GLUCOCORTICOIDS AND LYMPHOCYTE CATABOLISM

拟议研究的总体目标是了解机制 糖皮质激素通过该作用抑制免疫系统的功能。 特别是，我们将集中我们的研究阐明分子事件 参与胸腺细胞死亡的调节， 糖皮质激素 在几个实验室进行的研究，包括我们的 我们已经发现糖皮质激素调节的淋巴细胞溶解通过以下方式发生： 受体依赖机制激活固有的程序性呼叫死亡 这一过程通常被称为“凋亡”。 生物学上的标志 所有细胞凋亡形式的细胞死亡在深刻的核小体间 在任何可检测的损失之前发生的基因组DNA片段化， 称之为生存能力 因此，负责该DNA的核酸酶 降解是凋亡过程的关键成分。 期间 在上一个授予期，我们成功地进行了鉴定， 纯化和测定了一种新的抗人源性大肠杆菌的部分氨基酸序列， 具有固有的核小体间DNA切割的“独特”核酸酶 活动 在这项研究计划中，我们建议克隆这种基因， 核酸酶，并使用克隆的基因直接测试假设， 核小体间DNA断裂是细胞死亡的原因。 的 克隆的核酸酶基因的可用性将使我们能够解决 以前无法解决的问题，有关的监管模式， 糖皮质激素诱导的细胞凋亡。 此外，我们将定义分子 组织和细胞类型特异性凋亡的基础。 为了验证这一假设 并解决随之而来的问题，我们提出以下具体目标：（一） 大鼠糖皮质激素调节核酸酶基因的克隆及序列分析 胸腺细胞(II)为了验证克隆的核酸酶的真实性， 决定其在细胞凋亡形式的细胞死亡中的作用。(III)来定义 调节核酸酶基因表达的组织特异性机制， 凋亡和非凋亡细胞中的核酸酶活性。 知识 从这些研究中获得的信息应该可以定义精确的细胞 激活细胞凋亡生理过程的机制 提供免疫抑制机制的关键见解， 糖皮质激素

The overall goal of the proposed research is to understand the mechanism(s) by which glucocorticoids act to suppress the function of the immune system. In particular, we will focus our studies elucidating the molecular events that are involved in the regulation of thymocyte cell death by glucocorticoids. Studies undertaken in several laboratories including our own have revealed that glucocorticoid regulated lymphocytolysis occurs by receptor dependent mechanisms which activate inherent programmed call death processes commonly referred to as "apoptosis". The biological hallmark of all apoptosis forms of cell death in profound internucleosomal fragmentation of genomic DNA which occurs prior to any detectable loss in call viability. Accordingly, the nuclease responsible for this DNA degradation is a key component to the apoptotic process. During the previous granting period, we have succeeded in the identification, purification and determination of a partial amino acid sequence for a "unique" nuclease which has inherent internucleosomal DNA cleavage activity. In this research proposal, we propose to clone the gene for this nuclease and use the cloned gene to directly test the hypothesis that internucleosomal DNA fragmentation is the cause of cell death. The availability of the cloned nuclease gene will permit us to address previously unapproachable questions pertaining to the mode of regulation of apoptosis by glucocorticoids. Additionally, we will define the molecular basis for tissue and call type specific apoptosis. To test this hypothesis and address the ensuing issues, we propose the following specific aims: (I) To clone and sequence the glucocorticoid regulated nuclease from rat thymocytes. (II) To verify the authenticity of the cloned nuclease and determine its role in apoptotic forms of cell death. (III) To define the tissue specific mechanisms that regulate nuclease gene expression and nuclease activity in both apoptotic and non-apoptotic cells. The knowledge gained from these investigations should define the precise cellular mechanisms which activate the physiological process of apoptosis and provide key insights into the mechanisms of immunosuppression by glucocorticoids.