Lipid Transfer Proteins (AaLTP3 and AaLTP4) Are Involved in Sesquiterpene Lactone Secretion from Glandular Trichomes in Artemisia annua
Lipid Transfer Proteins (AaLTP3 and AaLTP4) Are Involved in Sesquiterpene Lactone Secretion from Glandular Trichomes in Artemisia annua
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DOI：
10.1093/pcp/pcz171
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发表时间：
2019-12-01
影响因子：
4.9
通讯作者：
Choi, Yong Eui
中科院分区：
文献类型：
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作者：
Adhikari, Prakash Babu;Han, Jung Yeon;Choi, Yong Eui
在青蒿属植物中，腺毛(GT)负责包括青蒿素/青蒿素B在内的倍半萜内酯的生物合成和分泌。植物中的非特异性脂转移蛋白(LTPS)结合并携带脂质分子穿过细胞膜，也被称为分泌蛋白。有趣的是，LTP基因的转录本在黄曲霉的GT中异常丰富。在本研究中，我们从韩国生态型的黄花蒿中分离到两个毛状体特异的LTP基因(AaLTP3和AaLTP4)。AaLTP3在枝条中大量表达，而AaLTP4在花中表达。由AaLTP3或AaLTP4启动子驱动的GUS信号表明，AaLTP3启动子在毛状非GT上活性，AaLTP4启动子在GTS上活性。对转基因烟草中增强的青色荧光蛋白(ECFP)与AaLTP3或AaLTP4蛋白融合的分析表明，ECFP对长GT分泌的脂类有很强的荧光。此外，短毛状体的头细胞及其分泌颗粒的花期也很明亮。免疫印迹分析表明，黄花苜蓿叶柄中的GT分泌物对AaLTP4抗体呈强阳性信号。与野生型相比，AaLTP3或AaLTP4的过表达导致倍半萜内酯(青蒿素B、青蒿素、双氢青蒿酸和青蒿酸)的产量增加。本研究表明，LTP基因(AaLTP3或AaLTP4)在青蒿GT中的脂类物质的截留和分泌过程中起着重要的作用，有助于通过基因工程提高倍半萜内酯的产量。
In Artemisia annua plants, glandular trichomes (GTs) are responsible for the biosynthesis and secretion of sesquiterpene lactones including artemisinin/arteannuin B. Nonspecific lipid transfer proteins (LTPs) in plants bind and carry lipidmolecules across the cell membrane and are also known as secretary proteins. Interestingly, the transcripts of LTP genes are exceptionally abundant in the GTs of A. annua. In the present study, we isolated two trichome-specific LTP genes (AaLTP3 and AaLTP4) from a Korean ecotype of A. annua. AaLTP3 was expressed abundantly in shoots, whereas AaLTP4 was expressed in flowers. The GUS signal driven by the AaLTP3 or AaLTP4 promoter in transgenic A. annua plants revealed that the AaLTP3 promoter was active on hair-like non-GTs and that the AaLTP4 promoter was active on GTs. Analysis of enhanced cyan fluorescent protein (ECFP) fluorescence fused with the AaLTP3 or AaLTP4 protein in transgenic tobacco revealed that ECFP florescence was very bright on secreted lipids of long GTs. Moreover, the florescence was also bright on the head cells of short trichomes and their secreted granules. Immunoblotting analysis of GT exudates in petioles of A. annua revealed a strong positive signal against the AaLTP4 antibody. Overexpression of AaLTP3 or AaLTP4 in transgenic A. annua plants resulted in enhanced production of sesquiterpene lactones (arteannuin B, artemisinin, dihydroartemisinic acid and artemisinic acid) compared with those of wild type. The present study shows that LTP genes (AaLTP3 or AaLTP4) play important roles in the sequestration and secretion of lipids in GTs of A. annua, which is useful for the enhanced production of sesquiterpene lactones by genetic engineering.