Wang Feibing, Zhu Hong, Kong Weili, Peng Rihe, Liu Qingchang, Yao Quanhong
Shanghai Key Laboratory of Agricultural Genetic Breeding, Biotech Research Institute, Shanghai Academy of Agricultural Sciences, Shanghai, 201106, China.
Beijing Key Laboratory of Crop Genetic Improvement/Laboratory of Crop Heterosis and Utilization, Ministry of Education, China Agricultural University, Beijing, 100193, China.
Planta. 2016 Jul;244(1):59-73. doi: 10.1007/s00425-016-2489-3. Epub 2016 Mar 5.
A basic helix-loop-helix (bHLH) transcription factor gene from Antirrhinum, AmDEL , increases flavonoids accumulation and enhances salt and drought tolerance via up-regulating flavonoid biosynthesis, proline biosynthesis and ROS scavenging genes in transgenic Arabidopsis. In plants, transcriptional regulation is the most important tools for increasing flavonoid biosynthesis. The AmDEL gene, as a basic helix-loop-helix transcription factor gene from Antirrhinum, has been shown to increase flavonoids accumulation in tomato. However, its role in tolerance to abiotic stresses has not yet been investigated. In this study, the codon-optimized AmDEL gene was chemically synthesized. Subcellular localization analysis in onion epidermal cells indicated that AmDEL protein was localized to the nucleus. Expression analysis in yeast showed that the full length of AmDEL exhibited transcriptional activation. Overexpression of AmDEL significantly increased flavonoids accumulation and enhanced salt and drought tolerance in transgenic Arabidopsis plants. Real-time quantitative PCR analysis showed that overexpression of AmDEL resulted in the up-regulation of genes involved in flavonoid biosynthesis, proline biosynthesis and ROS scavenging under salt and drought stresses. Meanwhile, Western blot and enzymatic analyses showed that the activities of phenylalanine ammonia lyase, chalcone isomerase, dihydroflavonol reductase, pyrroline-5-carboxylate synthase, superoxide dismutase and peroxidase were also increased. Further components analyses indicated that the significant increase of proline and relative water content and the significant reduction of H2O2 and malonaldehyde content were observed under salt and drought stresses. In addition, the rates of electrolyte leakage and water loss were reduced in transgenic plants. These findings imply functions of AmDEL in accumulation of flavonoids and tolerance to salt and drought stresses. The AmDEL gene has the potential to be used to increase the content of valuable flavonoids and improve tolerance to abiotic stresses in plants.
金鱼草中的一个碱性螺旋-环-螺旋(bHLH)转录因子基因AmDEL,通过上调转基因拟南芥中类黄酮生物合成、脯氨酸生物合成和活性氧清除基因,增加类黄酮积累并增强耐盐性和耐旱性。在植物中,转录调控是增加类黄酮生物合成的最重要工具。AmDEL基因作为金鱼草中的一个碱性螺旋-环-螺旋转录因子基因,已被证明可增加番茄中的类黄酮积累。然而,其在非生物胁迫耐受性中的作用尚未得到研究。在本研究中,对密码子优化后的AmDEL基因进行了化学合成。洋葱表皮细胞中的亚细胞定位分析表明,AmDEL蛋白定位于细胞核。酵母中的表达分析表明,AmDEL全长具有转录激活作用。AmDEL的过表达显著增加了转基因拟南芥植株中的类黄酮积累,并增强了其耐盐性和耐旱性。实时定量PCR分析表明,AmDEL的过表达导致盐胁迫和干旱胁迫下参与类黄酮生物合成、脯氨酸生物合成和活性氧清除的基因上调。同时,蛋白质免疫印迹和酶活性分析表明,苯丙氨酸解氨酶、查尔酮异构酶、二氢黄酮醇还原酶、吡咯啉-5-羧酸合成酶、超氧化物歧化酶和过氧化物酶的活性也有所增加。进一步的成分分析表明,在盐胁迫和干旱胁迫下,脯氨酸和相对含水量显著增加,过氧化氢和丙二醛含量显著降低。此外,转基因植物的电解质渗漏率和水分流失率降低。这些发现表明AmDEL在类黄酮积累以及耐盐和耐旱胁迫方面具有功能。AmDEL基因有潜力用于增加植物中珍贵类黄酮的含量并提高其对非生物胁迫的耐受性。