IL-7R and c-kit interactions in thymopoiesis

描述(申请人提供)：虽然胸腺内T细胞的发育 通过正选择和负选择的淋巴细胞已经被很好地表征 在过去的二十年里，规范发展的过程 前胸腺细胞和未成熟胸腺细胞来自造血干细胞(HSC) 人们对此知之甚少。共同淋巴祖细胞(CLP)、前胸腺细胞和 未成熟胸腺细胞表达两种细胞因子的受体，这两种细胞因子已知是 参与淋系分化的细胞因子：白介素7和c-kit配体 (九龙)。IL-7是一种基质来源的细胞因子，对淋巴祖细胞具有增殖、抗凋亡和分化作用。IL-7受体(IL-7R)由两个非共价连接的亚基IL-7ra和共同的伽马链(Gamma-c)组成，这两个亚基都是I型细胞因子受体超家族的成员。IL-7Ra和γ-c在 IL-7激活多条通路，包括JAK-STAT、P13K和ras-MAPK信号转导通路。通过同源重组产生IL-7-/-的小鼠的淋巴系缺陷比携带IL-7RcC-/-的小鼠要轻，这表明存在一种利用IL-7R的互补途径。KL是c-kit的配体，c-kit是一种受体酪氨酸激酶(RTK)，由HSC和包括CLP和未成熟胸腺细胞在内的未成熟祖细胞表达。C-KIT还激活多条通路，包括JAK-STAT 路径。尽管小鼠有缺陷的HSC和红系隔室 C-kit或KL基因座的突变是公认的，c-kit途径可以 对T淋巴细胞的生成也有显著的贡献。任何一种基因突变的小鼠 KL或c-kit有胸腺生成缺陷，可通过以下方法纠正 分别移植正常胸腺和正常HSC。我们有 产生的IL-7-/-Kit w4约/w4l和LL-7Ra-/-Kitw41/w41双重纯合 小鼠，两者都有完全的和协同的胸腺生成丧失， 提示IL-7R和c-kit信号通路之间存在相互作用。 初步分析表明，c-kit介导酪氨酸磷酸化和 与IL-7R相关。 在拟议的研究中要检验的假设是IL-7R和c-kit 信号通路在原始人T淋巴样分化中的直接作用 造血祖细胞。这些研究将调查相互作用是如何 在IL-7R和c-kit信号通路之间发生，包括生化和 受体关联的成像研究；唯一的、冗余的、 或JL-7R激活的STAT分子的协同作用(STAT5) 和c-kit(STAT1)；以及每条通路下游基因表达的变化。 体外研究将与两者在体内的相互作用相关。 适当的小鼠品系中的细胞因子受体通路。除了他们的 与淋巴分化的相关性，这些研究将提供一个模型，说明 细胞因子受体途径，其中一条涉及I型细胞因子受体和 另一种是RTK，在淋巴造血分化过程中相互作用。

DESCRIPTION (provided by applicant): While the intrathymic development of T lymphocytes via positive and negative selection has been well characterized over the past two decades, the processes which regulate the development of pro-thymocytes and immature thymocytes from hematopoietic stem cells (HSC) are poorly understood. Common lymphoid progenitors (CLP), prothymocytes and immature thymocytes express receptors for two cytokines which are known to be involved in lymphoid differentiation: interleukin-7 (IL-7) and c-kit ligand (KL). IL-7 is a stromal derived cytokine which has proliferative, anti-apoptotic and differentiative effects on lymphoid progenitors. The IL-7 receptor (IL-7R) consists of two non-covalently linked subunits, IL-7Ra and the common gamma chain (gamma-c), both of which are members of the type I cytokine receptor superfamily. The IL-7Ra and gamma-c heterodimerize in the presence of IL-7, resulting in activation of multiple pathways, including the JAK-STAT, P13K, and ras-MAPK signal transduction pathways. The lymphopoietic defect in mice rendered IL-7-/-by homologous recombination is less severe than that of mice with an IL-7RcC-/-genotype, suggesting the existence of a complementary pathway that utilizes the IL-7R. KL is the ligand for c-kit, a receptor tyrosine kinase (RTK) expressed by HSC and immature progenitors, including CLP and immature thymocytes. C-kit also activates multiple pathways, including the JAK-STAT pathway. Although the defective HSC and erythroid compartments of mice with mutations of the c-kit or KL loci are well appreciated, the c-kit pathway can also significantly contribute to T lymphopoiesis. Mice with mutations of either KL or c-kit have defective thymopoiesis, which can be corrected by transplantation of a normal thymus or normal HSC, respectively. We have generated IL-7-/- Kit w4 about/w4l and lL-7Ra-/- Kitw41/w41 doubly homozygous mice, both of which have a complete and synergistic loss of thymopoiesis, suggesting an interaction between the IL-7R and c-kit signaling pathways. Preliminary analyses show that c-kit mediates tyrosine phosphorylation and associates with IL-7R. The hypothesis to be tested in the proposed studies is that the IL-7R and c-kit signaling pathways directly interact in T lymphoid differentiation of primitive hematopoietic progenitors. The studies will investigate how interactions between the IL-7R and c-kit signaling pathways occur, including biochemical and imaging studies of receptor association; identification of unique, redundant, or synergistic interactions of STAT molecules activated by the JL-7R (STAT5) and c-kit (STAT1); and changes in gene expression downstream of each pathway. The in vitro studies will be correlated with in vivo interactions of the two cytokine receptor pathways in appropriate mouse strains. Besides their relevance to lymphoid differentiation, the studies will provide a model for how cytokine receptor pathways, one involving a Type I cytokine receptor and the other an RTK, interact during lymphohematopoietic differentiation.