Antigen recognition and activation of B-cell receptors of HIV-1 broadly neutralizing antibodies

项目摘要/摘要 B细胞抗原受体结合后B细胞表面早期事件的研究 (Bcr)可以评估抗原(Ag)在体内驱动B细胞活化的潜力。相比之下， 对于聚簇模型，解离活化模型(DAM)提出了一种交替的构象状态 静息的BCR以及B细胞膜上的早期事件如何启动B细胞的激活。在大坝里，B细胞 激活依赖于Ag解离不活跃的、静止的BCR寡聚体(BCR)的能力 构象开放)，并与共刺激分子促进BCR的纳米级重组，导致 完全激活。HIV-1包膜(Env)蛋白设计的最新进展提供了几个关键的潜力 然而，免疫原是如何被参与抗HIV-1的B细胞上的BCR感受到这种病毒抗原的 产生广谱中和抗体(BNAbs)的免疫反应还不是很清楚。整体而言 这项建议的目的是在定量的生物物理和膜动力学测量之间架起桥梁 在DAM的背景下，银-BCR相互作用对体外和体内B细胞激活的影响。为了实现这个目标，我们将 首次在纳米尺度上研究和定义了银诱导的早期BCR激活事件和 共刺激信号是完全激活表达BCR的B细胞所必需的，这些BCR指定原型HIV bNAbs或 它们的前身。我们假设，为了最佳地启动HIV Env免疫原，即那些能够最好地 激活B细胞并启动强大的亲和力成熟，BCR暴露于Env抗原的结果 首先是纳米尺度的BCR构象开放，其次是BCR的重组和共刺激 B细胞膜上信号放大的分子/激酶。在目标1中，我们将执行生物物理 分析和研究B细胞膜动力学以确定HIV env Ag-BCR相互作用的性质 表达bNab或生殖系前体BCRs的B细胞的最佳激活。在目标2中，我们将使用高分辨率 测定诱导纳米尺度BCR构象开放的银结合特性的方法，其作用 共刺激和调节分子与B细胞膜的时空重组 HIV Env AGS激活。在目标3中，我们将测试评估的环境AGS的激活和驱动能力 “更易诱导”(非无能或轻度无能)bNab B细胞的亲和力成熟 敲门(KI)曲目。这样的研究结果将是第一个允许使用系统方法进行预测的研究结果 基于这些确定的抗原诱导的早期B细胞，HIV免疫原启动不同的bNab前体的潜力 细胞激活事件。从长远来看，我们的研究将促进免疫原的设计和预先选择 在动物模型中进行测试，并加快艾滋病毒-1疫苗的开发。

Project Summary/Abstract Elucidating the early events on the B cell surface following antigen engagement of the B cell antigen receptor (BCR) can provide an assessment of the in vivo potential of an antigen (Ag) to drive B cell activation. In contrast to the “clustering model”, the “Dissociation Activation Model” (DAM) proposes an alternate conformational state of the resting BCR and how early events on the B cell membrane initiates B cell activation. In the DAM, B cell activation is dependent on the ability of an Ag to dissociate the inactive, resting BCR oligomers (BCR conformational opening), and promote nanoscale reorganization of BCR with co-stimulatory molecules, leading to full activation. Recent advances in HIV-1 Envelope (Env) protein designs have provided several key potential immunogens, however, how such viral Ags are sensed by BCRs on B cells that participates in anti-HIV-1 immune responses that generate broadly neutralizing antibodies (bnAbs) is not well understood. The overall objective of this proposal is to bridge the quantitative biophysical and membrane dynamics measurements of Ag-BCR interactions to ex-vivo and in-vivo B cell activation in the context of the DAM. Towards this goal, we will for the first time, study and define at nanoscale resolution, Ag-induced early BCR activation events and the costimulatory signaling required for full activation of B cells expressing BCRs specifying prototype HIV bnAbs or their precursors. We hypothesize that for optimal priming HIV Env immunogens, i.e. those capable of best activating B-cells and initiating robust affinity maturation, the exposure of the BCR to the Env antigens results first in nanoscale BCR conformational opening, followed by reorganization of BCRs and co-stimulatory molecules/kinases for signal amplification on the B cell membrane. In Aim 1, we will perform biophysical analyses and study B cell membrane dynamics to define the properties of HIV Env Ag-BCR interactions for optimal activation of B cells expressing bnAb or germline precursor BCRs. In Aim 2, we will use high resolution assays to determine Ag-binding properties that induce nanoscale BCR conformational opening, the role of costimulatory and regulatory molecules and the spatio-temporal reorganization of the B cell membrane following activation with HIV Env Ags. In Aim 3, we will test evaluated Env Ags for their ability to activate and drive affinity maturation in “more amenable to induce”, (non-anergic or more “mildly” anergic) bnAb B cells in naïve knock-in (KI) repertoires. Results of such studies would be the first to allow a systematic method for predicting the potential of HIV immunogens to prime distinct bnAb precursors based on these defined Ag-induced early B cell activation events. In the long-term our studies will facilitate design and pre-selection of immunogens for testing in animal models and accelerate HIV-1 vaccine development.