Wang Ju-Qiang, Kon Junko, Mogi Chihiro, Tobo Masayuki, Damirin Alatangaole, Sato Koichi, Komachi Mayumi, Malchinkhuu Enkhzol, Murata Naoya, Kimura Takao, Kuwabara Atsushi, Wakamatsu Kaori, Koizumi Hideki, Uede Toshimitsu, Tsujimoto Gozoh, Kurose Hitoshi, Sato Takashi, Harada Akihiro, Misawa Norihiko, Tomura Hideaki, Okajima Fumikazu
Laboratory of Signal Transduction and Department of Cell Biology, Institute for Molecular and Cellular Regulation, Gunma University, Maebashi 371-8512, Japan.
J Biol Chem. 2004 Oct 29;279(44):45626-33. doi: 10.1074/jbc.M406966200. Epub 2004 Aug 23.
T cell death-associated gene 8 (TDAG8) has been reported to be a receptor for psychosine. Ovarian cancer G-protein-coupled receptor 1 (OGR1) and GPR4, G-protein-coupled receptors (GPCRs) closely related to TDAG8, however, have recently been identified as proton-sensing or extracellular pH-responsive GPCRs that stimulate inositol phosphate and cAMP production, respectively. In the present study, we examined whether TDAG8 senses extracellular pH change. In the several cell types that were transfected with TDAG8 cDNA, cAMP was markedly accumulated in response to neutral to acidic extracellular pH, with a peak response at approximately pH 7.0-6.5. The pH effect was inhibited by copper ions and was reduced or lost in cells expressing mutated TDAG8 in which histidine residues were changed to phenylalanine. In the membrane fractions prepared from TDAG8-transfected cells, guanosine 5'-O-(3-thiotriphosphate) binding activity and adenylyl cyclase activity were remarkably stimulated in response to neutral and acidic pH. The concentration-dependent effect of extracellular protons on cAMP accumulation was shifted to the right in the presence of psychosine. The inhibitory psychosine effect was also observed for pH-dependent actions in OGR1- and GPR4-expressing cells but not for prostaglandin E(2)- and sphingosine 1-phosphate-induced actions in any pH in native and sphingosine 1-phosphate receptor-expressing cells. Glucosylsphingosine and sphingosylphosphorylcholine similarly inhibited the pH-dependent action, although to a lesser extent. Psychosine-sensitive and pH-dependent cAMP accumulation was also observed in mouse thymocytes. We concluded that TDAG8 is one of the proton-sensing GPCRs coupling to adenylyl cyclase and psychosine, and its related lysosphingolipids behave as if they were antagonists against protein-sensing receptors, including TDAG8, GPR4, and OGR1.
据报道，T细胞死亡相关基因8（TDAG8）是Psychosine的受体。然而，与TDAG8密切相关的G蛋白偶联受体卵巢癌G蛋白偶联受体1（OGR1）和GPR4，最近被鉴定为质子感应或细胞外pH响应性G蛋白偶联受体，分别刺激肌醇磷酸和cAMP的产生。在本研究中，我们检测了TDAG8是否能感知细胞外pH变化。在转染了TDAG8 cDNA的几种细胞类型中，cAMP响应细胞外pH从中性到酸性的变化而显著积累，在约pH 7.0 - 6.5时达到峰值响应。pH效应被铜离子抑制，并且在表达组氨酸残基被苯丙氨酸取代的突变TDAG8的细胞中降低或消失。在从转染了TDAG8的细胞制备的膜组分中，鸟苷5'-O-(3-硫代三磷酸)结合活性和腺苷酸环化酶活性响应中性和酸性pH而显著受到刺激。在存在Psychosine的情况下，细胞外质子对cAMP积累的浓度依赖性效应向右移动。在表达OGR1和GPR4的细胞中，对pH依赖性作用也观察到了Psychosine的抑制作用，但在天然细胞和表达鞘氨醇-1-磷酸受体的细胞中，在任何pH下对前列腺素E(2)和鞘氨醇-1-磷酸诱导的作用均未观察到这种抑制作用。葡萄糖基鞘氨醇和鞘氨醇磷酸胆碱同样抑制pH依赖性作用，尽管程度较小。在小鼠胸腺细胞中也观察到了Psychosine敏感和pH依赖性的cAMP积累。我们得出结论，TDAG8是与腺苷酸环化酶和Psychosine偶联的质子感应G蛋白偶联受体之一，其相关的溶血鞘脂类物质表现得就像是针对包括TDAG8、GPR4和OGR1在内的蛋白质感应受体的拮抗剂。
