Brezillon Stéphane, Lannoy Vincent, Franssen Jean-Denis, Le Poul Emmanuel, Dupriez Vincent, Lucchetti Jean, Detheux Michel, Parmentier Marc
Euroscreen, 802 Route de Lennik, 1070 Brussels, Belgium.
J Biol Chem. 2003 Jan 10;278(2):776-83. doi: 10.1074/jbc.M206396200. Epub 2002 Oct 24.
GPR7 and GPR8 are two structurally related orphan G protein-coupled receptors, presenting high similarities with opioid and somatostatin receptors. Two peptides, L8 and L8C, derived from a larger precursor, were recently described as natural ligands for GPR8 (Mori, M., Shimomura, Y., Harada, M., Kurihara, M., Kitada, C., Asami, T., Matsumoto, Y., Adachi, Y., Watanabe, T., Sugo, T., and Abe, M. (December, 27, 2001) World Patent Cooperation Treaty, Patent Application WO 01/98494A1). L8 is a 23-amino acid peptide, whereas L8C is the same peptide with a C terminus extension of 7 amino acids, running through a dibasic motif of proteolytic processing. Using as a query the amino acid sequence of the L8 peptide, we have identified in DNA databases a human gene predicted to encode related peptides and its mouse ortholog. By analogy with L8 and L8C, two peptides, named L7 and L7C could result from the processing of a 125-amino acid human precursor through the alternative usage of a dibasic amino acid motif. The activity of these four peptides was investigated on GPR7 and GPR8. In binding assays, L7, L7C, L8, and L8C were found to bind with low nanomolar affinities to the GPR7 and GPR8 receptors expressed in Chinese hamster ovary (CHO)-K1 cells. They inhibited forskolin-stimulated cAMP accumulation through a pertussis toxin-sensitive mechanism. The tissue distribution of prepro-L7 (ppL7) and prepro-L8 (ppL8) was investigated by reverse transcription-PCR. Abundant ppL7 transcripts were found throughout the brain as well as in spinal cord, spleen, testis, and placenta; ppL8 transcripts displayed a more restricted distribution in brain, with high levels in substantia nigra, but were more abundant in peripheral tissues. The ppL7 and ppL8 genes therefore encode the precursors of a class of peptide ligands, active on two receptor subtypes, GPR7 and GPR8. The distinct tissue distribution of the receptor and peptide precursors suggest that each ligand and receptor has partially overlapping but also specific roles in this signaling system.
GPR7和GPR8是两个结构相关的孤儿G蛋白偶联受体，与阿片受体和生长抑素受体高度相似。最近，从一个更大的前体衍生而来的两种肽L8和L8C被描述为GPR8的天然配体（森，M.，下村，Y.，原田，M.，栗原，M.，北田，C.，浅见，T.，松本，Y.，安达，Y.，渡边，T.，须古，T.，和阿部，M.（2001年12月27日）《世界专利合作条约》，专利申请WO 01/98494A1）。L8是一种23个氨基酸的肽，而L8C是同一肽，其C末端延伸了7个氨基酸，穿过一个蛋白水解加工的双碱性基序。以L8肽的氨基酸序列作为查询序列，我们在DNA数据库中鉴定出一个预测编码相关肽的人类基因及其小鼠直系同源基因。与L8和L8C类似，两种肽L7和L7C可能是由一个125个氨基酸的人类前体通过双碱性氨基酸基序的交替使用加工而成。研究了这四种肽对GPR7和GPR8的活性。在结合试验中，发现L7、L7C、L8和L8C以低纳摩尔亲和力与中国仓鼠卵巢（CHO）-K1细胞中表达的GPR7和GPR8受体结合。它们通过百日咳毒素敏感机制抑制福斯高林刺激的环磷酸腺苷（cAMP）积累。通过逆转录聚合酶链反应（RT-PCR）研究了前体蛋白L7（ppL7）和前体蛋白L8（ppL8）的组织分布。在整个大脑以及脊髓、脾脏、睾丸和胎盘中发现了丰富的ppL7转录本；ppL8转录本在大脑中的分布更为局限，在黑质中含量较高，但在周围组织中更为丰富。因此，ppL7和ppL8基因编码一类肽配体的前体，这些配体对两种受体亚型GPR7和GPR8具有活性。受体和肽前体不同的组织分布表明，每种配体和受体在这个信号系统中具有部分重叠但也有特定的作用。
