荒漠盐生植物盐穗木两个microRNA与靶标基因的耐盐抗旱功能研究

MicroRNAs(miRNAs) are an extensive class of small non-protein-coding endogenous RNA in animals and plants, which negatively regulate gene expression at the post-transcriptional levels by degrading mRNA or repressing mRNA translation, depending on the complementarities between a miRNA and its targeted mRNAs. Plant miRNAs play an important role in plant growth and development as well as response to environmental stresses. Halostachys caspica is just a kind of highly salt-tolerant short shrub that belongs to Chenopodiaceae in Xinjiang’s extremely saline-alkaline and arid environments. We choose two differentially expressed miRNAs (miR171b and miR169b) from H. caspica small RNA libraries to further carry out salt- and drought- tolerant research according to the data of the analysis of small RNA libraries (control and high salt treatment), H. caspica transcriptome and the expression of some candidate miRNAs and predicted target genes by real-time PCR with H. caspica assimilating branches treated with 600 mM NaCl stress. Through the correlation analysis of the two miRNAs and target genes and the molecular identification and physiological detection of transgenic Arabidopsis transformed H. caspica miR171b, 169b and their corresponding target genes, respectively, salt- and drought-tolerant physiological function on the two miRNAs and target genes can be confirmed; the regulation of the two miRNAs to target genes under salt stress can be uncovered primarily and there is a good perspective to improve salt and drought tolerance in crops and woods by microRNA- and target gene- based biotechnology, and finally to promote the development of local economy and the building of ecological environment.

microRNA(miRNA)是动植物中广泛存在的内源性非编码小RNA分子，在转录后水平负调控靶标基因的表达。miRNA在植物的生长发育和胁迫响应方面发挥着重要作用。盐穗木(Halostachys caspica)是新疆干旱盐碱荒漠区广布的一种极端耐盐的藜科盐生植物。我们结合本物种高盐（600 mM NaCl）胁迫下的小RNA文库分析、盐穗木转录组和候选的miRNA与预测的靶向基因在盐胁迫下的real-time PCR表达数据，选择了两个表达差异显著的盐穗木miR171b和miR169b做后续耐盐抗旱性研究。通过miRNA和靶基因的相关性分析及转基因植株的分子和生理检测，明确这两种miRNA和靶标基因的耐盐抗旱生理功能；初步地揭示这两种miRNA对靶标基因的调控；期望利用miRNA或者靶基因依据的生物技术提高作物和林木的非生物抗性，进而推动地方经济的发展和生态环境的建设。

盐穗木miR169b和miR171b是从高盐（600 mM NaCl）胁迫下的该物种根的小RNA文库中候选的两个差异表达的microRNA，预测它们的靶基因为NFYA1和SCL6。本项目鉴定盐穗木HcmiR169b/HcNFYA1和HcmiR171b/HcSCL6在盐旱胁迫下功能和初步调控机制。主要研究结果为：HcmiR169b/HcNFYA1在盐穗木非生物胁迫的表达呈现明显的负相关性；HcmiR169b在mRNA水平切割HcNFYA1；HcmiR169b的转基因拟南芥表现盐旱胁迫的敏感，而HcNFYA1赋予拟南芥盐旱胁迫的抗性；HcmiR169b/HcNFYA1通过ABA合成和信号转导及胁迫响应基因的表达参与盐旱胁迫的抗性。HcmiR171b对HcSCL6在转录水平切割，在盐穗木中也受到盐旱胁迫的诱导，无论是种萌期、幼苗期还是成苗期，HcmiR171b都降低了转基因拟南芥盐旱胁迫的抗性。另一对盐穗木HcmiR172e/HcTOE3也响应植物的非生物胁迫并具有靶向性。HcmiR172e的转基因拟南芥表现盐、旱、低温胁迫的敏感，而HcTOE3赋予拟南芥盐、旱和冰冻胁迫的抗性。该项成果丰富盐穗木microRNA与靶基因参与非生物胁迫的研究进展，具有非生物胁迫抗性的基因有望提高作物的抗性。