The Role of Kuzbanian and TACE in T Cell Development

描述(申请人提供)：信号转导的Notch通路 对细胞命运的决定很重要。我们实验室的研究首先 提示Notch信号在胸腺发育中发挥作用，导致 接收Notch信号的未成熟胸腺细胞发育为CD8T的模型 细胞，而没有收到Notch信号的T细胞成熟为CD4T细胞。 Notch-1受体的蛋白水解性加工是产生 Notch-1介导的信号和一种Notch配体Delta经历蛋白分解 在果蝇体内进行加工。然而，是否处理小鼠Notch同源基因 和Notch配体发生，所有负责这一过程的蛋白酶 都是未知的。金属蛋白酶Kuzbanian(Kuz)和TACE已被 与Notch受体和配体的加工有关。作为凹槽的加工 是Notch信令所必需的，Notch配体处理过程中的阻塞或 受体可能导致Notch信号的减少。我们将确定 KUZ和TACE的潜在Notch受体/配体底物在 胸腺和分析Notch受体/配体的功能后果 分裂对胸腺发育的影响。我们预测在过度表达A基因的胸腺细胞中 显性阴性的Kuz或TACE基因，可能比CD8T细胞更多的CD4T细胞 发展。反过来，在过度表达野生型Kuz或TACE的胸腺细胞中，更多 CD8T细胞比CD4T细胞会发育。这些预测将得到检验。 在体外和转基因小鼠中。已知缺口的表达和处理 胸腺中的受体和配体将通过定量Western进行评估 免疫印迹和流式细胞仪检测T细胞发育情况。相关性 在这些系统中处理和T细胞发育之间的关系将是 调查过了。

DESCRIPTION (provided by applicant): The Notch pathway of signal transduction is important for cell fate decisions. Studies from our laboratory first suggested a role for Notch signaling in thymic development, resulting in a model in which immature thymocytes receiving a Notch signal develop into CD8+ T cells, whereas T cells not receiving a Notch signal mature into CD4+ T cells. Proteolytic processing of the Notch-1 receptor is required for generation of Notch-1-mediated signals, and Delta, a Notch ligand, undergoes proteolytic processing in Drosophila. However, whether processing of mouse Notch homologues and Notch ligands occurs and all the proteases responsible for such processing are not known. The metalloproteases Kuzbanian (Kuz) and TACE have been implicated in processing of Notch receptors and ligands. As processing of Notch is required for Notch signaling, a block in processing of Notch ligands or receptors might result in a decrease in Notch signaling. We will identify the potential Notch receptor/ligand substrates of Kuz and TACE expressed in the thymus and analyze the functional consequences of Notch receptor/ligand cleavage on thymic development. We predict that in thymocytes overexpressing a dominant-negative Kuz or TACE gene, more CD4+ T cells than CD8 T cells might develop. Reciprocally, in thymocytes overexpressing wild-type Kuz or TACE, more CD8+ T cells than CD4+ T cells will develop. These predictions will be tested in vitro and in transgenic mice. Expression and processing of the known Notch receptors and ligands in the thymus will be assessed by quantitative Western blotting, and T cell development assessed by flow cytometry. Correlations between processing and T cell development in these systems will be investigated.