Members of RTP and REEP gene families influence functional bitter taste receptor expression
Members of RTP and REEP gene families influence functional bitter taste receptor expression
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DOI：
10.1074/jbc.m513637200
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发表时间：
2006-07-21
影响因子：
4.8
通讯作者：
Meyerhof, Wolfgang
中科院分区：
文献类型：
--
作者：
Behrens, Maik;Bartelt, Juliane;Meyerhof, Wolfgang
化学感觉受体的功能鉴定通常是通过在哺乳动物细胞系中的异源表达来实现的。然而，许多化学感受器基因，包括苦味受体(TAS2Rs)，只显示边缘细胞表面的表达。通常，这些问题可以通过使用由“出口标签”和受体序列本身组成的嵌合受体来规避。似乎化学感受器细胞在体内表达针对天然受体分子的细胞表面靶向因子。然而，对于TAS2R来说，这些因素仍然未知。本研究探讨RTP和REEP蛋白对人TAS2Rs在异种细胞中功能表达的影响。我们在HEK 293T细胞中表达了hTAS2Rs，并观察到对激动剂刺激的反应性有显著差异。免疫细胞化学显示苦味β-吡喃葡萄糖苷受体hTAS2R16在高尔基体内积聚。RTP和REEP蛋白的共表达改变了一些hTAS2R对激动剂刺激的反应，这可能是由于细胞表面生物素化实验证明的有效的细胞表面定位。HTAS2R16与RTP3或RTP4的免疫共沉淀表明，这些共因子影响hTAS2R16功能的机制可能涉及蛋白质-蛋白质的直接相互作用。最后，表达分析表明RTP和REEP基因在人的环状乳头和睾丸中表达，这两个部位都是TAS2R基因表达的部位。
Functional characterization of chemosensory receptors is usually achieved by heterologous expression in mammalian cell lines. However, many chemoreceptor genes, including bitter taste receptors (TAS2Rs), show only marginal cell surface expression. Usually, these problems are circumvented by using chimeric receptors consisting of "export tags" and the receptor sequence itself. It seems likely that chemoreceptor cells express factors for cell surface targeting of native receptor molecules in vivo. For TAS2Rs, however, such factors are still unknown. The present study investigates the influence of RTP and REEP proteins on the functional expression of human TAS2Rs in heterologous cells. We expressed hTAS2Rs in HEK 293T cells and observed dramatic differences in responsiveness to agonist stimulation. By immunocytochemistry we show accumulation of the bitter beta-glucopyranoside receptor hTAS2R16 in the Golgi compartment. Coexpression of RTP and REEP proteins changed the responses of some hTAS2Rs upon agonist stimulation, which is likely due to efficient cell surface localization as demonstrated by cell surface biotinylation experiments. The coimmunoprecipitation of hTAS2R16 and RTP3 or RTP4 suggests that the mechanism by which these cofactors influence hTAS2R16 function might involve direct protein-protein interaction. Finally, expression analyses demonstrate RTP and REEP gene expression in human circumvallate papillae and testis, both of which are sites of TAS2R gene expression.