MECHANISMS OF ACTIVATION OF ICE-LIKE CYSTEINE PROTEASES

描述(申请人提供)：caspase-1是抵抗病原体感染和细胞应激的先天免疫反应的重要组成部分。Caspase-1的激活可以导致非活性的前IL-12和前IL-18的处理，分别产生活性的细胞因子IL-12和IL-18，它们是刺激发热、免疫细胞的募集和激活、次级细胞因子的产生和细胞增殖的强有力的炎症介质。Caspase-1的激活也会导致一种称为下垂的炎性细胞死亡。申请人的实验室最近证明，这种形式的细胞死亡是由caspase-1接头蛋白ASC介导的，该蛋白组装了一种名为焦胞体的超分子结构，在下垂期间激活caspase-1，以响应不同的促炎和应激信号。他们还阐明了一种在家族性地中海热中突变的蛋白质PYRIN在人类自发性炎症性Papa综合征的caspase-1激活和炎症中的作用，并证明PYRIN是细胞骨架组织蛋白PSTPIP1的同源三聚体胞浆受体。PSTPIP1的连接通过揭开其PYD结构域而激活吡喃，然后与ASC相互作用，促进ASC齐聚成活性的焦小体。在这一应用中，提出了特定的目标，以扩展这些正在进行的研究，以进一步研究PSTPIP1和吡啉相互作用的调节和分子决定因素。特别是，目标将集中在表征蛋白酪氨酸磷酸酶PTP-PEST和酪氨酸激酶c-Abl的确切作用，这两个蛋白都已被证明与PSTPIP1相互作用，在吡喃介导的caspase-1激活途径中。此外，还建议鉴定新的ASC裂解体上游激活物，并进一步研究TLR2、TLR4、低温蛋白和caspase-1介导的蛋白分解在巨噬细胞裂解症中的作用。最后，建议进行研究，以探索将体外ASC嗜热体组装作为高通量筛选的潜在用途，以确定抑制和治疗炎症的小分子治疗药物。公共卫生相关性：炎症是对感染和细胞应激的先天免疫反应中的一个关键过程。这项研究的重点是了解关键细胞蛋白在复杂的分子和细胞炎症机制中的作用。这项研究的结果将有助于设计和发现有效的疗法来治疗与衰老、自身炎症性疾病和慢性炎症性疾病相关的炎症。

DESCRIPTION (provided by applicant): Caspase-1 is an important component of the innate immune response against pathogenic infection and cellular stresses. Activation of caspase-1 can lead to processing of the inactive pro-IL-12 and pro-IL-18 to produce the active cytokines IL-12 and IL-18, respectively, which are potent mediators of inflammation that stimulate fever, recruitment and activation of immune cells, production of secondary cytokines and cellular proliferation. Caspase-1 activation can also lead to an inflammatory form of cell death called pyroptosis. The applicant's laboratory demonstrated recently that this form of cell death is mediated by the caspase-1 adaptor protein ASC, which assembles a supramolecular structure termed the pyroptosome that activates caspase-1 during pyroptosis in response to diverse pro-inflammatory and stress signals. They have also elucidated the role of pyrin, a protein mutated in familial Mediterranean fever, in caspase-1 activation and inflammation in the human auto-inflammatory PAPA syndrome, and demonstrated that pyrin is a homotrimeric cytosolic receptor for the cytoskeleton- organizing protein PSTPIP1. Ligation by PSTPIP1 activates pyrin by unmasking of its PYD domain, which then interacts with ASC and facilitates ASC oligomerization into the active pyroptosome. In this application specific aims are proposed to extend these ongoing studies to further investigate the regulation and molecular determinants of the interaction between PSTPIP1 and pyrin. In particular, the aims will focus on characterizing the exact role of the protein-tyrosine phosphatase PTP-PEST and tyrosine kinase c-Abl, both of which have been shown to interact with PSTPIP1, in the pyrin-mediated caspase-1 activation pathway. It is also proposed to characterize novel upstream activators of the ASC pyroptosome and further investigate the role TLR2, TLR4, cryopyrin and caspase-1-mediated proteolysis in macrophage pyroptosis. Finally, studies are proposed to investigate the potential use of the in vitro ASC pyroptosome assembly as a high throughput screen to identify small molecule therapeutics to inhibit and treat inflammation. PUBLIC HEALTH RELEVANCE: Inflammation is a key process in the innate immune response to infections and cellular stress. This research focuses on understanding the role of key cellular proteins in the complex molecular and cellular inflammatory mechanisms. The results of this research will help in the design and discovery of effective therapeutics to treat inflammation associated with aging, auto-inflammatory disease and chronic inflammatory diseases.