Department of Pharmacology, Drug Development and Therapeutics, Institute of Biomedicine, University of Turku, Turku, Finland ; Department of Anesthesia/CCM, Stanford University Medical School, Stanford, California, United States of America.
PLoS One. 2013 Oct 2;8(10):e76366. doi: 10.1371/journal.pone.0076366. eCollection 2013.
Receptor expression enhancing proteins (REEPs) were identified by their ability to enhance cell surface expression of a subset of G protein-coupled receptors (GPCRs), specifically GPCRs that have proven difficult to express in heterologous cell systems. Further analysis revealed that they belong to the Yip (Ypt-interacting protein) family and that some REEP subtypes affect ER structure. Yip family comparisons have established other potential roles for REEPs, including regulation of ER-Golgi transport and processing/neuronal localization of cargo proteins. However, these other potential REEP functions and the mechanism by which they selectively enhance GPCR cell surface expression have not been clarified. By utilizing several REEP family members (REEP1, REEP2, and REEP6) and model GPCRs (α2A and α2C adrenergic receptors), we examined REEP regulation of GPCR plasma membrane expression, intracellular processing, and trafficking. Using a combination of immunolocalization and biochemical methods, we demonstrated that this REEP subset is localized primarily to ER, but not plasma membranes. Single cell analysis demonstrated that these REEPs do not specifically enhance surface expression of all GPCRs, but affect ER cargo capacity of specific GPCRs and thus their surface expression. REEP co-expression with α2 adrenergic receptors (ARs) revealed that this REEP subset interacts with and alter glycosidic processing of α2C, but not α2A ARs, demonstrating selective interaction with cargo proteins. Specifically, these REEPs enhanced expression of and interacted with minimally/non-glycosylated forms of α2C ARs. Most importantly, expression of a mutant REEP1 allele (hereditary spastic paraplegia SPG31) lacking the carboxyl terminus led to loss of this interaction. Thus specific REEP isoforms have additional intracellular functions besides altering ER structure, such as enhancing ER cargo capacity, regulating ER-Golgi processing, and interacting with select cargo proteins. Therefore, some REEPs can be further described as ER membrane shaping adapter proteins.
受体表达增强蛋白（REEPs）是通过其增强一组 G 蛋白偶联受体（GPCR）细胞表面表达的能力来鉴定的，特别是那些在异源细胞系统中难以表达的 GPCR。进一步的分析表明，它们属于 Yip（Ypt-interacting protein）家族，并且一些 REEP 亚型会影响内质网结构。Yip 家族的比较已经确定了 REEPs 的其他潜在作用，包括调节内质网-高尔基体运输和货物蛋白的加工/神经元定位。然而，这些其他潜在的 REEP 功能以及它们选择性增强 GPCR 细胞表面表达的机制尚未阐明。通过利用几种 REEP 家族成员（REEP1、REEP2 和 REEP6）和模型 GPCR（α2A 和 α2C 肾上腺素能受体），我们研究了 REEP 对 GPCR 质膜表达、细胞内加工和运输的调节。我们使用免疫定位和生化方法相结合，证明了这组 REEP 主要定位于内质网，而不是质膜。单细胞分析表明，这些 REEPs 并非特异性地增强所有 GPCR 的表面表达，而是影响特定 GPCR 的内质网货物容量，从而影响其表面表达。REEP 与 α2 肾上腺素能受体（AR）的共表达表明，这组 REEP 与 α2C 相互作用并改变其糖基化加工，但不影响 α2A AR，表明与货物蛋白选择性相互作用。具体来说，这些 REEPs 增强了α2C AR 的最小/非糖基化形式的表达并与之相互作用。最重要的是，表达缺乏羧基末端的突变 REEP1 等位基因（遗传性痉挛性截瘫 SPG31）导致这种相互作用丧失。因此，除了改变内质网结构之外，特定的 REEP 同工型还具有其他细胞内功能，例如增强内质网货物容量、调节内质网-高尔基体加工以及与选择性货物蛋白相互作用。因此，一些 REEPs 可以进一步描述为内质网膜成型衔接蛋白。
