Effect of dioxins on regulation of tyrosine hydroxylase gene expression by aryl hydrocarbon receptor: a neurotoxicology study
Effect of dioxins on regulation of tyrosine hydroxylase gene expression by aryl hydrocarbon receptor: a neurotoxicology study
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DOI：
10.1186/1476-069x-8-24
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发表时间：
2009-06-06
影响因子：
6
通讯作者：
Ishihara-Sugano, Mitsuko
中科院分区：
文献类型：
--
作者：
Akahoshi, Eiichi;Yoshimura, Seiko;Ishihara-Sugano, Mitsuko
背景：二恶英和相关化合物被怀疑会导致神经系统紊乱。流行病学研究表明，暴露于这些化合物引起神经发育障碍，如学习障碍和注意缺陷多动障碍，这被认为是密切相关的多巴胺能功能障碍。虽然其作用的分子机制尚未完全研究，但该过程的主要参与者是芳烃受体（AhR）。本研究旨在探讨2，3，7，8-四氯二苯并-对-二恶英（TCDD）暴露对AhR调节多巴胺合成限速酶TH基因表达的影响。RT-PCR和Western blotting检测TCDD诱导的TH表达。通过使用siRNA方法抑制AhR表达证实AhR参与TCDD诱导的TH基因表达。通过高效液相色谱法测量包括多巴胺在内的儿茶酚胺。报告基因分析用于鉴定TH基因启动子区的调控基序。结果：TCDD通过激活AhR在mRNA和蛋白水平上诱导TH的形成。诱导的TH蛋白是功能性的，其表达增加多巴胺的合成。报告基因分析和EMSA表明AhR直接调控TH基因的表达。在TH基因上游-285 ~-167 bp处发现了一段调控序列，称为芳烃受体反应元件III（AHRE-III）。在TCDD暴露下，AhR复合物结合到AHRE-III以及异生素反应元件（XRE），虽然AHRE-III是不相同的XRE，传统的AhR-binding motif.Conclusion：我们的研究结果表明，TCDD直接调节多巴胺系统TH基因的反式激活通过AhR-AHRE-III介导的途径。AhR介导的通路可能具有特定的AhR介导的基因组控制通路，其在多巴胺能障碍的发展中将TCDD作用的效应传递至靶细胞。
Background: Dioxins and related compounds are suspected of causing neurological disruption. Epidemiological studies indicated that exposure to these compounds caused neurodevelopmental disturbances such as learning disability and attention deficit hyperactivity disorder, which are thought to be closely related to dopaminergic dysfunction. Although the molecular mechanism of their actions has not been fully investigated, a major participant in the process is aryl hydrocarbon receptor (AhR). This study focused on the effect of 2, 3, 7, 8-tetrachlorodibenzo-p-dioxin (TCDD) exposure on the regulation of TH, a rate-limiting enzyme of dopamine synthesis, gene expression by AhR.Methods: N2a-R beta cells were established by transfecting murine neuroblastoma Neuro2a with the rat AhR cDNA. TH expression induced by TCDD was assessed by RT-PCR and Western blotting. Participation of AhR in TCDD-induced TH gene expression was confirmed by suppressing AhR expression using the siRNA method. Catecholamines including dopamine were measured by high-performance liquid chromatography. A reporter gene assay was used to identify regulatory motifs in the promoter region of TH gene. Binding of AhR with the regulatory motif was confirmed by an electrophoretic mobility shift assay (EMSA).Results: Induction of TH by TCDD through AhR activation was detected at mRNA and protein levels. Induced TH protein was functional and its expression increased dopamine synthesis. The reporter gene assay and EMSA indicated that AhR directly regulated TH gene expression. Regulatory sequence called aryl hydrocarbon receptor responsive element III (AHRE-III) was identified upstream of the TH gene from -285 bp to -167 bp. Under TCDD exposure, an AhR complex was bound to AHRE-III as well as the xenobiotic response element (XRE), though AHRE-III was not identical to XRE, the conventional AhR-binding motif.Conclusion: Our results suggest TCDD directly regulate the dopamine system by TH gene transactivation via an AhR-AHRE-III-mediated pathway. The AhR- mediated pathway could have a particular AhR- mediated genomic control pathway transmitting the effects of TCDD action to target cells in the development of dopaminergic disabilities.