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.

项目总结/摘要 粘附G蛋白偶联受体（aGPCR）是一组33个特征不明显的非嗅觉GPCR 具有独特的特征，包括自催化加工和大的细胞外区域。aGPCR涉及 在无数的生物过程中，其中一些与疾病有关，特别是广泛的 癌的许多aGPCR的细胞外区域含有已知参与细胞凋亡的结构域。 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是孤儿受体，没有已知的配体。 这些aGPCR的配体的鉴定应该有助于理解它们的生理功能和作用， 癌症发展鉴定参与配体-受体相互作用的配体的一个主要挑战是， 细胞-细胞界面的缺点是它们倾向于非常弱地结合它们的受体，并且目前可用的方法不 要有足够的敏感度。为了克服这一点，我们建议采用一种新的和高度敏感的细胞为基础的 方法鉴定17个未充分研究的孤儿aGPCR的蛋白质配体。该方法的可行性 已经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来筛选跨膜蛋白的集合，以鉴定配体候选物， aGPCR。在这项研究中确定的配体候选者将为更深入地表征 它们与各自的aGPCR的结合以及在未来研究中结合的功能后果。