Involvement of Nonclassical MHC in Early T Cell Ontogeny in Xenopus

非经典 MHC 参与非洲爪蟾早期 T 细胞个体发育

• 批准号: 7701182
• 负责人: JACQUES Robert
• 金额: $ 7.7万
• 依托单位: UNIVERSITY OF ROCHESTER
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-08-15 至 2011-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7701182
• 关键词: Accounting; Adopted; Adult; Amphibia; Antibodies; Biological; Biological Metamorphosis; Bypass; CD8B1 gene; Cell Differentiation process; Cell Lineage; Cell Maturation; Cell surface; Cells; Communities; Complex; Dependence; Dependency; Development; Education; Embryo; Epithelial; Flow Cytometry; Future; Gene Transfer Techniques; Generations; Genes; Gills; Goals; Growth; Hybridomas; Immune; Immune system; Immunity; Immunization; Immunocompetent; Immunology; In Situ Hybridization; J segment gene; Jaw; Larva; Life; Ligands; Liver; Lymphocyte; Mature T-Lymphocyte; Mediating; Messenger RNA; Modeling; Molecular Profiling; Mus; Pathway interactions; Pattern; Peptides; Peripheral; Phage Display; Phylogenetic Analysis; Physiological; Play; Process; Proteins; Protocols documentation; RNA Interference; Rana; Reagent; Recombinants; Regulation; Relative (related person); Research Personnel; Role; Signal Transduction; Spleen; Staging; Surface; T-Cell Development; T-Cell Receptor; T-Lymphocyte; TCF Transcription Factor; Technology; Testing; Thymus Gland; Vertebrates; Work; Xenopus; Xenopus laevis; base; comparative; genome sequencing; improved; in vivo; insight; neonate; novel; pathogen; public health relevance; receptor; response; technology development; thymocyte; transcription factor; tumor; xenopus development

DESCRIPTION (Provided by Applicant): The overall goal of this application is to explore the involvement of non-classical MHC class Ib (class Ib) molecules in thymocyte differentiation during early development by a comparative approach in the Xenopus. Increasing evidence suggests that in the absence or suboptimal expression of 'classical' MHC class Ia (class Ia) as it occurs in neonates, class Ib molecules can generate survival signals to differentiating thymocytes. Not much is known about the physiological relevance of this alternative thymic selection process. It is hypothesized that in the absence (or suboptimal level) of class Ia expression early in development, thymic differentiation and T cell education critically depends on class Ib. Both the thymic dependence of T cells and the factors that play key roles in the regulation of T cell development are broadly conserved among jawed vertebrates. Therefore, the investigators propose to use the Xenopus and its immunocompetent but naturally class Ia-deficient larval stage as a model because it provides the advantage of having the early developing thymus of larvae externally visible, easily accessible, and amenable to experimental manipulation. Importantly, the hypothesis involving class Ib can be tested in the natural absence of class Ia, and the development of T cells can be studied in a biological context. Finally, the investigators have recently characterized a novel Xenopus nonclassical MHC class Ib (XNC) gene subfamily, XNC10, whose expression is tightly associated with the CD8 T-cell lineage during thymocyte differentiation since early in ontogeny. Therefore, they propose to test the hypothesis that class Ib genes are critically involved in thymic differentiation and T cell education during ontogeny of Xenopus. This will be achieved by the two following aims: 1. To determine the expression pattern of XNC10 and a selected set of immune genes during ontogeny: This will involve in situ hybridization as well as immunohistology and flow cytometry analysis for which they will generate new antibodies against XNC10 and other thymocytes markers (e.g., CD4, CD8¿, TCR¿, a); and 2. To assess the function of XNC10 in T cell differentiation and thymic education, which will involve in vivo knockdown by RNA interference using transgenesis to reveal the function of XNC10 in CD8 T cell development. This application will provide fundamental, and not phylogenetically restricted, insight on the relative contributions of class Ia versus class Ib-mediated thymocyte education pathways during ontogeny. Technologies developed and reagents obtained will be of high benefit for the scientific community and will considerably increase the potential of X. laevis as a non-mammalian alternative model. PUBLIC HEALTH RELEVANCE: The goal of this project is to better understand the possible role of certain MHC nonclassical class I molecules in assisting the development of fully mature T cells during early stage of life and ultimately helping their response against pathogens or tumors prone to escape the host's immune defenses. For this, the investigators will use an evolutionary approach that takes advantage of the frog X. laevis, which is a unique non-mammalian comparative model that permits easy access of the thymus early during development.

描述（由申请人提供）：本申请的总体目标是通过非洲爪蟾的比较方法来探索非经典 MHC Ib 类（Ib 类）分子在早期发育过程中参与胸腺细胞分化。当新生儿中出现“经典”MHC Ia 类（Ia 类）缺失或表达欠佳时，Ib 类分子可以产生分化胸腺细胞的生存信号。人们对这种替代性胸腺选择过程的生理学相关性了解甚多，人们再次认识到，在发育早期 Ia 类表达缺失（或达不到最佳水平）的情况下，胸腺分化和 T 细胞教育关键取决于 Ib 类胸腺依赖性。 T 细胞的功能以及在 T 细胞发育调节中起关键作用的因子在有颌脊椎动物中广泛保守，因此，研究人员建议使用非洲爪蟾及其具有免疫活性但天然存在 Ia 类缺陷的细胞。幼虫阶段作为模型，因为它提供了幼虫早期发育胸腺外部可见、易于接近且易于实验操作的优点。重要的是，涉及 Ib 类的假设可以在 Ia 类自然不存在的情况下进行检验。最后，研究人员最近鉴定了非洲爪蟾非经典 MHC Ib 类 (XNC) 基因亚家族 XNC10，其表达与 CD8 密切相关。自个体发育早期以来，胸腺细胞分化过程中的 T 细胞谱系因此，他们提出检验 Ib 类基因在非洲爪蟾个体发育过程中胸腺分化和 T 细胞教育中至关重要的假设。这将通过以下两个目标来实现。确定 XNC10 和一组选定的免疫基因在个体发育过程中的表达模式：这将涉及原位杂交以及免疫组织学和流式细胞术分析，为此他们将产生针对 XNC10 的新抗体。和其他胸腺细胞标记物（例如 CD4、CD8¿ ,TCR?? ，a)；和2.评估XNC10在T细胞分化和胸腺教育中的功能，这将涉及使用转基因通过RNA干扰进行体内敲低，以揭示XNC10在CD8 T细胞发育中的功能。 该应用将为个体发育过程中 Ia 类与 Ib 类介导的胸腺细胞教育途径的相对贡献提供基本的、不受系统发育限制的见解。所开发的技术和获得的试剂将为科学界带来很大好处，并将大大增加潜力。 X. laevis 作为非哺乳动物替代模型。 公共健康相关性：该项目的目标是更好地了解某些 MHC 非经典 I 类分子在生命早期阶段协助完全成熟 T 细胞发育的可能作用，并最终帮助它们对易于逃脱的病原体或肿瘤做出反应。为此，研究人员将使用一种利用青蛙 X. laevis 的进化方法，这是一种独特的非哺乳动物比较模型，可以在发育早期轻松进入胸腺。