Key genes during ethylene-induced adventitious root development in cucumber (Cucumis sativus L.). Key genes during ethylene-induced adventitious root development in cucumber (Cucumis sativus L.)

Our previous studies have shown that exogenous ethylene (ETH) may induce plant adventitious root development in cucumber. In this study, transcriptome technique was used to explore the key genes in ETH-induced rooting. The results revealed that ETH regulated 1415 diferentially expressed genes (DEGs) during rooting, among which 687 DEGs were up-regulated and 728 DEGs were down-regulated. According to Kyoto encyclopedia of genes and genomes (KEGG) pathway analysis, the critical pathways involved in ETH-induced adventitious root development were selected for further study, including carbon metabolism [starch and sucrose metabolism, glycolysis / gluconeogenesis, citrate cycle (TCA cycle), oxidative phosphorylation, fatty acid biosynthesis and fatty acid degradation], secondary metabolite biosynthesis (phenylalanine metabolism and flavonoid biosynthesis) and plant hormone signal transduction. In carbon metabolism, ETH reduced t the expression of CsHK2, CsPK2 and CsCYP86A1, whereas enhanced the expression of CsBAM1 and CsBAM3. Moreover, ETH negatively regulated the transcript level of CsPAL and CsF3’M and positively mediated that of CsPAO in secondary metabolite biosynthesis pathway. Additionally, ETH could induce adventitious rooting by negatively regulating auxin and ETH signal transduction-related genes (CsLAX5, CsGH3.17, CsSUAR50 and CsERS) and positively regulating ABA and BR signaling transduction-related genes (CsPYL1, CsPYL5, CsPYL8, CsBAK1 and CsXTH3) . Furthermore, the results of real-time PCR about the mRNA levels of these genes were consistent with transcriptome results. Therefore, ETH may induce adventitious root development by regulating carbon metabolism-related genes, secondary metabolite biosynthesis-related genes and plant hormone signal transduction-related genes. Overall design: Cucumber explants at 48 h after the control (distilled water) and ethylene donor (0.5 μM ethrel) tratments were collected and stored at −80 °C until being used, in triplicate, and RNA was extracted and sequenced by paired-end sequencing, using Illumina.

本课题组前期研究证实，外源乙烯（exogenous ethylene, ETH）可诱导黄瓜不定根的发育。本研究采用转录组测序技术，探究ETH诱导黄瓜不定根形成过程中的关键基因。研究结果显示，在不定根形成阶段，ETH共调控了1415个差异表达基因（differentially expressed genes, DEGs），其中687个基因呈上调表达，728个基因呈下调表达。基于京都基因与基因组百科全书（Kyoto Encyclopedia of Genes and Genomes, KEGG）通路富集分析，本研究筛选出ETH诱导黄瓜不定根发育所涉及的关键通路开展后续研究，涵盖碳代谢通路[淀粉与蔗糖代谢、糖酵解/糖异生、三羧酸循环（citrate cycle, TCA cycle）、氧化磷酸化、脂肪酸生物合成与脂肪酸降解]、次生代谢产物生物合成通路（苯丙氨酸代谢与黄酮类生物合成）以及植物激素信号转导通路。在碳代谢通路中，ETH可下调CsHK2、CsPK2及CsCYP86A1的基因表达水平，而上调CsBAM1与CsBAM3的表达量。此外，在次生代谢产物生物合成通路中，ETH对CsPAL与CsF3’M的转录水平呈负调控作用，对CsPAO的转录水平则呈正调控作用。进一步研究发现，ETH可通过负调控生长素与ETH信号转导相关基因（CsLAX5、CsGH3.17、CsSUAR50及CsERS），同时正调控脱落酸（abscisic acid, ABA）与油菜素内酯（brassinosteroid, BR）信号转导相关基因（CsPYL1、CsPYL5、CsPYL8、CsBAK1及CsXTH3），从而诱导不定根形成。针对上述基因mRNA水平的实时荧光定量PCR（real-time PCR）检测结果与转录组分析结果高度吻合。综上，ETH或可通过调控碳代谢相关基因、次生代谢产物生物合成相关基因以及植物激素信号转导相关基因，诱导黄瓜不定根的发育。实验设计：收集经蒸馏水（对照组）与乙烯供体试剂（0.5 μM 乙烯利）处理48小时后的黄瓜外植体，设置3次生物学重复，样品于-80℃低温保存待用；随后提取总RNA，采用Illumina平台进行双端测序。