Research Centre for Translational Medicine at East Hospital, School of Life Sciences and Technology, Tongji University, Shanghai, China.
Research Centre for Translational Medicine at East Hospital, School of Life Sciences and Technology, Tongji University, Shanghai, China.
Arch Biochem Biophys. 2018 May 15;646:24-30. doi: 10.1016/j.abb.2018.03.031. Epub 2018 Mar 27.
Relaxin family peptide receptor 3 (RXFP3) is implicated in the regulation of food intake and stress response upon activation by its cognate agonist relaxin-3. As an A-class G protein-coupled receptor, RXFP3 is an integral plasma membrane protein with seven transmembrane domains, yet influence of the membrane lipids on its function remains unknown. In the present study, we disclosed that cholesterol, an essential membrane lipid for mammalian cells, modulated the binding properties of human RXFP3 with its ligands. We first demonstrated that depletion of cholesterol from host human embryonic kidney (HEK) 293T cells by methyl-β-cyclodextrin altered ligand-binding properties of the overexpressed human RXFP3, such as increasing its binding potency with some antagonists and decreasing its binding affinity with a NanoLuc-conjugated R3/I5 tracer. Thereafter, we demonstrated that two B-chain residues, B5Tyr and B12Arg, were primarily responsible for the increased binding potency of these antagonists with human RXFP3 under the cholesterol depletion condition. Our results suggest that cell membrane cholesterol interacts with human RXFP3 and modulates its ligand-binding properties, providing new insights into the influence of membrane lipids on RXFP3 function.
松弛素家族肽受体 3（RXFP3）在被其同源激动剂松弛素-3 激活后，参与调节食物摄入和应激反应。作为 A 类 G 蛋白偶联受体，RXFP3 是一种完整的质膜蛋白，具有七个跨膜结构域，但膜脂对其功能的影响尚不清楚。在本研究中，我们揭示了胆固醇作为哺乳动物细胞的必需膜脂，调节了人 RXFP3 与其配体的结合特性。我们首先证明，用甲基-β-环糊精从宿主人胚肾 293T 细胞中耗尽胆固醇会改变过表达的人 RXFP3 的配体结合特性，例如增加其与一些拮抗剂的结合效力，并降低其与 NanoLuc 缀合的 R3/I5 示踪剂的结合亲和力。此后，我们证明了两个 B 链残基，B5Tyr 和 B12Arg，在胆固醇耗尽条件下，这些拮抗剂与人类 RXFP3 的结合效力增加主要归因于这两个 B 链残基。我们的研究结果表明，细胞膜胆固醇与人 RXFP3 相互作用并调节其配体结合特性，为膜脂对 RXFP3 功能的影响提供了新的见解。
