UV-B辐射对颠茄氮代谢及次生代谢产物含量的影响

Effects of UV-B on the Activities of Nitrogen Metabolism and Secondary Metabolites of Atropa belladonna

有害的中波紫外线(ultraviolet B,UV-B;280~320 nm)辐射影响植物的生长与发育。但也有研究证明,UV-B辐射可诱导生物碱合成。然而,UV-B辐射能否提高颠茄(Atropa belladonna L.)中托品烷类生物碱(tropane alkaloids,TAs)的含量尚未见报道。本研究以颠茄实生苗为材料,研究UV-B不同照射度强、时间(d数)对颠茄的氮代谢、生物碱含量及TAs代谢途径中的几个关键酶基因表达量的影响。结果表明,随着辐射天数的增加(5~30 d),低强度(LU,5μW/cm^2)UV-B处理与对照(无辐射)比较,硝态氮、莨菪碱、东莨菪碱含量无显著差异。然而,中等强度(MU,10μW/cm^2)和高强度(HU,15μW/cm^2)UV-B辐射,明显增加硝态氮含量,谷氨酰胺合成酶(glutamine synthetase,GS)、谷氨酸脱氢酶(glutamine dehydrogenase,GDH)活性明显高于对照。重要的是,中、高强度UV-B辐射显著降低了颠茄的叶片与茎中莨菪碱和东莨菪碱含量。荧光定量PCR揭示,莨菪碱合成的关键酶腐胺N-甲基转移酶(putrescine N-methyltransferase,PMT)编码基因、莨菪碱-6-β-羟化酶(hyoscyamine-6-β-hydroxylase,H6H)基因表达呈高度组织特异性,主要是在根部表达。与对照比较,低强度照射25 d引起pmt在根部的表达量显著上调,而中、高强度照射导致其下调;h6h在根部的相对表达量随着处理强度的增加逐渐降低;托品酮还原酶Ⅰ(tropinone reductaseⅠ,TRⅠ)编码基因在叶片中的表达量较高,随照射强度的增加而升高。上述结果表明,低强度UV-B辐射促进氮代谢,有利于莨菪碱合成;而长期中、高强度UV-B辐射,尽管促进了谷氨酸代谢,但却使pmt和h6h表达降低,不利于莨菪碱和东莨菪碱的积累。总之,本研究结果显示,不同UV-B辐射强度和时间,对颠茄合成TAs的影响不同,可为大田试验生产莨菪碱提供有益的参考。

Ultraviolet B （UV-B） radiation has harmful effects on plant growth and development. Alkaloid synthesis can be induced by UV-B radiation. However, it has not been reported whether UV-B radiation increases production of tropane alkaloids （TAs） of the Atropa belladonna. In this research we used A. belladonna seedlings to investigate nitrogen metabolism, alkaloid contents, key enzyme geneexpression of TA metabolic pathways in response to different doses of UV-B radiation and treatment durations. The results showed that low-intensity UV-B radiation （LU, 5 μW/cm2） did not affect the content of nitrate nitrogen, hyoscyamine or scopolamine with increased treatment time. However, medium-intensity （ MU, 10 μW/cm2 ） and high-intensity UV-B radiation （ HU, 15 μW/cm2 ） significantly increased nitrate nitrogen contents, glutamine synthetase （GS） activities and glutamine dehydrogenase （GDH） activities. Importantly, MU and HU significantly reduced the contents of belladonna hyoscyamine and scopolamine in leaves and stems. Using real-time fluorescence quantitative PCR, we found that the coding genes of putrescine N-methyltransferase （pmt） and hyoscyamine-6-β- hydroxylase （h6h） showed significant tissue-specific expression patterns, especially in roots. Compared with the control group, LU for 25 days increased the pmt expression levels significantly, but MU and HU reduced pmt expression levels. The relative gene expression levels of h6h reduced gradually with increased irradiation dosage. The relative gene expression levels of trI , the coding gene of tropinone reductase I , were higher in the leaves, which were elevated with increased intensity. The results above showed that LU is favorable for nitrogen metabolism and hyoscyamine synthesis. MU and HU promote glutamate metabolism, but reduce the gene expression levels of pmt and h6h, thus not favorable to accumulate hyoscyamine or scopolamine. In summary, distinct UV-B radiation intensity or durations have different effects for TA synthesis in A. belladonna. This research could provide a beneficial reference for field oroduction of hvoscvamine.