Cell surface receptors are critical for cell signaling and constitute a quarter of all human genes. Despite their importance and abundance, receptor interaction networks remain understudied because of difficulties associated with maintaining membrane proteins in their native conformation and their typically weak interactions. To overcome these challenges, we developed an extracellular vesicle-based method for membrane protein display that enables purification-free and high-throughput detection of receptor–ligand interactions in membranes. We demonstrate that this platform is broadly applicable to a variety of membrane proteins, enabling enhanced detection of extracellular interactions over a wide range of binding affinities. We were able to recapitulate and expand the interactome for prominent members of the B7 family of immunoregulatory proteins such as PD-L1/CD274 and B7-H3/CD276. Moreover, when applied to the orphan cancer-associated fibroblast protein, LRRC15, we identified a membrane-dependent interaction with the tumor stroma marker TEM1/CD248. Furthermore, this platform enabled profiling of cellular receptors for target-expressing as well as endogenous extracellular vesicles. Overall, this study presents a sensitive and easy to use screening platform that bypasses membrane protein purification and enables characterization of interactomes for any cell surface–expressed target of interest in its native state.

细胞表面受体对细胞信号传导至关重要，占所有人类基因的四分之一。尽管它们的重要性和丰富性，受体相互作用网络仍然没有得到充分研究，因为将膜蛋白维持在其天然构象中存在困难，而且它们的相互作用通常很弱。为了克服这些挑战，我们开发了一种基于细胞外囊泡的膜蛋白展示方法，该方法能够对膜中受体-配体相互作用进行免纯化和高通量检测。我们证明该平台广泛适用于各种膜蛋白，从而能够在广泛的结合亲和力范围内增强细胞外相互作用的检测。我们能够概括和扩展 B7 免疫调节蛋白家族的重要成员（如 PD-L1/CD274 和 B7-H3/CD276）的相互作用组。此外，当应用于孤儿癌症相关成纤维细胞蛋白 LRRC15 时，我们发现了与肿瘤基质标记物 TEM1/CD248 的膜依赖性相互作用。此外，该平台能够分析细胞受体以表达目标以及内源性细胞外囊泡。总体而言，这项研究提供了一种灵敏且易于使用的筛选平台，该平台绕过了膜蛋白纯化，并能够表征任何细胞表面表达的天然状态目标的相互作用组。我们确定了与肿瘤基质标志物 TEM1/CD248 的膜依赖性相互作用。此外，该平台能够分析用于表达靶标的细胞受体以及内源性细胞外囊泡。总体而言，这项研究提供了一种灵敏且易于使用的筛选平台，该平台绕过了膜蛋白纯化，并能够表征任何细胞表面表达的天然状态目标的相互作用组。我们确定了与肿瘤基质标志物 TEM1/CD248 的膜依赖性相互作用。此外，该平台能够分析用于表达靶标的细胞受体以及内源性细胞外囊泡。总体而言，这项研究提供了一种灵敏且易于使用的筛选平台，该平台绕过了膜蛋白纯化，并能够表征任何细胞表面表达的天然状态目标的相互作用组。