Characterization of miRNAs in Response to Short-Term Waterlogging in Three Inbred Lines of Zea mays

Waterlogging of plants leads to low oxygen levels (hypoxia) in the roots and causes a metabolic switch from aerobic respiration to anaerobic fermentation that results in rapid changes in gene transcription and protein synthesis. Our research seeks to characterize the microRNA-mediated gene regulatory networks associated with short-term waterlogging. MicroRNAs (miRNAs) are small non-coding RNAs that regulate many genes involved in growth, development and various biotic and abiotic stress responses. To characterize the involvement of miRNAs and their targets in response to short-term hypoxia conditions, a quantitative real time PCR (qRT-PCR) assay was used to quantify the expression of the 24 candidate mature miRNA signatures (22 known and 2 novel mature miRNAs, representing 66 miRNA loci) and their 92 predicted targets in three inbred Zea mays lines (waterlogging tolerant Hz32, mid-tolerant B73, and sensitive Mo17). Based on our studies, miR159, miR164, miR167, miR393, miR408 and miR528, which are mainly involved in root development and stress responses, were found to be key regulators in the post-transcriptional regulatory mechanisms under short-term waterlogging conditions in three inbred lines. Further, computational approaches were used to predict the stress and development related cis-regulatory elements on the promoters of these miRNAs; and a probable miRNA-mediated gene regulatory network in response to short-term waterlogging stress was constructed. The differential expression patterns of miRNAs and their targets in these three inbred lines suggest that the miRNAs are active participants in the signal transduction at the early stage of hypoxia conditions via a gene regulatory network; and crosstalk occurs between different biochemical pathways.植物涝害会导致根部缺氧（hypoxia），引发代谢从有氧呼吸向无氧发酵转变，进而使基因转录与蛋白质合成发生快速变化。本研究旨在表征与短期涝害相关的microRNA（miRNA）介导的基因调控网络。MicroRNA（miRNA）是一类小型非编码RNA，可调控诸多参与生长发育、各类生物及非生物胁迫响应的基因。为明确miRNA及其靶标在响应短期低氧胁迫中的作用，我们采用实时定量聚合酶链式反应（quantitative real time PCR，qRT-PCR）技术，对3个玉米（Zea mays）自交系（耐涝系Hz32、中间耐受系B73以及敏感系Mo17）中的24个候选成熟miRNA标记（包含22个已知成熟miRNA与2个新发现的成熟miRNA，对应66个miRNA基因座）及其92个预测靶标的表达水平进行了定量检测。基于本研究结果，在这3个自交系中，主要参与根系发育与胁迫响应的miR159、miR164、miR167、miR393、miR408及miR528被证实是短期涝害条件下转录后调控机制的关键调控因子。进一步，我们通过计算预测方法分析了这些miRNA启动子区域中与胁迫及发育相关的顺式调控元件，并构建了一条响应短期涝害胁迫的潜在miRNA介导基因调控网络。这3个自交系中miRNA及其靶标的差异表达模式表明，miRNA通过基因调控网络在低氧胁迫早期的信号转导过程中发挥活跃作用，且不同生化通路之间存在交叉对话。