Guo Xiaohao, Ran Le, Huang Xinyu, Wang Xiuchen, Zhu Jiantang, Tan Yuanyuan, Shu Qingyao
State Key Laboratory of Rice Biology and Breeding, and Zhejiang Provincial Key Laboratory of Crop Germplasm Innovation and Exploitation, The Advanced Seed Institute, Zhejiang University, Hangzhou, China.
School of Biological Science and Technology, University of Jinan, Jinan, China.
Front Plant Sci. 2024 Oct 1;15:1478200. doi: 10.3389/fpls.2024.1478200. eCollection 2024.
Melatonin, a tryptophan-derived indoleamine metabolite with important roles in plant growth and defense, has recently been regarded as a new plant hormone. Maize is one of the most important cereal crops in the world. Although the melatonin receptor gene, , has already been identified, the genetic basis of melatonin biosynthesis in maize has still not been elucidated. (SNAT) is the enzyme that converts serotonin to N-acetylserotonin (NAS) or 5-methoxytryptamine (5MT) to melatonin in Arabidopsis and rice, but no SNAT encoding gene has been identified yet in maize.
The bioinformatics analysis was used to identify maize SNAT genes and the enzyme activity of the recombinant proteins was determined through in vitro assay. The expression levels of and under drought and heat stresses were revealed by public RNA-seq datasets and qRT-PCR analysis.
We first identified three maize SNAT genes, ZmSNAT1, ZmSNAT2, and ZmSNAT3, through bioinformatics analysis, and demonstrated that ZmSNAT2 was present in only eight of the 26 cultivars analyzed. We then determined the enzyme activity of ZmSNAT1 and ZmSNAT3 using their recombinant proteins through in vitro assay. The results showed that both ZmSNAT1 and ZmSNAT3 could convert serotonin to NAS and 5-MT to melatonin. Recombinant ZmSNAT1 catalyzed serotonin into NAS with a higher catalytic activity ( , 8.6 mM; , 4050 pmol/min/mg protein) than ZmSNAT3 ( , 11.51 mM; , 142 pmol/min/mg protein). We further demonstrated that the 228th amino acid Tyr (Y228) was essential for the enzymatic activity of ZmSNAT1. Finally, we revealed that the expression of ZmSNAT1 and ZmSNAT3 varied among different maize cultivars and different tissues of a plant, and was responsive to drought and heat stresses.
In summary, the present study identified and characterized the first two functional SNAT genes in maize, laying the foundation for further research on melatonin biosynthesis and its regulatory role in plant growth and response to abiotic stresses.
褪黑素是一种由色氨酸衍生的吲哚胺代谢产物，在植物生长和防御中具有重要作用，最近被视为一种新的植物激素。玉米是世界上最重要的谷类作物之一。尽管已经鉴定出褪黑素受体基因，但玉米中褪黑素生物合成的遗传基础仍未阐明。在拟南芥和水稻中，血清素N-乙酰基转移酶（SNAT）是一种将血清素转化为N-乙酰血清素（NAS）或5-甲氧基色胺（5MT）转化为褪黑素的酶，但尚未在玉米中鉴定出编码SNAT的基因。
利用生物信息学分析鉴定玉米SNAT基因，并通过体外测定确定重组蛋白的酶活性。通过公共RNA测序数据集和定量逆转录聚合酶链反应（qRT-PCR）分析揭示了干旱和热胁迫下ZmSNAT1和ZmSNAT3的表达水平。
我们首先通过生物信息学分析鉴定了三个玉米SNAT基因，即ZmSNAT1、ZmSNAT2和ZmSNAT3，并证明ZmSNAT2仅存在于所分析的26个品种中的8个品种中。然后，我们通过体外测定使用它们的重组蛋白确定了ZmSNAT1和ZmSNAT3的酶活性。结果表明，ZmSNAT1和ZmSNAT3都可以将血清素转化为NAS，并将5-MT转化为褪黑素。重组ZmSNAT1催化血清素生成NAS的催化活性（Km，8.6 mM；Vmax，4050 pmol/min/mg蛋白）高于ZmSNAT3（Km，11.51 mM；Vmax，142 pmol/min/mg蛋白）。我们进一步证明第228位氨基酸酪氨酸（Y228）对ZmSNAT1的酶活性至关重要。最后，我们揭示了ZmSNAT1和ZmSNAT3的表达在不同玉米品种和植物的不同组织中有所不同，并且对干旱和热胁迫有响应。
总之，本研究鉴定并表征了玉米中前两个功能性SNAT基因，为进一步研究褪黑素生物合成及其在植物生长和对非生物胁迫响应中的调节作用奠定了基础。
