Molecular Mechanism of Granule-mediated Apoptosis

细胞毒性细胞颗粒介导的细胞凋亡是一种独特的细胞信号转导形式，需要细胞内 穿孔蛋白穿孔蛋白传递颗粒相关丝氨酸蛋白酶(颗粒酶)。这个 对颗粒分泌途径中的遗传缺陷的鉴定清楚地表明，颗粒- 介导的细胞凋亡对免疫稳态起着至关重要的作用。获得性细胞毒性细胞效应器功能障碍 病毒蛋白可能引发淋巴增殖和自身反应，并降低宿主防御能力。 响应性。病毒感染的细胞或肿瘤细胞也可能通过抑制颗粒介导的凋亡来抵抗 这条途径的几乎所有方面。描绘了这一鲜为人知的现象的多个步骤 对于理解一些疾病状态的免疫发病机制变得越来越重要。 为了了解穿孔素对颗粒酶的传递，需要对颗粒酶的分子特性进行研究 最近的步骤--围绕着约束、内部化和早期贩运的事件 颗粒成分。颗粒酶B仍然与颗粒相关的蛋白多糖-丝氨酸结合，在 细胞毒性细胞的分泌物。出乎意料的是，颗粒酶与丝氨酸进行了静电交换 靶向细胞硫酸糖胺多聚糖，然后促进与未定义受体的结合(S)以获得最佳 内部化。此外，如果足够的量，颗粒酶似乎表现出膜溶解活性。 实现了内体浓缩。标题为连体穿孔素-颗粒酶递送假说-两者 穿孔素和颗粒酶预计会从丝氨酸甘氨酸静电交换到细胞表面。 并共同促进内溶递送过程。将先前的努力扩展到糖生物学 颗粒酶B-细胞表面相互作用和强调成像技术，特别是新的方法， 直接可视化颗粒酶和穿孔素与靶细胞的相互作用，具体目的是： 1.颗粒酶B从丝氨酸转移到靶细胞表面的研究 2.确定穿孔素在颗粒酶传递中的作用--分子分析 3.在细胞毒细胞介导的过程中，评估靶细胞的质膜和内体完整性 细胞凋亡

Cytotoxic cell granule-mediated apoptosis is a unique form of cell signaling that entails the intracellular delivery of granule-associated serine proteases (granzymes) by the pore forming protein perforin. The identification of genetic defects in the granule secretion pathway have clearly established that granule- mediated apoptosis is crucial for immune homeostasis. Acquired disruption of cytotoxic cell effector function by viral proteins are likely to trigger lymphoproliferation and autoreactivity as well as diminish host defense responsiveness. A virus-infected or tumor cell may also resist granule mediated apoptosis by inhibiting virtually any aspect of this pathway. Delineating the multiple steps of this poorly understood phenomenon has become increasingly important to comprehend the immunopathogenesis of a number of disease states. To achieve an understanding of granzyme delivery by perforin requires molecular characterization of the most proximal steps - the events that surround the binding, internalization and early trafficking of these granule components. Granzyme B remains bound to the granule-associated proteoglycan, serglycin, after secretion by a cytotoxic cell. Unexpectedly, the granzyme undergoes electrostatic exchange from serglycin to target cell sulfated glycosaminoglycans which then facilitate binding to undefined receptor(s) for optimal internalization. In addition, the granzyme appears to manifest membranolytic activity if a sufficient endosomal concentration is achieved. Entitled the conjoined Perforin-Granzyme delivery hypothesis - both perforin and granzymes are predicted to undergo electrostatic exchange to the cell surface from serglycin and together contribute to endosomolytic delivery process. Extending previous efforts to the glycobiology of granzyme B-cell surface interactions and emphasizing imaging techniques, particularly new methods that directly visualize the interaction of the granzyme and perforin with target cells, the Specific Aims are: 1. Study the transfer of granzyme B from serglycin to the target cell surface 2. Ascertain role of perforin in granzyme delivery - a molecular analysis 3. Assess plasma membrane and endosomal integrity of the target cell during cytotoxic cell-mediated apoptosis