CLONING AND CHARACTERIZATION OF A FUNCTIONALLY DISTINCT CORTICOTROPIN-RELEASING FACTOR-RECEPTOR SUBTYPE FROM RAT-BRAIN
CLONING AND CHARACTERIZATION OF A FUNCTIONALLY DISTINCT CORTICOTROPIN-RELEASING FACTOR-RECEPTOR SUBTYPE FROM RAT-BRAIN
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DOI：
10.1073/pnas.92.3.836
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发表时间：
1995-01-31
影响因子：
11.1
通讯作者：
OLTERSDORF, T
中科院分区：
文献类型：
--
作者：
LOVENBERG, TW;LIAW, CW;OLTERSDORF, T
本研究报道了一个cDNA克隆的分离，该克隆编码促肾上腺皮质激素释放因子（CRF）受体家族的第二个成员，命名为CRF（2）受体。在PCR范例中使用简并序列的寡核苷酸鉴定cDNA。利用从大鼠脑中获得的PCR片段从大鼠下丘脑cDNA文库中分离全长cDNA，该cDNA编码411个氨基酸的蛋白质，在整个编码区与已知的CRF（1）受体具有约70%的同一性。当在小鼠Ltk（-）细胞中表达时，该受体刺激cAMP产生以响应CRF和已知的CRF样激动剂。CRF和非哺乳动物CRF相关肽sauvagine和尾加压素I以剂量依赖性方式刺激腺苷酸环化酶活性，其效力等级顺序不同于CRF（1）受体：sauvagine >尾加压素大于或等于大鼠/人CRF >绵羊CRF。逆转录-聚合酶链反应（RT-PCR）分析CRF（2）受体mRNA的组织分布表明，CRF（2）受体mRNA存在于大鼠脑中，在肺和心脏中也可检测到。原位杂交研究表明，在脑内的下丘脑腹内侧核，外侧隔，杏仁核，和内嗅皮质的特异性表达，但在垂体中的表达不显着。通过PCR鉴定了CRF（2）受体的另一个剪接变体，其具有不同的N-末端结构域，编码431个氨基酸的推定蛋白质。因此，数据表明存在另一种功能性CRF受体，与CRF（1）受体在药理学特征和组织分布方面存在显著差异，这将预测两种受体之间的重要功能差异。
The present study reports the isolation of a cDNA clone that encodes a second member of the corticotropin-releasing factor (CRF) receptor family, designated as the CRF(2) receptor. The cDNA was identified using oligonucleotides of degenerate sequence in a PCR paradigm. A PCR fragment obtained from rat brain was utilized to isolate a full-length cDNA from a rat hypothalamus cDNA library that encoded a 411-amino acid protein with approximate to 70% identity to the known CRF(1) receptor over the entire coding region. When expressed in mouse Ltk(-) cells, this receptor stimulates cAMP production in response to CRF and known CRF-like agonists. CRF and the nonmammalian CRF-related peptides sauvagine and urotensin I stimulate adenylate cyclase activity in a dose-dependent manner with a rank order of potency different from that of the CRF(1) receptor: sauvagine > urotensin greater than or equal to rat/human CRF > ovine CRF. Tissue distribution analysis of the mRNAs by reverse transcriptase-PCR shows CRF(2) receptor mRNA is present in rat brain and detectable in lung and heart. In situ hybridization studies indicate specific expression within the brain in the ventromedial nuclei of the hypothalamus, the lateral septum, the amygdala, and entorhinal cortex, but there is unremarkable expression in the pituitary. An additional splice variant of the CRF(2) receptor with a different N-terminal domain has been identified by PCR, encoding a putative protein of 431 amino acids. Thus, the data demonstrate the presence of another functional CRF receptor, with significant differences in the pharmacological profile and tissue distribution from the CRF(1) receptor, which would predict important functional differences between the two receptors.