Zebrafish arylalkylamine-N-acetyltransferase genes -: targets for regulation of the circadian clock
Zebrafish arylalkylamine-N-acetyltransferase genes -: targets for regulation of the circadian clock
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DOI：
10.1677/jme.1.01893
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发表时间：
2006-04-01
影响因子：
3.5
通讯作者：
Gothilf, Y
中科院分区：
文献类型：
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作者：
Appelbaum, L;Vallone, D;Gothilf, Y
褪黑激素产生的每日节律由芳基烷基胺-N-乙酰转移酶（AANAT）活性的变化控制。斑马鱼具有两个aanat，aanat 1和aanat 2;前者仅在视网膜中表达，后者在视网膜和松果体中都表达。在这里，他们的差异表达和调节进行了研究，使用转录定量和瞬时和稳定的体内和体外转染试验。在松果体中，aanat 2启动子表现出生物钟控制的活性，如在AANAT 2：EGFP转基因鱼中增强型绿色荧光蛋白（EGFP）mRNA的昼夜节律所示。在体内瞬时表达分析的aanat 2启动子表明，E盒和感光细胞保守元件（PCE）是必需的表达在松果体。在视网膜中，这两个基因的表达的特点是一个强大的昼夜节律的转录水平。在持续黑暗中，视网膜aanat 2的节律性表达持续存在，而aanat 1节律消失;表明前者受生物钟控制，后者也是光驱动的。在含有光致生物钟的PAC-2斑马鱼细胞系中，稳定转染的aanat 1和aanat 2启动子均表现出生物钟控制的昼夜节律，这是E盒驱动表达的特征。在NIH-3 T3细胞中的瞬时共转染实验显示，这两个，E-box和PCE-containing，启动子是由BMAL/CLOCK和orthedenticle同源框5的协同作用驱动的。这项研究揭示了两个相关基因调控的共同机制，但描述了它们的差异阶段和光响应，这可能是由其他基因特异性调控机制和组织特异性转录因子谱驱动的。
Daily rhythms of melatonin production are controlled by changes in the activity of arylalkylamine-N-acetyltransferase (AANAT). Zebrafish possess two aanats, aanat1 and aanat2; the former is expressed only in the retina and the latter is expressed in both the retina and the pineal gland. Here, their differential expression and regulation were studied using transcript quantification and transient and stable in vivo and in vitro transfection assays. In the pineal gland, the aanat2 promoter exhibited circadian clock-controlled activity, as indicated by circadian rhythms of Enhanced green fluorescent protein (EGFP) mRNA in AANAT2:EGFP transgenic fish. In vivo transient expression analyses of the aanat2 promoter indicated that E-box and photoreceptor conserved elements (PCE) are required for expression in the pineal gland. In the retina, the expression of both genes was characterized by a robust circadian rhythm of their transcript levels. In constant darkness, the rhythmic expression of retinal aanat2 persisted while the aanat1 rhythm disappeared; indicating that the former is controlled by a circadian clock and the latter is also light driven. In the light-entrainable clock-containing PAC-2 zebrafish cell line, both stably transfected aanat1 and aanat2 promoters exhibited a clock-controlled circadian rhythm, characteristic for an E-box-driven expression. Transient co-transfection experiments in NIH-3T3 cells revealed that the two, E-box- and PCE-containing, promoters are driven by the synergistic action of BMAL/CLOCK and orthehodenticle homeobox 5. This study has revealed a shared mechanism for the regulation of two related genes, yet describes their differential phases and photic responses which may be driven by other gene-specific regulatory mechanisms and tissue-specific transcription factor profiles.