IL-7R and c-kit interactions in thymopoiesis

描述（由申请人提供）：虽然T细胞的胸腺内发育 通过阳性和阴性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 来自造血干细胞（HSC）的前胸腺细胞和未成熟胸腺细胞是 不太了解。常见的淋巴祖细胞（CLP），前胸腺细胞和 未成熟的胸腺细胞表达两种细胞因子的受体， 参与淋巴样分化：白细胞介素-7（IL-7）和c-kit配体 （KL）。IL-7是基质来源的细胞因子，其对淋巴祖细胞具有增殖、抗凋亡和分化作用。IL-7受体（IL-7 R）由两个非共价连接的亚基IL-7 Ra和共同γ链（γ-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-7 Ra和γ-c在以下存在下异二聚化： IL-7，导致多种途径的激活，包括JAK-STAT、P13 K和ras-MAPK信号转导途径。通过同源重组使IL-7-/-的小鼠中的淋巴细胞生成缺陷比具有IL-7 RcC-/-基因型的小鼠中的淋巴细胞生成缺陷不那么严重，这表明存在利用IL-7 R的互补途径。KL是c-kit的配体，c-kit是一种由HSC和未成熟祖细胞（包括CLP和未成熟胸腺细胞）表达的受体酪氨酸激酶（RTK）。C-kit还激活多种途径，包括JAK-STAT 通路虽然有缺陷的HSC和红细胞区室的小鼠与 c-kit或KL基因座的突变是很好理解的，c-kit途径可以 也显著有助于T淋巴细胞生成。突变的小鼠 KL或c-kit具有缺陷的胸腺生成，这可以通过以下方法来纠正： 分别移植正常胸腺或正常HSC。我们有 产生IL-7-/- Kit w 4约/w 41和IL-7 Ra-/-Kit w 41/w 41双纯合 小鼠，这两种小鼠都具有胸腺生成的完全和协同损失， 提示IL-7 R和c-kit信号通路之间的相互作用。 初步分析表明，c-kit介导酪氨酸磷酸化， 与IL-7 R相关。 在拟议的研究中要检验的假设是，IL-7 R和c-kit 信号通路直接在T淋巴细胞分化中相互作用， 造血祖细胞这些研究将调查如何相互作用 IL-7 R和c-kit信号通路之间发生，包括生物化学和 受体结合的成像研究;识别独特的，冗余的， 或由JL-7 R（STAT 5）激活的STAT分子的协同相互作用 和c-kit（STAT 1）;以及每种途径下游基因表达的变化。 体外研究将与两者的体内相互作用相关 细胞因子受体途径中的合适的小鼠品系。除了他们 与淋巴分化的相关性，这些研究将提供一个模型， 细胞因子受体途径，一个涉及I型细胞因子受体和 其他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.