Department of Plant and Environmental Sciences, University of Copenhagen, Thorvaldsensvej 40, 1871, Frederiksberg, Denmark.
Department of Plant and Environmental Sciences, University of Copenhagen, Thorvaldsensvej 40, 1871, Frederiksberg, Denmark.
Plant Physiol Biochem. 2023 Aug;201:107797. doi: 10.1016/j.plaphy.2023.107797. Epub 2023 May 24.
Cucurbitaceae species are used in traditional medicine around the world. Cucurbitacins are highly oxygenated triterpenoids found in Cucurbitaceae species and exhibit potent anticancer activity alone and in combination with other existing chemotherapeutic drugs. Therefore, increasing production of these specialized metabolites is of great relevance. We recently showed that hairy roots of Cucurbita pepo can be used as a platform for metabolic engineering of cucurbitacins to modify their structure and increase their production. To study the changes in cucurbitacin accumulation upon formation of hairy roots, an empty vector (EV) control and Cucurbitacin inducing bHLH transcription factor 1 (CpCUCbH1)-overexpressing hairy roots of C. pepo were compared to untransformed (WT) roots. Whilst CpCUCbH1-overexpression increased production of cucurbitacins I and B by 5-fold, and cucurbitacin E by 3-fold when compared to EV lines, this increase was not significantly different when compared to WT roots. This indicated that Rhizobium rhizogenes transformation lowered the cucurbitacins levels in hairy roots, but that increasing expression of cucurbitacin biosynthetic genes by CpCUCbH1-overexpression restored cucurbitacin production to WT levels. Subsequent metabolomic and RNA-seq analysis indicated that the metabolic profile and transcriptome of hairy roots was significantly changed when compared to WT roots. Interestingly, it was observed that 11% of the differentially expressed genes were transcription factors. It was noteworthy that the majority of transcripts showing highest Pearson correlation coefficients to the Rhizobium rhizogenes genes rolB, rolC and ORF13a, were predicted to be transcription factors. In summary, hairy roots are an excellent platform for metabolic engineering of plant specialized metabolites, but these extensive transcriptome and metabolic profile changes should be considered in subsequent studies.
葫芦科植物在世界各地的传统医学中被使用。葫芦素是在葫芦科植物中发现的高度氧化的三萜类化合物，具有单独和与其他现有化疗药物联合使用的强大抗癌活性。因此，增加这些特殊代谢物的产量具有重要意义。我们最近表明，南瓜的毛状根可作为葫芦素代谢工程的平台，以修饰其结构并增加其产量。为了研究毛状根形成时葫芦素积累的变化，将空载体 (EV) 对照和过表达葫芦素诱导 bHLH 转录因子 1 (CpCUCbH1) 的南瓜毛状根与未转化 (WT) 根进行了比较。虽然与 EV 系相比，CpCUCbH1 的过表达将葫芦素 I 和 B 的产量增加了 5 倍，将葫芦素 E 的产量增加了 3 倍，但与 WT 根相比，这种增加没有显著差异。这表明根瘤农杆菌转化降低了毛状根中的葫芦素水平，但通过 CpCUCbH1 过表达增加葫芦素生物合成基因的表达将葫芦素产量恢复到 WT 水平。随后的代谢组学和 RNA-seq 分析表明，与 WT 根相比，毛状根的代谢谱和转录组发生了显著变化。有趣的是，观察到 11%的差异表达基因是转录因子。值得注意的是，与根瘤农杆菌基因 rolB、rolC 和 ORF13a 表现出最高 Pearson 相关系数的大多数转录本被预测为转录因子。总之，毛状根是植物特殊代谢物代谢工程的优秀平台，但在后续研究中应考虑这些广泛的转录组和代谢谱变化。
