Gong Ji-Ming, Lee David A, Schroeder Julian I
Division of Biological Sciences and Center for Molecular Genetics, Cell and Developmental Biology Section, University of California at San Diego, La Jolla, CA 92093-0116, USA.
Proc Natl Acad Sci U S A. 2003 Aug 19;100(17):10118-23. doi: 10.1073/pnas.1734072100. Epub 2003 Aug 8.
Phytochelatin synthases (PCS) mediate cellular heavy-metal resistance in plants, fungi, and worms. However, phytochelatins (PCs) are generally considered to function as intracellular heavy-metal detoxification mechanisms, and whether long-distance transport of PCs occurs during heavy-metal detoxification remains unknown. Here, wheat TaPCS1 cDNA expression was either targeted to Arabidopsis roots with the Arabidopsis alcohol dehydrogenase (Adh) promoter (Adh::TaPCS1/cad1-3) or ectopically expressed with the cauliflower mosaic virus 35S promoter (35S::TaPCS1/cad1-3) in the PC-deficient mutant cad1-3. Adh::TaPCS1/cad1-3 and 35S::TaPCS1/cad1-3 complemented the cadmium, mercury, and arsenic sensitivities of the cad1-3 mutant. Northern blot, RT-PCR, and Western blot analyses showed Adh promoter-driven TaPCS1 expression only in roots and thus demonstrated lack of long-distance TaPCS1 mRNA and protein transport in plants. Fluorescence HPLC analyses showed that under Cd2+ stress, no PCs were detectable in cad1-3. However, in Adh::TaPCS1/cad1-3 plants, PCs were detected in roots and in rosette leaves and stems. Inductively coupled plasma atomic emission spectrometer analyses showed that either root-specific or ectopic expression of TaPCS1 significantly enhanced long-distance Cd2+ transport into stems and rosette leaves. Unexpectedly, transgenic expression of TaPCS1 reduced Cd2+ accumulation in roots compared with cad1-3. The reduced Cd2+ accumulation in roots and enhanced root-to-shoot Cd2+ transport in transgenic plants were abrogated by l-buthionine sulfoximine. The presented findings show that (i) transgenic expression of TaPCS1 suppresses the heavy-metal sensitivity of cad1-3, (ii) PCs can be transported from roots to shoots, and (iii) transgenic expression of the TaPCS1 gene increases long-distance root-to-shoot Cd2+ transport and reduces Cd2+ accumulation in roots.
植物螯合肽合酶（PCS）介导植物、真菌和蠕虫细胞对重金属的抗性。然而，植物螯合肽（PCs）通常被认为是细胞内重金属解毒机制，在重金属解毒过程中PCs是否发生长距离运输尚不清楚。在这里，小麦TaPCS1 cDNA的表达要么通过拟南芥乙醇脱氢酶（Adh）启动子靶向拟南芥根（Adh::TaPCS1/cad1-3），要么在PC缺陷型突变体cad1-3中用花椰菜花叶病毒35S启动子（35S::TaPCS1/cad1-3）异位表达。Adh::TaPCS1/cad1-3和35S::TaPCS1/cad1-3互补了cad1-3突变体对镉、汞和砷的敏感性。Northern印迹、RT-PCR和Western印迹分析表明，Adh启动子驱动的TaPCS1仅在根中表达，因此证明了植物中不存在TaPCS1 mRNA和蛋白质的长距离运输。荧光HPLC分析表明，在Cd2+胁迫下，在cad1-3中未检测到PCs。然而，在Adh::TaPCS1/cad1-3植物中，在根、莲座叶和茎中检测到了PCs。电感耦合等离子体原子发射光谱仪分析表明，TaPCS1的根特异性表达或异位表达显著增强了Cd2+向茎和莲座叶的长距离运输。出乎意料的是，与cad1-3相比，TaPCS1的转基因表达降低了根中Cd2+的积累。转基因植物中根中Cd2+积累的减少和根到地上部Cd2+运输的增强被L-丁硫氨酸亚砜胺消除。所呈现的研究结果表明：（i）TaPCS1的转基因表达抑制了cad1-3对重金属的敏感性；（ii）PCs可以从根运输到地上部；（iii）TaPCS1基因的转基因表达增加了根到地上部长距离的Cd2+运输，并减少了根中Cd2+的积累。
