Age-related changes in the proteome and lipidome of the immunological synapse

描述（由申请方提供）：老年人中，与T细胞功能相关的T细胞功能下降导致感染性和肿瘤性疾病的发生率较高、恢复期较长以及发病率和死亡率升高。如通过IL-2产生和细胞增殖所测量的，受损的T细胞功能显著促进免疫衰老。IL-2产生中与免疫相关的减少与信号分子向免疫突触的募集缺陷有关，免疫突触是在T细胞和抗原呈递细胞之间形成的粘附连接。这已经通过使用免疫荧光显微镜来监测单个蛋白质向免疫突触的移位而得到证实。越来越多的证据表明，质膜被横向分隔成小的脂质微区，这些脂质微区聚结在活化的T细胞受体（TCR）周围，在那里它们促进IL-2产生所需的信号蛋白的募集。所得到的结构包含脂筏和蛋白质支架，并且在免疫突触内作为小的~ 200 nm的“信号体”存在。免疫突触的整体组成中与神经元相关的差异尚未以系统的方式进行评估。此外，负责受损的蛋白质募集到免疫突触的机制仍然未知。影响质膜组成的变化，特别是影响鞘脂代谢物神经酰胺的变化，对脂筏结构有显著影响，并可能影响信号体的组装。尚未检查质膜鞘脂组成中的神经递质相关变化。然而，我们的初步数据表明，来自老年小鼠的T细胞具有显著更高水平的特定鞘脂。细胞组成的这些变化可能导致衰老T细胞形成有效免疫突触的能力降低。我们假设TCR诱导的信号体在其蛋白质和脂质募集模式中表现出与年龄相关的差异，并且老年T细胞鞘脂组成的变化影响调节IL-2产生的下游信号级联。我们将使用增强的磁性免疫分离程序结合最先进的蛋白质组学和脂质组学方法来验证这一假设。我们将通过诱导细胞鞘脂中与年龄相关的变化以及测量免疫突触形成和T细胞活化来进一步验证这一假设。这些研究将提供第一次全面调查的蛋白质和脂质含量的信号复合物形成的抗原刺激。从这些免疫突触的蛋白质组学和脂质组学分析中获得的信息将构成构建T细胞中与年龄相关的变化的“地图”的基础。该图谱将增强我们对免疫衰老分子基础的理解，并将促进逆转与衰老相关的有害变化的策略的发展。与年龄相关的T细胞（对抵抗病原体和肿瘤很重要的免疫细胞）功能下降是老年人健康和生活质量的重要决定因素。这些缺陷的潜在机制尚不完全清楚。该项目将利用最先进的方法来构建T细胞脂质和蛋白质中与年龄相关的关键变化的“地图”。该图谱将增强我们对免疫衰老分子基础的理解，并将促进逆转与衰老相关的有害变化的策略的发展。

DESCRIPTION (provided by applicant): Age-associated declines in T cell function contribute to the higher incidences, longer recovery periods, and elevated morbidity and mortality from infectious and neoplastic diseases in the elderly. Impaired T cell function, as measured by IL-2 production and cell proliferation, contributes significantly to immune senescence. Age-related reductions in IL-2 production are associated with defects in the recruitment of signaling molecules to immune synapses, the adhesive junction that forms between a T cell and an antigen- presenting cell. This has been demonstrated by using immunofluorescence microscopy to monitor the translocation of individual proteins to the immune synapse. Accumulating evidence indicates that the plasma membrane is laterally compartmentalized into small lipid microdomains that coalesce around the activated T cell receptor (TCR), where they facilitate the recruitment of signaling proteins required for IL-2 production. The resulting structures incorporate both lipid rafts and protein scaffolds, and exist as small ~200nm 'signalosomes' within the immune synapse. Age-related differences in overall composition the immune synapse have not been evaluated in a systematic manner. Furthermore, the mechanisms responsible for the impaired recruitment of proteins to the immune synapse remain unknown. Changes affecting the composition of the plasma membrane, particularly changes affecting the sphigolipid metabolite ceramide, have significant impacts on lipid raft structure, and could influence the assembly of signalosomes. Age-related changes in the sphingolipid composition of the plasma membrane have not been examined. However, our preliminary data indicate that T cells from old mice have significantly higher levels of specific sphingolipids. These changes in cellular composition may contribute to the reduced ability of aged T cells to form effective immune synapse. We hypothesize that TCR-induced signalosomes exhibit age-related differences in their patterns of protein and lipid recruitment, and that changes in the sphingolipid composition of aged T cells impact the downstream signaling cascades that regulate IL-2 production. We will test this hypothesis using an enhanced magnetic immunoisolation procedure in conjunction with state-of-the-art proteomic and lipidomic methods. We will further test this hypothesis by inducing age-related changes in cellular sphingolipids and measuring immune synapse formation and T cell activation. These studies will provide the first comprehensive investigation of the protein and lipid content of the signaling complexes formed in response to antigenic stimulation. The information obtained from these proteomic and lipidomic analyses of the immune synapse will form the basis for constructing a 'map' of age-related changes in T cells. This map will enhance our understanding of the molecular bases of immune senescence, and will promote the development of strategies to reverse the detrimental changes associated with aging. The age related decline in T cell (immune cells important for fighting against pathogens and tumors) function is an important determinant of the health and quality of life in elderly. The underlying mechanisms of these defects are not completely known. This project will utilize state-of-the-art methods to construct a 'map' of the key age-related changes in lipids and proteins of T cells. This map will enhance our understanding of the molecular bases of immune senescence, and will promote the development of strategies to reverse the detrimental changes associated with aging.