Alexandre David, Vaudry Hubert, Grumolato Luca, Turquier Valérie, Fournier Alain, Jégou Sylvie, Anouar Youssef
European Institute for Peptide Research (IFRMP 23), Laboratory of Cellular and Molecular Neuroendocrinology, Institut National de la Santé et de la Recherche Médicale, University of Rouen, 76821 Mont Saint Aignan, France.
Endocrinology. 2002 Jul;143(7):2680-92. doi: 10.1210/endo.143.7.8880.
Pituitary adenylate cyclase-activating polypeptide (PACAP) exerts its various effects through activation of two types of G protein-coupled receptors, a receptor with high affinity for PACAP named PAC1-R and two receptors exhibiting similar affinity for both PACAP and vasoactive intestinal polypeptide named VPAC1-R and VPAC2-R. Here, we report the characterization of PAC1-R and novel splice variants in the frog Rana ridibunda. The frog PAC1-R has 78% homology with human PAC1-R and is highly expressed in the central nervous system. Two splice variants of the frog receptor that display additional amino acid cassettes in the third intracellular loop were characterized. PAC1-R25 carries a 25-amino acid insertion that matches the hop cassette of the mammalian receptor, whereas PAC1-R41 carries a cassette with no homology to any mammalian PAC1-R variant. A third splice variant of PAC1-R, exhibiting a completely different intracellular C-terminal domain, named PAC1-Rmc has also been identified. Determination of cAMP formation in cells transfected with the cloned receptors showed that PACAP activated PAC1-R, PAC1-R25, and PAC1-R41 with similar potency. In contrast, PACAP failed to stimulate adenylate cyclase in cells transfected with PAC1-Rmc. Fusion of PAC1-R or PAC1-Rmc with the green fluorescent protein revealed that both receptors are expressed and targeted to the plasma membrane in transfected cells. The different PAC1-R variants are highly expressed in the frog brain and spinal cord and to a lesser extent in peripheral tissues, where only certain isoforms could be detected. The present data indicate that in frog, PACAP may act through different PAC1-R splice variants that differ in their G(s) protein coupling and their abundance in various tissues.
垂体腺苷酸环化酶激活多肽（PACAP）通过激活两种类型的G蛋白偶联受体发挥其多种作用，一种是对PACAP具有高亲和力的受体，称为PAC1-R，另两种对PACAP和血管活性肠肽具有相似亲和力的受体，称为VPAC1-R和VPAC2-R。在此，我们报告了青蛙泽蛙中PAC1-R及其新剪接变体的特征。青蛙PAC1-R与人类PAC1-R具有78%的同源性，并且在中枢神经系统中高度表达。我们对青蛙受体的两种剪接变体进行了特征分析，它们在第三个细胞内环中显示出额外的氨基酸盒。PAC1-R25带有一个25个氨基酸的插入片段，与哺乳动物受体的跳跃盒匹配，而PAC1-R41携带一个与任何哺乳动物PAC1-R变体均无同源性的盒。还鉴定出了PAC1-R的第三种剪接变体，其细胞内C末端结构域完全不同，命名为PAC1-Rmc。对转染了克隆受体的细胞中cAMP形成的测定表明，PACAP以相似的效力激活PAC1-R、PAC1-R25和PAC1-R41。相比之下，PACAP未能刺激转染了PAC1-Rmc的细胞中的腺苷酸环化酶。PAC1-R或PAC1-Rmc与绿色荧光蛋白的融合显示，两种受体均在转染细胞中表达并靶向质膜。不同的PAC1-R变体在青蛙脑和脊髓中高度表达，在周围组织中表达程度较低，在周围组织中只能检测到某些亚型。目前的数据表明，在青蛙中，PACAP可能通过不同的PAC1-R剪接变体起作用，这些变体在与G(s)蛋白偶联以及在各种组织中的丰度方面存在差异。
