Non-proteasomal protease(s) that define C-termini of MHC class I epitopes

描述（由申请方提供）：细胞毒性CD 8 + T细胞通过杀死病毒感染或恶性改变的细胞在宿主防御中发挥关键作用。它们通过结合在MHC I类分子背景下呈递的特异性肽（表位）来执行该功能。这些表位必须满足严格的长度（8-10个氨基酸）和序列要求，以便成功地结合MHC I类。许多必要的加工活性归因于26 S蛋白酶体，一种位于细胞质中的酶复合物。特别是肽的C-末端修剪通常被认为是蛋白酶体的专有结构域。然而，我们的初步研究表明，在蛋白酶体抑制剂的存在下，某些肽的C-末端延伸可以在胞质溶胶中被修剪，产生活性表位。这表明胞质溶胶中的另一种酶活性有助于C-末端肽加工。这种可能性开启了对关键免疫途径的全新理解。我们的长期目标是更好地了解这种替代途径，并最终将这些新知识应用于疫苗开发和肿瘤免疫学等领域。当前的目标是确定这种活性的来源，并确认其在抗原加工中的作用。这将通过由体外呈递测定、最先进的蛋白质鉴定技术以及最终的siRNA敲低实验指导的常规蛋白质纯化技术来实现。免疫系统的细胞毒性T细胞在消除感染和攻击癌细胞方面发挥关键作用。他们通过展示在其“目标”表面的肽来识别他们要杀死的细胞。这里提出的研究重点是如何在细胞内产生肽。我们越了解肽的产生（也称为“抗原加工”），我们就越能控制免疫系统这种重要细胞类型的招募。

DESCRIPTION (provided by applicant): Cytotoxic CD8+ T-cells play a key role in host defense by killing virally-infected or malignancy-altered cells. They perform this function by binding to specific peptides (epitopes) presented in the context of MHC class I molecules. These epitopes must meet stringent length (8-10 amino acids) and sequence requirements in order to successfully bind MHC class I. Much of the necessary processing activity has been attributed to the 26S proteasome, an enzymatic complex located in the cytoplasm. C-terminal trimming of peptides, in particular, is generally accepted to be exclusively the domain of the proteasome. However, our preliminary studies demonstrate that C-terminal extensions of certain peptides can be trimmed in the cytosol, generating the active epitope, in the presence of proteasome inhibitor. This indicates that another enzymatic activity in the cytosol contributes to C-terminal peptide processing. This possibility opens up a whole new understanding of a critical immunologic pathway. Our long term goal is to gain a better understanding of this alternative pathway and, ultimately, to apply this new knowledge to areas such as vaccine development and tumor immunology. The immediate goal is to identify the source of this activity and confirm its role in antigen processing. This will be accomplished through conventional protein purification techniques guided by in vitro presentation assays, state of the art protein identification techniques and, ultimately, siRNA knockdown experiments. NARRATIVE: Cytotoxic T cells of the immune system can play a critical role in eliminating infections and in attacking cancer cells. They identify the cells that they are to kill by the peptides that are displayed at the surfaces of their "targets". The research proposed here focuses on how the peptides are generated within the cell. The better we understand peptide production (also termed "antigen processing"), the more control we will have over the recruitment of this important cell type of the immune system.