule remodeling and/or conformational changes. Targeting these molecules will create new opportunities for the development of γδ T cell–based immunotherapies for diseases in which pAgs are produced, including infections, autoimmunity, and cancer. Butyrophilin 2A1 plays a critical role in phosphoantigen recognition by human T cells. Human γδ T cells become activated in response to microbial and cancer-derived phosphoantigens, but the molecular mechanism for phosphoantigen recognition remained unclear. We show that a critical component of this recognition is the cell-surface molecule butyrophilin 2A1 (BTN2A1), which binds to the γδ TCR and in conjunction with BTN3A1, signals the presence of phosphoantigens to γδ T cells. Gamma delta (γδ) T cells are essential to protective immunity. In humans, most γδ T cells express Vγ9Vδ2+ T cell receptors (TCRs) that respond to phosphoantigens (pAgs) produced by cellular pathogens and overexpressed by cancers. However, the molecular targets recognized by these γδTCRs are unknown. Here, we identify butyrophilin 2A1 (BTN2A1) as a key ligand that binds to the Vγ9+ TCR γ chain. BTN2A1 associates with another butyrophilin, BTN3A1, and these act together to initiate responses to pAg. Furthermore, binding of a second ligand, possibly BTN3A1, to a separate TCR domain incorporating Vδ2 is also required. This distinctive mode of Ag-dependent T cell activation advances our understanding of diseases involving pAg recognition and creates opportunities for the development of γδ T cell–based immunotherapies.

中文翻译：

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Butyrophilin 2A1 对 γδ T 细胞的磷酸抗原反应性至关重要

一种识别磷酸抗原的奇怪方式 与经过充分研究的 αβ T 细胞相比，αβ T 细胞识别主要组织相容性复合体 (MHC) 和 MHC 样分子呈递的肽抗原，γδ T 细胞如何识别抗原在很大程度上仍然是个谜。一类主要的 γδ T 细胞，称为 Vγ9Vδ2+，被微生物和癌细胞产生的小的磷酸化非肽抗原或磷酸化抗原激活。里高等人。发现这些细胞需要在其细胞表面结合两个免疫球蛋白超家族成员，即嗜酪蛋白 2A1 (BTN2A1) 和 BTN3A1 来识别这些磷酸抗原。BTN2A1 直接结合 T 细胞受体 (TCR) 的 Vγ9+ 结构域，而第二个配体，可能是 BTN3A1，结合 TCR 另一侧的 Vδ2 和 γ 链区域。更好地理解这种意想不到的 T 细胞抗原识别形式应该会为未来的 γδ T 细胞介导的免疫疗法提供信息和增强。科学，这个问题 p。eaay5516 一个关键配体参与 γδ 免疫 T 细胞识别病原体或癌症相关的磷酸抗原。引言 T 细胞是免疫系统的一个关键组成部分，它可以通过称为 T 细胞受体 (TCR) 的同源细胞表面受体识别外来分子（抗原）。T 细胞的两个主要家族，称为 αβ 和 γδ T 细胞，由它们用于产生各自 TCR 的不同基因位点定义。αβ T 细胞通常识别与称为主要组织相容性复合体 (MHC) 分子的抗原呈递分子相关联的靶细胞表面上展示的抗原。关于 γδ T 细胞如何识别抗原知之甚少，尽管很明显它们对于保护性免疫也是必不可少的。在人类中，许多 γδ T 细胞（归类为 Vγ9Vδ2 T 细胞）对由细胞病原体和癌症产生的称为磷酸化抗原 (pAg) 的小磷酸化非肽抗原有反应。反过来，γδ T 细胞被激活、增殖，迅速产生促炎细胞因子，如 IFN-γ，并发挥细胞毒活性。pAg 识别似乎涉及细胞表面分子，即嗜酪蛋白 3A1 (BTN3A1)，它在该过程中发挥了必要但不充分的作用。因此，支持 Vγ9Vδ2 T 细胞识别 pAg 的分子基础仍不清楚，是一个长期存在的难题，阻碍了对这些重要免疫细胞的研究。基本原理 与 αβ T 细胞相反，Vγ9Vδ2 T细胞不受MHC限制，可以识别多种癌症和传染病表达的pAg。因此，它们代表了开发新免疫疗法的有吸引力的目标。需要更清楚地了解 pAg 识别的分子基础，以最佳地利用这些细胞进行免疫治疗。我们采取了多管齐下的方法来研究哪些分子是 γδ T 细胞检测 pAg 所必需的。我们使用全基因组筛选来鉴定介导 pAg 驱动的 γδ T 细胞活化的分子。此外，我们询问这些分子是否直接与 Vγ9Vδ2 TCR 结合，以及它们如何与 BTN3A1 结合使用。结果 在我们的全基因组筛选中鉴定出的最重要的候选分子是嗜酪蛋白 2A1 (BTN2A1)，该分子不同于 BTN3A1，但与 BTN3A1 相关。我们表明，如果没有 BTN2A1，Vγ9Vδ2 T 细胞不能被细菌或哺乳动物 pAg 激活，并且 BTN2A1 表达是 Vγ9Vδ2 T 细胞介导的肿瘤细胞杀伤所必需的。BTN3A1 和测试的其他嗜酪蛋白家族成员都不能弥补 BTN2A1 的损失。BTN2A1可以直接与Vγ9Vδ2 TCR结合，并与靶细胞表面的BTN3A1紧密结合。我们还确定了 BTN2A1 的跨膜和/或细胞内结构域的重要作用。此外，pAg 介导的 γδ T 细胞活化需要 BTN2A1 和 BTN3A1 的共表达，它们似乎共同传达了 Vγ9Vδ2 T 细胞对 pAg 的识别和响应。最后，我们表明 BTN2A1 与 TCR 的 Vγ9 结构域的一侧结合，并且还揭示了在包含 Vδ2 的 TCR 的单独区域上存在关键的假定第二配体结合域。这些结合位点中的任何一个的破坏都会消除 Vγ9Vδ2 T 细胞对 pAg 的反应能力。结论 我们的研究结果表明，γδ T 细胞识别 pAg 的方式与任何其他免疫细胞识别抗原的方式完全不同。我们提出了一个模型，其中 BTN2A1 和 BTN3A1 共同结合 Vγ9Vδ2 TCR 以响应 pAg。这种 pAg 可能会修饰 BTN2A1-BTN3A1 复合物以使其具有刺激性，这可能通过 BTN 分子重塑和/或构象变化发生。靶向这些分子将为开发基于 γδ T 细胞的免疫疗法创造新的机会，用于治疗产生 pAg 的疾病，包括感染、自身免疫和癌症。Butyrophilin 2A1 在人类 T 细胞的磷酸抗原识别中起关键作用。人类 γδ T 细胞响应微生物和癌症衍生的磷酸抗原而被激活，但磷酸抗原识别的分子机制仍不清楚。我们表明这种识别的一个关键组成部分是细胞表面分子嗜乳蛋白 2A1 (BTN2A1)，它与 ​​γδ TCR 结合并与 BTN3A1 结合，向γδ T 细胞发出磷酸抗原的存在信号。Gamma delta (γδ) T 细胞对保护性免疫至关重要。在人类中，大多数 γδ T 细胞表达 Vγ9Vδ2+ T 细胞受体 (TCR)，这些受体对细胞病原体产生的磷酸抗原 (pAg) 做出反应，并由癌症过度表达。然而，这些γδTCRs识别的分子靶点是未知的。这里，我们将嗜酪蛋白 2A1 (BTN2A1) 鉴定为与 Vγ9+ TCR γ 链结合的关键配体。BTN2A1 与另一种嗜酪蛋白 BTN3A1 结合，它们共同作用以启动对 pAg 的反应。此外，还需要将第二个配体（可能是 BTN3A1）与包含 Vδ2 的单独 TCR 结构域结合。这种独特的 Ag 依赖性 T 细胞激活模式促进了我们对涉及 pAg 识别的疾病的理解，并为开发基于 γδ T 细胞的免疫疗法创造了机会。