Crosstalk and the cytoskeleton in dendritic cell antigen presentation

 描述（由申请人提供）：在这里，我们试图了解树突状细胞（DC）肌动蛋白细胞骨架如何控制抗原呈递给同源T细胞，以及两种细胞之间的双向通讯如何重新排列细胞骨架网络以协调免疫应答。DC通过向应答T细胞传递环境特异性信息来启动抗原呈递。正确的信息驱动T细胞动态地将它们的肌动蛋白网络重新定向到DC-T细胞接触的位点，并组装肌动蛋白连接的细胞内信号传导组分。这导致诱导细胞行为和基因表达变化的长期持续信号事件，从而导致适应性免疫的发生。虽然肌动蛋白网络在T细胞中的作用已经得到很好的证实，但对C细胞骨架在抗原呈递和免疫中的作用知之甚少。目前还不清楚DC和T细胞之间的串扰如何通过免疫突触（IS）处的受体-配体接合重新排列DC细胞骨架以影响无数的T细胞反应。先前的研究强烈支持DC肌动蛋白网络在抗原呈递中发挥重要作用。此外，我们的初步工作突出了CD 40信号在DC中重排肌动蛋白细胞骨架网络以促进有效抗原呈递中的作用。我们假设，通过将DC肌动蛋白细胞骨架调制成刚性和刚性网络，能够将通信中继到同源应答T细胞。然后，DC网络的这种转变将支持应答T细胞信号传导分子的反式迁移、组装和活化。在这个项目中，我们将描述CD 40信号转导在指导DC上调和招募肌动蛋白捆绑蛋白Fascin到DC IS中的作用。在目标1A中，我们将定义CD 40连接在调节肌成束蛋白差异表达、磷酸化-激活状态和肌成束蛋白在IS的定位中的作用。目的1B将描述CD 40下游的细胞内信号事件与在IS聚集的CD 40-CD 40 L在调节成束蛋白活性中的作用。最后，在目标1C中，我们评估了野生型与显性阴性突变体肌成束蛋白拯救应答T细胞的CD 40缺陷DC引发的能力。结果将提供一个更好的理解的DC细胞骨架在抗原呈递中发挥的作用，并提出了一个更大的框架的研究模型，描述：（a）极化货物运输沿着肌动蛋白捆绑网络;（B）受体-配体的流动性和膜流动性在肌动蛋白捆绑网站;和（c）微管网络动态DC和各自的贡献T细胞反应。这些研究中发现的信息可能为可调人工抗原呈递材料的开发和基于DC的免疫疗法的合理设计提供蓝图。

 DESCRIPTION (provided by applicant): Here we seek to understand how the dendritic cell (DC) actin cytoskeleton governs antigen presentation to cognate T cells and also how bidirectional communication between the two cells can rearrange cytoskeletal networks to orchestrate immune responses. DC initiates antigen presentation by imparting context specific information to responder T cells. Correct information drives T cells to dynamically reorient their actin network towards the site of DC-T cell contact and assemble actin-linked intracellular signaling components. This results in long-term sustained signaling events that induce cell behavioral and gene expression changes, thereby leading to onset of adaptive immunity. Although contribution of the actin network in T cells is well established, little is understood of C cytoskeletal roles in antigen presentation and immunity. It is also unclear how crosstalk between DC and T cells through receptor-ligand engagement at the immunological synapse (IS) could rearrange the DC cytoskeleton to influence the myriad of T cell responses. Previous studies strongly support the DC actin network playing significant roles in antigen presentation. Additionally, our preliminary work highlights the role of CD40 signaling in rearranging the actin cytoskeletal network in DC to promote efficient antigen presentation. We hypothesize that the ability to relay communication to cognate responder T cells occurs through modulation of the DC actin cytoskeleton into stiff and rigid networks. This transition of the DC network would then support in trans mobility, assembly, and activation of responder T cell signaling molecules. In this project, we will delineate the role of CD40 signaling in directing DC to upregulate and recrui the actin-bundling protein fascin to the DC IS. In Aim 1A we will define the role of CD40 ligation in modulating differential fascin expression, phosphorylation-activation states, and localization o fascin at the IS. Aim 1B will delineate the downstream intracellular signaling events of CD40 vs. the role of CD40-CD40L clustering at the IS in regulation of fascin activities. Finally, in Aim 1C we evaluate wild type vs. dominant negative mutant fascin ability to rescue CD40-defecient DC priming of responder T cells. Results will provide a better understanding of the role the DC cytoskeleton plays in antigen presentation and propose a model for a larger framework of studies describing: (a) polarized cargo transport along actin-bundled networks; (b) receptor- ligand mobility and membrane fluidity at actin-bundled sites; and (c) microtubule network dynamics in DC and respective contribution to T cell responses. Information uncovered in these studies may provide blueprints for development of tunable artificial antigen presenting material and rational design of DC-based immunotherapies.