Targeting a Membrane Protease that Controls NK Cell Maturation

靶向控制 NK 细胞成熟的膜蛋白酶

• 批准号: 10506509
• 负责人: Joel L Pomerantz
• 金额: $ 24.56万
• 依托单位: JOHNS HOPKINS UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-06-01 至 2024-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10506509
• 关键词: Alzheimer' s Disease; Antiviral Response; Aspartate; Aspartic Endopeptidases; Binding; Biological; Biological Assay; Cancerous; Cell Lineage; Cell Maturation; Cell physiology; Cells; Cellular Assay; Defect; Development; Disease; Effector Cell; Engineering; Event; Exhibits; Family; Health; Human; ITGAM gene; Immune; Immune system; In Vitro; Knock-in Mouse; Knowledge; Leukocytes; Lipid Bilayers; Lymphocyte; Lymphocyte Function; Lymphoid Cell; Malignant Neoplasms; Membrane; Metalloproteases; Molecular; Molecular Target; Mouse Strains; Mus; Natural Killer Cells; Pathway interactions; Peptide Hydrolases; Process; Protein Analysis; Proteins; Proteolysis; Role; Serine Protease; Site; Stress; Substrate Interaction; T-Lymphocyte; Tumor Cell Line; arm; base; cell behavior; cytotoxic; cytotoxicity; design; immune system function; in vivo; mutant; neoplastic cell; novel; novel therapeutics; pathogen; presenilin; rhomboid; signal peptide peptidase; therapy design; tumor

PROJECT SUMMARY/ABSTRACT A variety of important biological pathways in health and disease are regulated by intramembrane proteolysis, the process that achieves the targeted cleavage of protein substrates within or proximal to the lipid bilayer. Four classes of intramembrane proteases have been described, including the site 2 protease (S2P) metalloprotease family, the rhomboid serine protease family, the Rce1 glutamyl protease family, and the aspartyl protease family that includes presenilins and the related Signal Peptide Peptidase (SPP)-like (SPPL) subfamily. The aspartyl proteases of the SPPL subfamily, including SPP, SPPL2a, SPPL2b, SPPL2c, and SPPL3, are perhaps the least understood subset of intramembrane proteases. Only recently have the biological roles of some of these proteases emerged, and only a handful of relevant biological substrates have been identified. How these intramembrane proteases specifically recognize and cleave their substrates remains mysterious. We discovered a critical role for SPPL3 in NK cell maturation and cytotoxicity. SPPL3 is required in a cell-autonomous manner for the maturation of NK cells from the immature CD27+CD11b- stage to the CD27+CD11b+ and CD27-CD22b+ stages, and for normal NK cell cytotoxicity toward tumor cell targets. Mice engineered to express only SPPL3 D271A in the NK lineage revealed that the proteolytic function of SPPL3 is required in NK cell maturation and function. Like other aspartyl intramembrane proteases of the SPPL subfamily, very little is known about how SPPL3 recognizes and cleaves its substrates. All SPPL proteases possess YD and GXGD motifs that contain the catalytic aspartates, but other regions in the protein that are required for substrate recognition and cleavage have not been identified. Furthermore, although some SPPL3 substrates have been identified, the relevant substrate that must be cleaved by SPPL3 during NK cell maturation is currently unknown. To expand our basic understanding of SPPL3 and related aspartyl intramembrane proteases, and to advance understanding of the key checkpoint in NK cell maturation controlled by SPPL3, we will use newly developed substrate binding and cleavage assays to define SPPL3 determinants required for substrate selection and to identify SPPL3 substrates in NK cells. Our results will expand understanding of intramembrane aspartyl proteases and illuminate molecular events that control NK cell development and function.

项目概要/摘要 健康和疾病中的多种重要生物途径均受到膜内蛋白水解作用的调节， 该过程实现脂质双层内或邻近脂质双层的蛋白质底物的靶向裂解。四 膜内蛋白酶的类别已被描述，包括位点 2 蛋白酶 (S2P) 金属蛋白酶 家族、菱形丝氨酸蛋白酶家族、Rce1 谷氨酰蛋白酶家族和天冬氨酰蛋白酶家族 其中包括早老素和相关的信号肽肽酶 (SPP) 样 (SPPL) 亚家族。天冬氨酰 SPPL 亚家族的蛋白酶，包括 SPP、SPPL2a、SPPL2b、SPPL2c 和 SPPL3，可能是最少的 了解膜内蛋白酶的子集。直到最近才发现其中一些的生物学作用 蛋白酶出现了，但仅鉴定了少数相关的生物底物。这些如何 膜内蛋白酶特异性识别并切割其底物仍然是个谜。我们发现 SPPL3 在 NK 细胞成熟和细胞毒性中发挥关键作用。细胞自主方式需要 SPPL3 用于 NK 细胞从未成熟的 CD27+CD11b- 阶段成熟到 CD27+CD11b+ 和 CD27-CD22b+ 阶段，以及正常 NK 细胞对肿瘤细胞靶标的细胞毒性。小鼠被设计为仅表达 SPPL3 NK 谱系中的 D271A 揭示了 SPPL3 的蛋白水解功能是 NK 细胞成熟和 功能。与 SPPL 亚家族的其他天冬氨酰膜内蛋白酶一样，人们对如何实现知之甚少。 SPPL3 识别并切割其底物。所有 SPPL 蛋白酶都具有 YD 和 GXGD 基序，其中包含 催化天冬氨酸，但蛋白质中底物识别和切割所需的其他区域 尚未被识别。此外，虽然已经鉴定了一些 SPPL3 底物，但相关 NK 细胞成熟过程中必须被 SPPL3 裂解的底物目前尚不清楚。扩大我们的基础 了解 SPPL3 和相关的天冬氨酰膜内蛋白酶，并促进对 SPPL3 控制 NK 细胞成熟的关键检查点，我们将使用新开发的底物结合和 裂解测定以确定底物选择所需的 SPPL3 决定簇并鉴定 SPPL3 底物 在 NK 细胞中。我们的结果将扩大对膜内天冬氨酰蛋白酶的理解并阐明分子 控制 NK 细胞发育和功能的事件。