小麦小分子RNA TaMIR167和TaMIR1139应答和抵御低磷逆境的分子机理

Small RNA molecules (miRNAs) play crucial roles on responding and withstanding the low-Pi stress in plants by regulating the target genes' expression via the mechanisms of post-transcriptional modification and translational repression. Our previous studies have elucidated that two miRNAs in wheat (Triticum aestivum L.) referred to TaMIR167 and TaMIR1139 are significantly up-regulated by deficient-Pi, which further results in the down-regulated expressions of TaARF6 (auxin response factor 6), TaARF8 (auxin response factor 8), and TaUPL (Ulp1 protease-like), three target genes of TaMIR167, and TaMYB1 (MYB type transcription factor 1), TaNADH6 (NADH dehydrogenase subunit 6), and TaPT (phosphate/phosphoenolpyruvate translocator protein), three target genes of TaMIR1139. In this current project, based on the previous results obtained from TaMIR167, TaMIR1139 and their interacted target genes, we will systematically dissect the temporal and spatial expression pattern of the tested miRNAs and their target genes under various Pi-supply conditions based on the approaches of qRT-PCR and in situ hybridization. Based on the generated transgenic plants with over- and down-regulated expressions of the tested miRNAs and their target genes by DNA recombinant techniques and further genetic transformation via mediation of Agrobacterium tumefaciens, we will reveal the transcriptional profilings, protein expression patterns, and metabolic profilings regulated by TaMIR167 and TaMIR1139, and their interacted target genes. We will also separately dissect the roles of the tested miRNAs and target genes on plant's response and endurance to low-Pi stress. Moreover, based on comprehensive analysis of the research data, such as classification of the functions of differential expressed genes, proteins and metabolic substances regulated by the tested miRNAs and their interacted target genes, combined with statistical regression analysis and correlation analysis, we will elucidate the molecular networks and corresponding biochemical pathways in plants responding and withstanding to low-Pi stress mediated by the tested wheat miRNAs. The implementation of this current project could be useful to shed light up on the molecular mechanism that plants respond and withstand to low-Pi stress from the small RNA scope. Also, the results from this project could also provide important theoretical reference for generating novel crop germplasms with high-P use efficiency.

小分子RNA（miRNAs）在转录后、翻译水平调节其作用靶基因转录本数量，进而在调控植株应答和抵御低磷逆境中发挥重要作用。申请人前期发现，低磷下小麦小分子RNA TaMIR167和TaMIR1139的表达上调，造成TaMIR167靶基因TaARF6、TaARF8及TaUPL和TaMIR1139靶基因TaMYB1、TaNADH6及TaPT的转录本减少。本项目以TaMIR167和TaMIR1139及其作用靶基因为基础，阐明供试miRNAs及其靶基因应答低磷逆境的时空模式；以上、下调表达TaMIR167和TaMIR1139及其靶基因的转基因植株为材料，全面揭示供试miRNAs及其靶基因调控的转录谱、蛋白谱和代谢谱特征，系统解析供试miRNAs调控小麦应答低磷逆境的分子网络和生化代谢途径。本课题的完成，将从miRNAs角度系统阐明小麦应答和抵御低磷逆境的分子机理，为小麦磷高效遗传改良提供理论依据。

系统揭示miRNA介导植株抵御低磷逆境机理，对于改善作物磷利用效率具有重要意义。本课题对小麦miR167和miR1139介导植株抵御低磷逆境能力进行研究，研究内容和主要结果如下。.1. TaMIR167作用靶基因生长素响应基因ARF6和ARF8，miR167a及其靶基因应答低磷逆境模式相反，表明ARF6和ARF8受到miR167a转录后调节。超表达miR167a和下调表达ARF6及ARF8株系，低磷下植株长势较对照变差，表明该miRNA/靶基因模块在调控植株应答低磷逆境中发挥重要功能。miR167a和靶基因株系低磷下单株干重和磷累积量减少，细胞保护酶活性下降，与其对转录谱和蛋白谱进行较大调控有关。miR167a调控磷转运蛋白基因PT1转录，参与对植株响应低磷逆境的调控。.2. TaZAT8为miR167a调控下游基因，对低磷胁迫明显应答。与对照相比，超表达TaZAT8转化株系植株长势明显改善，含磷量增加，抵御低磷逆境能力增强。这与其调控磷转运蛋白基因PT1和PT2转录有关。TaZAT8显著增强低磷逆境下细胞保护酶活性和特定细胞保护酶基因表达水平。TaZAT8通过调控部分介导生长素转运PIN基因转录，改善低磷逆境下根体建成，增强植株抵御低磷逆境能力。.3. TaMIR11397个靶基因，分别归属于信号转导、激素响应、转运和生化代谢功能类别。靶基因应答低磷特征与TaMIR1139相反，证实各靶基因受到miRNA的转录后调节。超表达TaMIR1139株系低磷逆境下表型、植株干质量和含磷量明显改善、光合能力增强。TaMIR1139对低磷逆境下植株转录谱和蛋白谱具有较大调控效应。该小分子RNA介导植株抵御低磷逆境能力，与期调控特定磷素吸收PT基因转录有关。.4. TabHLH1为TaMIR1139下游基因，对低磷和低氮逆境显著应答。超表达该基因显著增强植株抵御低磷及低氮能力，与对照相比，转化株系植株个体增大，干质量和磷累积量增加，这与其对磷转运蛋白PT1转录调控，进而改善植株低磷逆境下磷素吸收有关。TabHLH1通过调控特定细胞保护酶基因转录，对细胞保护酶活性和活性氧内稳态产生较大调控，参与TaMIR1139介导植株抵御低磷逆境过程。