Immune profiling by high throughput sequencing of complete antigen receptors

描述(由申请人提供)：适应性免疫系统有能力对几乎无限的病原体和环境威胁做出反应。识别这些不同靶点所需的广泛分子多样性是通过T和B淋巴细胞中抗原受体基因片段的重排和组合关联而产生的。这些可变基因片段的DNA序列决定了编码的异源二聚体T细胞受体和B细胞免疫球蛋白的抗原特异性。虽然免疫反应不是仅仅由特异性决定的，但抗原受体是淋巴细胞识别和参与的主要部位，直接将这一复杂的遗传信息与宿主防御、疫苗接种和自身免疫功能联系起来。因此，全面的免疫球蛋白和T细胞受体的序列清单是了解，甚至可能操纵获得性免疫反应的关键一步。然而，在一个复杂的淋巴细胞群体中确定不同抗原受体基因的不同序列面临许多挑战，最显著的是获取两个亚单位链的配对序列信息，这两个亚单位链有助于TH异二聚体抗原受体的特异性。这项应用的目标是开发一种高通量策略，克服这些挑战，并能够对配对的抗原受体亚单位进行序列分析。结合分子生物学、微流控技术和高通量测序，我将建立一种快速分析T细胞受体和B细胞免疫球蛋白序列的有效方法。这种方法将被用来研究稳定状态和对抗原攻击反应的淋巴细胞库的分子和功能。它还将用于在系统性红斑狼疮模型中研究特定抗原受体特异性在自身免疫发病机制中的作用。这些研究将极大地促进我们对健康和疾病中适应性免疫系统的理解。此外，该项目中开发的工具将对研究感染、疫苗接种和自身免疫等领域的免疫功能具有广泛的实用价值。它们还可用于在多种生物系统中以单细胞分辨率分析复杂种群 与公共卫生相关：免疫系统的T和B细胞保护身体免受感染，但也可能出现故障并导致疾病。这些重要的细胞在它们识别的靶点上极其多样化；这种多样性给用传统技术大量研究这些细胞带来了许多挑战。这个项目开发了一种新技术，用来研究复杂的T和B细胞群体，并调查这些细胞在感染、疫苗接种和自身免疫性疾病的背景下如何发挥作用。

DESCRIPTION (provided by applicant): The adaptive immune system has the capacity to respond to a virtually limitless array of pathogens and environmental threats. The extensive molecular diversity required to recognize such varied targets is generated through the rearrangement and combinatorial association of antigen receptor gene segments in T and B lymphocytes. The DNA sequence of these variable gene segments determines the antigen specificity of encoded heterodimeric T cell receptors and B cell immunoglobulins. Although an immune response is not dictated by specificity alone, the antigen receptor serves as the principal site for lymphocyte recognition and engagement, directly linking this complex set of genetic information to functions in host defense, vaccination, and autoimmunity. Therefore, a comprehensive sequence inventory of the immunoglobulin and T cell receptor repertoire is a key step towards understanding, and perhaps manipulating, the adaptive immune response. However, determining the distinct sequences of diverse antigen receptor genes within a complex lymphocyte population presents a number of challenges, most notably in acquiring paired sequence information for both of the subunit chains that contribute to the specificity of th heterodimeric antigen receptor. The goal of this application is to develop a high throughput strategy that overcomes these challenges and enables sequence analysis of paired antigen receptor subunits. Using a combination of molecular biology, microfluidics technology, and high throughput sequencing, I will establish an effective method for rapidly profiling T cell receptor and B cell immunoglobulin sequences. This approach will be used for molecular and functional studies of the lymphocyte repertoire in the steady-state and in response to antigenic challenge. It will also be used to investigate the contribution of particular antigen receptor specificities t autoimmune pathogenesis in a model of systemic lupus erythematosus. These studies will significantly advance our understanding of the adaptive immune system in health and disease. Furthermore, the tools developed in this project will have broad utility for studying immune function in areas such as infection, vaccination, and autoimmunity. They can also be adapted for profiling complex populations at single cell resolution in a wide variety of biological systems PUBLIC HEALTH RELEVANCE: T and B cells of the immune system protect the body from infection but can also malfunction and cause disease. These important cells are extremely diverse in the targets they recognize; such diversity presents many challenges to studying these cells in large numbers with traditional techniques. This project develops a new technology with which to study complex populations of T and B cells, and investigates how these cells function in the context of infection, vaccination, and autoimmune disease.