Enhanced production of melatonin by ectopic overexpression of human serotonin N-acetyltransferase plays a role in cold resistance in transgenic rice seedlings
Enhanced production of melatonin by ectopic overexpression of human serotonin N-acetyltransferase plays a role in cold resistance in transgenic rice seedlings
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DOI：
10.1111/j.1600-079x.2010.00783.x
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发表时间：
2010-09-01
影响因子：
10.3
通讯作者：
Back, Kyoungwhan
中科院分区：
文献类型：
--
作者：
Kang, Kiyoon;Lee, Kyungjin;Back, Kyoungwhan
5-羟色胺N-乙酰转移酶（SNA）是脊椎动物褪黑激素生物合成的限速酶，负责N-乙酰5-羟色胺的产生;然后该分子通过羟吲哚-O-甲基转移酶转化为褪黑激素。我们利用农杆菌介导的基因转化法，通过在组成型泛素启动子下表达人SNA基因，获得了转基因水稻植株。本文研究了SNA在褪黑激素生物合成中的作用以及褪黑激素在水稻中的生理作用。通过Southern、北方和RT-PCR分析证实了转基因水稻在T（1）转基因水稻中的整合和表达。在转基因水稻植物中观察到了高水平的SNA特异性酶活性，而野生型则显示出微量的SNA酶活性。SNA蛋白的功能性表达与转基因水稻中N-乙酰5-羟色胺和褪黑激素合成的增加密切相关。实验同时使用外源性治疗的血清素和衰老的离体叶片，其中包含一个池的血清素，显着增强褪黑激素的生物合成，表明内源性血清素水平发挥了瓶颈作用的褪黑激素的生物合成途径。最后，转基因水稻幼苗与高水平的褪黑激素表现出较高的叶绿素合成在冷胁迫，这表明褪黑激素在抗冷胁迫的作用。
Serotonin N-acetyltransferase (SNA), a rate-limiting enzyme in melatonin biosynthesis in vertebrates, is responsible for the production of N-acetylserotonin; this molecule is then converted to melatonin by hydroxyindole-O-methyltransferase. We generated transgenic rice plants via expression of the human SNA gene under the constitutive ubiquitin promoter using Agrobacterium-mediated gene transformation. We investigated the role of SNA in the biosynthesis of melatonin and the physiological role of melatonin in rice plants. The integration and expression of the transgene were confirmed in T(1) transgenic rice seedlings by Southern, Northern, and RT-PCR analyses. High SNA-specific enzyme activities were observed in the transgenic rice plants, whereas the wild type revealed a trace level of SNA enzyme activity. The functional expression of SNA protein was closely associated with the elevated synthesis of N-acetylserotonin and melatonin in the transgenic rice plants. Experiments using both exogenous treatment of serotonin and senescent detached leaves, which contain a pool of serotonin, significantly enhanced melatonin biosynthesis, indicating that endogenous serotonin levels play a bottleneck role in the pathway of melatonin biosynthesis. Finally, the transgenic rice seedlings with high levels of melatonin showed elevated chlorophyll synthesis during cold stress, suggesting a role for melatonin in cold-stress resistance.