Complete cDNA sequence of a cytochrome P-450 inducible by glucocorticoids in human liver.
Complete cDNA sequence of a cytochrome P-450 inducible by glucocorticoids in human liver.
复制标题
人类肝脏中糖皮质激素诱导的细胞色素 P-450 的完整 cDNA 序列。
DOI：
10.1073/pnas.83.14.5311
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发表时间：
1986
影响因子：
11.1
通讯作者：
Guzelian,PS
中科院分区：
文献类型：
--
作者：
Molowa,DT;Schuetz,EG;Wrighton,SA;Watkins,PB;Kremers,P;Mendez-Picon,G;Parker,GA;Guzelian,PS
HLp是一种人肝细胞色素P-450，与糖皮质激素诱导的大鼠肝细胞色素P-450p和兔肝细胞色素P-450 LM3c免疫化学相关。为了研究HLp的结构和调控，我们使用一种识别纯化HLp的单克隆抗体筛选了lambda gt11中的人肝脏cDNA文库。我们分离并测序了两个重叠的cDNA克隆，它们跨越了编码504个氨基酸的蛋白质的mRNA的整个2011个碱基。预测该蛋白的氨基末端氨基酸序列与纯化HLp测定的前20个残基相同。HLp mRNA与大鼠和家兔的相关蛋白序列同源性超过70%，但与其他已发表的细胞色素P-450基因同源性低于40%。此外，人类和大鼠基因组DNA的Southern blot分析显示，分别有50和60千碱基的DNA可与HLp DNA杂交。对5例患者的人肝脏RNA进行Blot分析，发现与HLp cdna杂交的主要条带（2.2千碱基）和次要条带（3.0千碱基）。在三名接受地塞米松（一种强效糖皮质激素）治疗的患者中，来自同一肝脏的微粒体中这些杂交mrna的表观浓度以及免疫反应性HLp蛋白的数量呈剂量依赖关系增加。此外，在含有地塞米松的培养基中培养120小时的人肝癌细胞系（Hep G2）培养物中分离的RNA和微体样本中，与在无类固醇培养基中培养的培养液相比，两种可与HLp cdna杂交的mRNA的诱导量增加了6倍，免疫反应性HLp蛋白的诱导量增加了3倍。我们得出结论，HLp是一个保守的糖皮质激素诱导的细胞色素P-450基因家族的人类代表，其诱导机制涉及HLp mRNA的积累。
HLp is a human liver cytochrome P-450 that is immunochemically related to the glucocorticoid-inducible liver cytochrome P-450p in the rat and its homologue in the rabbit, P-450 LM3c. To investigate the structure and regulation of HLp, we used a monoclonal antibody that recognizes purified HLp to screen a human liver cDNA library in lambda gt11. We isolated and sequenced two overlapping cDNA clones that span the entire 2011 bases of an mRNA that codes for a protein of 504 amino acids. The predicted amino-terminal amino acid sequence of this protein is identical to the first 20 residues determined from purified HLp. HLp mRNA shares more than 70% sequence homology with related proteins from the rat and rabbit but less than 40% homology with other published cytochrome P-450 genes. Moreover, Southern blot analysis of human and rat genomic DNA revealed 50 and 60 kilobases of DNA, respectively, hybridizable to the HLp cDNAs. Blot analysis of human liver RNA from five patients revealed major (2.2 kilobase) and minor (3.0 kilobase) bands that hybridized to HLp cDNAs. The apparent concentration of these hybridizable mRNAs as well as the amounts of immunoreactive HLp protein in microsomes from the same liver were increased in a dose-dependent relationship in three patients who received dexamethasone, a potent glucocorticoid. Furthermore, in samples of RNA and of microsomes isolated from cultures of a human hepatoma cell line (Hep G2) incubated for 120 hr in medium containing dexamethasone, there was a 6-fold induction of the two mRNA species hybridizable to HLp cDNAs and a 3-fold induction of immunoreactive HLp protein as compared to the values for cultures incubated in steroid-free medium. We conclude that HLp is a human representative of a conserved glucocorticoid-inducible cytochrome P-450 gene family whose mechanism of induction involves accumulation of HLp mRNA.