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稀有鮈鲫3-羟基-3-甲基戊二酰辅酶A还原酶全长cDNA的克隆及其在五氯酚暴露雄性鱼中的表达谱分析

[Cloning of full-length cDNA of HMGR from Gobiocypris rarus and analysis of its expression profiles in male exposed to pentachlorophenol].

作者信息

Deng Chuan, Mao Si-Yu, Xiong Li, Zhang Xiao-Zheng, Li Wei, Gao Xiang, Liu Qiu-Ping, Chen Yun, Liu Yan

出版信息

Huan Jing Ke Xue. 2014 Aug;35(8):3183-91.

PMID:25338397
Abstract

3-Hydroxy-3-methylglutaryl coenzyme A reductase (HMGR) is the first rate-limiting enzyme in the mevalonate (MVA) pathway. The full-length cDNA of HMGR was cloned from Gobiocypris rarus, and HMGR expression profiles in different tissues and in response to different treatments of pentachlorophenol (PCP) were analyzed by real-time PCR, to investigate the endocrine disruption mechanism of PCP, which altered steroid hormone precursors (cholesterol) levels by modulating gene transcription profiles of HMGR. Based on the homologous clone strategy and rapid-amplification of cDNA ends (RACE) technology, the full-length 3 101-base-pair (bp) cDNA of HMGR was isolated from the livers of rare minnow (Gobiocypris rarus) for the first time, and was designated as GrHMGR (GenBank accession number KF885724). GrHMGR encoded a protein of 884 amino acids and phylogenetic tree analysis indicated that the deduced protein GrHMGR had extensive sequence similarities to other fish HMGRs. Real-time PCR analyses indicated that GrHMGR mRNA expression was tightly controlled in a tissue-specific fashion, with the sites of expression being brain, gonads and liver, and the highest site of expression being gonads. After male rare minnows were exposed to different concentrations of PCP, significant decrease in GrHMGR gene expression with increased PCP concentration in the brain and gonads were observed, together with the differential gene expression trend in the liver. Furthermore, it was found that the decrease of HMGR could reduce the synthesis of cholesterol. This proved that PCP might disrupt the pathway of cholesterol synthesis and then influenced the endocrine system of rare minnow.

摘要

3-羟基-3-甲基戊二酰辅酶A还原酶(HMGR)是甲羟戊酸(MVA)途径中的首个限速酶。从稀有鮈鲫中克隆了HMGR的全长cDNA,并通过实时定量PCR分析了HMGR在不同组织中的表达谱以及对五氯苯酚(PCP)不同处理的响应,以研究PCP的内分泌干扰机制,PCP通过调节HMGR的基因转录谱改变类固醇激素前体(胆固醇)水平。基于同源克隆策略和cDNA末端快速扩增(RACE)技术,首次从稀有鮈鲫肝脏中分离出全长3101个碱基对(bp)的HMGR cDNA,并将其命名为GrHMGR(GenBank登录号KF885724)。GrHMGR编码一个884个氨基酸的蛋白质,系统发育树分析表明推导的蛋白质GrHMGR与其他鱼类的HMGR具有广泛的序列相似性。实时定量PCR分析表明,GrHMGR mRNA表达以组织特异性方式受到严格控制,表达部位为脑、性腺和肝脏,表达量最高的部位是性腺。雄性稀有鮈鲫暴露于不同浓度的PCP后,观察到脑和性腺中GrHMGR基因表达随PCP浓度升高而显著降低,肝脏中也有不同的基因表达趋势。此外,发现HMGR的降低会减少胆固醇的合成。这证明PCP可能会破坏胆固醇合成途径,进而影响稀有鮈鲫的内分泌系统。

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