Structural Biology of Apoptosomes and Related Signaling Complexes

凋亡体和相关信号复合物的结构生物学

• 批准号: 7544974
• 负责人: CHRISTOPHER W AKEY
• 金额: $ 23.56万
• 依托单位: BOSTON UNIVERSITY MEDICAL CAMPUS
• 依托单位国家: 美国
• 财政年份: 2001
• 资助国家: 美国
• 起止时间: 2001-07-01 至 2010-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7544974
• 关键词: Animals; Apical; Apoptosis; Apoptotic; Authorization documentation; Bacteria; Baculoviruses; Binding; Boston; Cardiac; Caspase; Cell Death; Cell Line; Cell Proliferation; Cells; Cellular Stress; Cessation of life; Chronic; Complex; Crohn' s disease; Crystallization; Cues; Defect; Degenerative Disorder; Development; Disclosure; Drosophila genus; Electrons; Enzymes; Eukaryota; Event; Face; Family; Genes; Goals; Grant; Head; Homologous Gene; Homology Modeling; Human; Human Resources; Immune response; Immunity; In Vitro; Individual; Inflammation; Inflammatory; Instruction; Intestines; Ischemia; Last Name; Lead; Longitudinal Studies; Major Depressive Disorder; Malignant Neoplasms; Maps; Mediator of activation protein; Methods; Michigan; Mitochondria; Modeling; Modus; Multienzyme Complexes; Muscle Cells; Names; Nucleotides; Organism; Pathway interactions; Phase; Play; Principal Investigator; Printing; Process; Proteins; Recruitment Activity; Registries; Research Personnel; Research Project Grants; Resolution; Role; Science; Series; Signal Transduction; Skeletal Muscle; Structure; Syndrome; Technology; Testing; Texas; Time; Tissues; Universities; Work; X-Ray Crystallography; apoptotic protease-activating factor 1; cell growth; cell killing; cytochrome c; graduate student; human embryonic stem cell; improved; medical schools; member; novel; nucleoside triphosphatase; particle; pathogen; pro-caspase-9; programs; research study; response; small bowel Crohn' s disease; structural biology; three dimensional structure; time use; transcription factor

Programmed cell death (apoptosis) is a process whereby individual cells are terminated to benefit the organism. However, many cancers inhibit apoptotic pathways to allow cell proliferation, while degenerative diseases may up-regulate apoptosis to kill cells prematurely. In the intrinsic cell death pathway, cytochrome c is released from mitochondria and interacts with Apaf-1 in the presence of dATP. This triggers apoptosome assembly and the resulting platform recruits and activates procaspase-9. This holo-enzyme then activates executioner procaspases which kill the cell. Significantly, Apaf-1 assembly has recently been implicated as a possible contributing factor to major depression syndrome (MDD). In Drosophila, a similar cell killing machine is formed by Dark, an Apaf-1 Related Killer which sequentially activates the procaspases Drone and DrICE. In Specific Aims 1 and 2, we will determine high resolution, 3-dimensional structures of the Apaf-1 and Dark apoptosomes. In addition, structures will be obtained of "holo-enzymes" with prodomains from their apical procaspases. In these studies, electron cryo-microscopy and single particle methods, X-ray crystallography and homology modeling will be used to create atomic models. These studies will provide a detailed picture of human and Drosophila apoptosomes and will also reveal conformational changes that occur when procaspases bind. Thus, Specific Aims 1 and 2 will give a clearer understanding of how these large platforms assemble and function in the intrinsic cell death pathway. In pro-inflammatory pathways, the Apaf-1 related protein NOD2 senses bacteria and assembles a signaling platform. Through a series of steps, this complex activates the transcription factor NFkB, which up-regulates genes involved in innate or adaptive immunity. Defects in NOD2 are associated with chronic inflammation of the small intestine (Crohn's disease) and with Blau's syndrome. In Specific Aim 3, a NOD2 signaling platform will be assembled and the first 3-dimensional structure of this complex will be determined. In the long term, these studies will help us to understand how NOD2 functions in the innate immune response to bacteria in the gastro-intestinal tract.

程序性细胞死亡（细胞凋亡）是个体细胞被终止以有益于生物体的过程。然而，许多癌症抑制凋亡途径以允许细胞增殖，而退行性疾病可能上调凋亡以过早杀死细胞。在内源性细胞死亡途径中，细胞色素c从线粒体释放，并在dATP存在下与Apaf-1相互作用。这触发了核糖体组装，并且所产生的平台募集并激活了半胱氨酸天冬氨酸蛋白酶原-9。这种全酶然后激活刽子手前半胱氨酸蛋白酶，杀死细胞。值得注意的是，Apaf-1组装最近被认为是重性抑郁综合征（MDD）的一个可能的促成因素。在果蝇中，一个类似的细胞杀伤机器是由Dark形成的，Dark是一种Apaf-1相关的杀手，它依次激活前半胱氨酸蛋白酶Drone和DrICE。 在具体目标1和2中，我们将确定Apaf-1和Dark染色体的高分辨率三维结构。此外，结构将获得的“全酶”的前域从他们的顶端procaspase。在这些研究中，电子冷冻显微镜和单粒子方法，X射线晶体学和同源建模将用于创建原子模型。这些研究将提供一个详细的图片人类和果蝇染色体，也将揭示构象变化时发生的前半胱氨酸蛋白酶结合。因此，具体目标1和2将更清楚地了解这些大平台如何在内在细胞死亡途径中组装和发挥作用。 在促炎途径中，Apaf-1相关蛋白NOD 2感知细菌并组装信号平台。通过一系列步骤，这种复合物激活转录因子NFkB，其上调参与先天性或适应性免疫的基因。NOD 2缺陷与小肠慢性炎症（克罗恩病）和Blau综合征有关。在具体目标3中，将组装NOD 2信号平台，并确定该复合物的第一个三维结构。从长远来看，这些研究将有助于我们了解NOD 2在胃肠道细菌的先天免疫反应中的作用。