Granzyme a Pathway of CTL-Mediated Cell Death

颗粒酶是 CTL 介导的细胞死亡的途径

• 批准号: 7080452
• 负责人: Judy Lieberman
• 金额: $ 54.3万
• 依托单位: IMMUNE DISEASE INSTITUTE, INC.
• 依托单位国家: 美国
• 财政年份: 1999
• 资助国家: 美国
• 起止时间: 1999-07-01 至 2009-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7080452
• 关键词: DNA damage; apoptosis; cell mediated cytotoxicity; cysteine endopeptidases; cytolysins; cytotoxic T lymphocyte; deoxyribonuclease I; enzyme mechanism; expression cloning; genetically modified animals; laboratory mouse; mitochondria; molecular cloning; natural killer cells; pore forming protein; serine proteinases; stress proteins

DESCRIPTION (provided by applicant): Cytotoxic T lymphocytes (CTL) and natural killer (NK) cells exocytose cytotoxic granules to induce apoptosis of virally infected or tumor targets. Cytotoxic granules contain perforin, a pore-forming protein, and a group of serine proteases, called granzymes (Gzm), in a proteoglycan matrix. Perforin and either GzmA or GzmB is sufficient for granule-mediated lysis by CTL, but mice deficient in either Gzm may be impaired in defense against some viral infections. GzmB activates a ubiquitous apoptotic cascade induced by cleavage of caspases and can also directly cleave some key caspase pathway substrates. However, CTL granule mediated cytolysis is virtually unimpaired in the setting of complete caspase blockade. GzmA, the most abundant Gzm, activates a novel caspase-independent apoptotic pathway. The goal of this proposal is to continue to unravel the mechanism of action of GzmA, building on the research accomplished during the previous funding. GzmA, delivered into target cells via perforin, induces single-stranded DNA damage as Nell as apoptotic morphology and loss of cell membrane integrity. A special target of the GzmA cell death pathway is a 270-440 kDa endoplasmic reticulum-associated complex, called the SET complex, which contains three GzmA substrates, the nucleosome assembly protein SET, the DNA bending protein HMG-2 and the apurinic endonuclease (Ape1) in base excision repair (BER). GzmA cleavage of Ape1 makes target cell repair of GzmA-induced damage unlikely. The SET complex also contains the GzmA-activated DNase, NM23-H1, which is activated when GzmA cleaves its specific inhibitor, the SET protein. The competing renewal of this grant will investigate the kinetics of GzmA-mediated damage within target cells; determine the mitochondrial effects of GzmA; identify the remaining components of the SET complex and seek to understand further the role of the SET complex in normal cellular function and in GzmA-mediated cell death; and investigate the effect of GzmA on other DNA repair pathways, besides BER.

描述(由申请人提供)：细胞毒性T淋巴细胞(CTL)和自然杀伤(NK)细胞分泌细胞毒颗粒，以诱导病毒感染或肿瘤靶点的细胞凋亡。细胞毒性颗粒含有穿孔素，一种造孔蛋白，以及蛋白多糖基质中的一组丝氨酸蛋白酶，称为颗粒酶(GZM)。穿孔素和GzmA或GzmB对CTL的颗粒介导的裂解是足够的，但缺乏Gzm的小鼠对某些病毒感染的防御可能会受到损害。GzmB可激活caspase裂解所诱导的普遍存在的细胞凋亡级联反应，也可直接裂解一些关键的caspase途径底物。然而，在caspase被完全阻断的情况下，CTL颗粒介导的细胞溶解几乎没有受到损害。GzmA是GZM中含量最丰富的一种，它激活了一条新的caspase非依赖的凋亡途径。这项提案的目标是在先前资助期间完成的研究的基础上，继续解开GZMA的行动机制。GzmA通过穿孔素传递到靶细胞，诱导单链DNA损伤和Nell一样的凋亡形态和细胞膜完整性的丧失。GzmA细胞死亡的特殊靶点 途径是一个270-440 kDa的内质网相关复合体，称为SET复合体，它包含三个GzmA底物，核小体组装蛋白SET，DNA弯曲蛋白HMG-2和碱基切除修复(BER)中的无嘌呤核酸内切酶(APE1)。GzmA裂解APE1使靶细胞不可能修复GzmA诱导的损伤。SET复合体还包含GzmA激活的DNA酶NM23-H1，当GzmA切割其特定的抑制物SET蛋白时，该酶被激活。这项资助的竞争性更新将研究GzmA介导的靶细胞内损伤的动力学；确定GzmA对线粒体的影响；确定SET复合体的剩余成分并寻求进一步了解SET复合体在正常细胞功能和GzmA介导的细胞死亡中的作用；以及研究GzmA对除BER之外的其他DNA修复途径的影响。