MOLECULAR-CLONING OF A HUMAN CANNABINOID RECEPTOR WHICH IS ALSO EXPRESSED IN TESTIS
MOLECULAR-CLONING OF A HUMAN CANNABINOID RECEPTOR WHICH IS ALSO EXPRESSED IN TESTIS
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DOI：
10.1042/bj2790129
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发表时间：
1991-10-01
影响因子：
4.1
通讯作者：
PARMENTIER, M
中科院分区：
文献类型：
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作者：
GERARD, CM;MOLLEREAU, C;PARMENTIER, M
从人脑干cDNA文库中分离到一个具有鸟嘌呤-核苷酸结合蛋白（g蛋白）偶联受体全部特征的受体蛋白cDNA克隆。所使用的探针（HGMP08）是一个600 bp的DNA片段，以人类基因组DNA为模板，用已知g蛋白偶联受体中保守序列对应的退化寡核苷酸引物，经低强度PCR扩增而成。推断出的氨基酸序列编码了一个含有472个残基的蛋白质，与最近克隆的大鼠大麻素受体有97.3%的相似性[Matsuda， Lolait, Brownstein, Young & Bronner (1990) Nature (London) 346,561 -564]。正如预期的那样，在大脑中发现了大量的转录本，但在睾丸中也发现了少量的转录本。同样的探针被用于筛选人类睾丸cDNA文库。对获得的cDNA克隆进行了部分测序，证明了在两种组织中表达的大麻素受体的身份。合成大麻素配体[H-3]CP55940在转染重组受体克隆的Cos-7细胞膜上特异性结合。在稳定转染的CHO-K1细胞系中，大麻素激动剂介导了福斯克林诱导的环AMP积累的剂量依赖性和立体选择性抑制。在哺乳动物细胞中表达人类大麻素受体的能力应该有助于开发更具选择性的药物，并且应该促进寻找内源性大麻素配体。
A cDNA clone encoding a receptor protein which presents all the characteristics of a guanine-nucleotide-binding protein (G-protein)-coupled receptor was isolated from a human brain stem cDNA library. The probe used (HGMP08) was a 600 bp DNA fragment amplified by a low-stringency PCR, using human genomic DNA as template and degenerate oligonucleotide primers corresponding to conserved sequences amongst the known G-protein-coupled receptors. The deduced amino acid sequence encodes a protein of 472 residues which shares 97.3 % identity with the rat cannabinoid receptor cloned recently [Matsuda, Lolait, Brownstein, Young & Bronner (1990) Nature (London) 346, 561-564]. Abundant transcripts were detected in the brain, as expected, but lower amounts were also found in the testis. The same probe was used to screen a human testis cDNA library. The cDNA clones obtained were partially sequenced, demonstrating the identity of the cannabinoid receptors expressed in both tissues. Specific binding of the synthetic cannabinoid ligand [H-3]CP55940 was observed on membranes from Cos-7 cells transfected with the recombinant receptor clone. In stably transfected CHO-K1 cell lines, cannabinoid agonists mediated a dose-dependent and stereoselective inhibition of forskolin-induced cyclic AMP accumulation. The ability to express the human cannabinoid receptor in mammalian cells should help in developing more selective drugs, and should facilitate the search for the endogenous cannabinoid ligand(s).