Endocannabinoid signaling plays a regulatory role in various (neuro)biological functions. 2-arachidonoylglycerol (2-AG) is the most abundant endocannabinoid, and although its canonical biosynthetic pathway involving phosphoinositide-specific phospholipase C and diacylglycerol lipase α is known, alternative pathways remain unsettled. Here, we characterize a noncanonical pathway implicating glycerophosphodiesterase 3 (GDE3, from GDPD2 gene). Human GDE3 expressed in HEK293T cell membranes catalyzed the conversion of lysophosphatidylinositol (LPI) into monoacylglycerol and inositol-1-phosphate. The enzyme was equally active against 1-acyl and 2-acyl LPI. When using 2-acyl LPI, where arachidonic acid is the predominant fatty acid, LC-MS analysis identified 2-AG as the main product of LPI hydrolysis by GDE3. Furthermore, inositol-1-phosphate release into the medium occurred upon addition of LPI to intact cells, suggesting that GDE3 is actually an ecto-lysophospholipase C. In cells expressing G-protein–coupled receptor GPR55, GDE3 abolished 1-acyl LPI–induced signaling. In contrast, upon simultaneous ex-pression of GDE3 and cannabinoid receptor CB2, 2-acyl LPI evoked the same signal as that induced by 2-AG. These data strongly suggest that, in addition to degrading the GPR55 LPI ligand, GDE3 can act as a switch between GPR55 and CB2 signaling. Coincident with a major expression of both GDE3 and CB2 in the spleen, spleens from transgenic mice lacking GDE3 displayed doubling of LPI content compared with WT mice. Decreased production of 2-AG in whole spleen was also observed, supporting the in vivo relevance of our findings. These data thus open a new research avenue in the field of endocannabinoid generation and reinforce the view of GPR55 and LPI being genuine actors of the endocannabinoid system.

中文翻译：

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甘油磷酸二酯酶 3 (GDE3) 是一种溶血磷脂酰肌醇特异性外磷脂酶 C，充当内源性大麻素信号转导开关。

内源性大麻素信号在各种（神经）生物学功能中发挥调节作用。2-花生四烯酸甘油 (2-AG) 是最丰富的内源性大麻素，虽然其典型的生物合成途径涉及磷酸肌醇特异性磷脂酶 C 和二酰基甘油脂肪酶 α 是已知的，但替代途径仍未解决。在这里，我们表征了涉及甘油磷酸二酯酶 3（GDE3，来自 GDPD2 基因）的非经典途径。在 HEK293T 细胞膜中表达的人类 GDE3 催化溶血磷脂酰肌醇 (LPI) 转化为单酰基甘油和 1-磷酸肌醇。该酶对 1-酰基和 2-酰基 LPI 的活性相同。当使用 2-酰基 LPI 时，其中花生四烯酸是主要脂肪酸，LC-MS 分析确定 2-AG 是 GDE3 水解 LPI 的主要产物。此外，向完整细胞中加入 LPI 后，1-磷酸肌醇释放到培养基中，表明 GDE3 实际上是一种外溶血磷脂酶 C。在表达 G 蛋白偶联受体 GPR55 的细胞中，GDE3 消除了 1-酰基 LPI 诱导的信号传导。相反，在同时表达 GDE3 和大麻素受体 CB2 时，2-酰基 LPI 诱发了与 2-AG 诱导的信号相同的信号。这些数据强烈表明，除了降解 GPR55 LPI 配体外，GDE3 还可以作为 GPR55 和 CB2 信号之间的开关。与脾脏中 GDE3 和 CB2 的主要表达相一致，与 WT 小鼠相比，来自缺乏 GDE3 的转基因小鼠的脾脏的 LPI 含量增加了一倍。还观察到整个脾脏中 2-AG 的产生减少，支持我们发现的体内相关性。