Identify cell surface protein ligand candidates for understudied adhesion GPCRs using a sensitive cell-based approach

使用敏感的基于细胞的方法识别用于正在研究的粘附 GPCR 的细胞表面蛋白配体候选物

• 批准号: 10452320
• 负责人: Zhengyu Ma
• 金额: $ 14.2万
• 依托单位: ALFRED I. DU PONT HOSP FOR CHILDREN
• 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/10452320
• 关键词: Adhesions; Affinity; Antibodies; Antigen-Presenting Cells; Back; Binding; Biological Assay; Biological Process; C-terminal; CD3 Antigens; Cell Line; Cell Surface Proteins; Cell membrane; Cell surface; Cells; Characteristics; Collection; Custom; Data; Development; Disease; Drug Design; Enzyme-Linked Immunosorbent Assay; Flow Cytometry; Future; G Protein-Coupled Receptor Signaling; G-Protein-Coupled Receptors; Genes; Human; Individual; Integral Membrane Protein; Interleukin-10; Interleukin-4; Lead; Ligand Binding; Ligands; Malignant Neoplasms; Measures; Mediating; Methods; Microscope; Modeling; Monitor; Movement; Mus; Musculoskeletal Diseases; N-terminal; Nature; Noise; Open Reading Frames; Orphan; Pattern; Physiological; Play; Production; Protein Array; Proteins; Puromycin; Receptor Cell; Receptor Signaling; Retroviral Vector; Role; Sensitivity and Specificity; Signal Transduction; Stains; Surface; T-Cell Receptor; T-Lymphocyte; Tertiary Protein Structure; Testing; Th2 Cells; adhesion receptor; base; cDNA Library; cytokine; extracellular; human disease; novel; protein protein interaction; receptor; receptor binding; stable cell line; therapeutic target; two-dimensional

PROJECT SUMMARY/ABSTRACT Adhesion G protein-coupled receptors (aGPCRs) are a group of 33 poorly characterized non-olfactory GPCRs with distinct features including autocatalytic processing and large extracellular regions. aGPCRs are involved in a myriad of biological processes and some of them are associated with diseases, especially a wide range of cancers. The extracellular regions of many of the aGPCRs contain domains that are known to be involved in protein-protein interactions, suggesting that binding to cell surface protein ligands may play an important role in their mechanisms of action. The majority of aGPCRs, however, are orphan receptors without known ligands. Identification of ligands for these aGPCRs should help understand their physiological functions and roles in cancer development. A major challenge for identifying ligands involved in ligand-receptor interactions at the cell-cell interface is that they tend to bind their receptors very weakly and currently available methods do not have enough sensitivity. To overcome this, we propose to employ a novel and highly sensitive cell-based approach to identify protein ligands for 17 understudied orphan aGPCRs. The feasibility of the approach has been tested using model ligands and receptors that are known bind to each other with low affinities. We will use the cell-based approach to screen a collection of transmembrane proteins to identify ligand candidates for the aGPCRs. Ligand candidates identified in this study will pave the way for more in-depth characterization of their bindings to respective aGPCRs and the functional consequences of the bindings in future studies.

项目概要/摘要 粘附 G 蛋白偶联受体 (aGPCR) 是一组 33 个特征不明确的非嗅觉 GPCR 具有独特的特征，包括自催化处理和大的细胞外区域。 aGPCR 参与其中 参与无数的生物过程，其中一些与疾病相关，尤其是多种 癌症。许多 aGPCR 的胞外区域包含已知参与以下过程的结构域： 蛋白质-蛋白质相互作用，表明与细胞表面蛋白质配体的结合可能在 他们的行动机制。然而，大多数 aGPCR 是没有已知配体的孤儿受体。 这些 aGPCR 配体的鉴定应有助于了解它们的生理功能和作用 癌症的发展。识别参与配体-受体相互作用的配体的一个主要挑战 细胞-细胞界面的一个特点是它们往往与受体的结合非常弱，而目前可用的方法不能 有足够的灵敏度。为了克服这个问题，我们建议采用一种新颖且高度敏感的基于细胞的 方法鉴定 17 个待研究的孤儿 aGPCR 的蛋白质配体。该方法的可行性已 使用已知以低亲和力彼此结合的模型配体和受体进行了测试。我们将 使用基于细胞的方法筛选跨膜蛋白集合，以确定候选配体 aGPCR。本研究中确定的候选配体将为更深入的表征铺平道路 它们与各自 aGPCR 的结合以及未来研究中结合的功能后果。