Data from: Effects of overexpressing a native gene encoding 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS) on glyphosate resistance in Arabidopsis thaliana

Widespread overuse of the herbicide glyphosate, the active ingredient in RoundUp®, has led to the evolution of glyphosate-resistant weed biotypes, some of which persist by overproducing the herbicide's target enzyme, 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS). EPSPS is a key enzyme in the shikimic acid pathway for biosynthesis of aromatic amino acids, lignin, and defensive compounds, but little is known about how overproducing EPSPS affects downstream metabolites, growth, or lifetime fitness in the absence of glyphosate. We are using Arabidopsis as a model system for investigating phenotypic effects of overproducing EPSPS, thereby avoiding confounding effects of genetic background or other mechanisms of herbicide resistance in agricultural weeds. Here, we report results from the first stage of this project. We designed a binary vector expressing a native EPSPS gene from Arabidopsis under control of the CaMV35S promoter (labelled OX, for over-expression). For both OX and the empty vector (labelled EV), we obtained nine independent T3 lines. Subsets of these lines were used to characterize glyphosate resistance in greenhouse experiments. Seven of the nine OX lines exhibited enhanced glyphosate resistance when compared to EV and wild-type control lines, and one of these was discarded due to severe deformities. The remaining six OX lines exhibited enhanced EPSPS gene expression and glyphosate resistance compared to controls. Glyphosate resistance was correlated with the degree of EPSPS over-expression for both vegetative and flowering plants, indicating that glyphosate resistance can be used as a surrogate for EPSPS expression levels in this system. These findings set the stage for examination of the effects of EPSPS over-expression on fitness-related traits in the absence of glyphosate. We invite other investigators to contact us if they wish to study gene expression, downstream metabolic effects, and other questions with these particular lines.除草剂草甘膦（glyphosate）是农达（RoundUp®）的有效成分，其广泛滥用已促使抗草甘膦杂草生物型演化产生；其中部分抗性杂草通过过量合成该除草剂的靶标酶——5-烯醇丙酮酰莽草酸-3-磷酸合酶（5-enolpyruvylshikimate-3-phosphate synthase，EPSPS）来维持生存。EPSPS是莽草酸途径中合成芳香族氨基酸、木质素以及防御性化合物的关键酶，但目前学界对无草甘膦胁迫时，过量合成EPSPS如何影响下游代谢物、植株生长乃至终生适合度的机制仍知之甚少。本研究以拟南芥（Arabidopsis）作为模式系统，探究过量合成EPSPS带来的表型效应，以此规避农业杂草中遗传背景差异或其他抗除草剂机制所带来的混淆效应。本文报道该项目第一阶段的研究结果。我们构建了一个双元载体，在花椰菜花叶病毒35S启动子（CaMV35S promoter）的调控下表达拟南芥内源EPSPS基因（该过表达株系标记为OX，即over-expression的缩写）。我们分别获得了OX株系与空载体（empty vector，标记为EV）的9个独立T3代株系。选取其中部分株系开展温室试验，以鉴定其草甘膦抗性水平。9个OX株系中，有7个相较于EV株系与野生型对照株系表现出更强的草甘膦抗性，但其中1个因存在严重畸形被排除。剩余的6个OX株系相较于对照组，均表现出EPSPS基因表达量上调以及草甘膦抗性增强。无论是营养生长期还是开花期植株，草甘膦抗性均与EPSPS过量表达程度呈正相关，这表明在本研究系统中，可通过草甘膦抗性作为EPSPS表达水平的替代检测指标。本研究结果为后续探究无草甘膦胁迫下，EPSPS过量表达对适合度相关性状的影响奠定了基础。若其他研究者希望利用本研究的这批株系开展基因表达、下游代谢效应等相关研究，可与我们联系。