花生AhCLE基因家族鉴定及对土壤紧实和氮素复合胁迫响应分析

Identification of the AhCLE Gene Family in Peanut and Its Response to Complex Stresses of Soil Compaction and Nitrogen

花生是我国重要的油料作物,其生长过程面临多种土壤环境逆境胁迫。CLE(CLAVATA3/Embryo Surrounding Region)多肽是目前研究最深入的植物多肽(多肽激素),具有类似传统植物激素的调节功能,广泛参与植物生长发育及抗逆过程。目前关于花生CLE(AhCLE)家族基因鉴定及抗逆功能的研究尚未见报导。本研究基于花生基因组注释信息,系统鉴定AhCLE家族成员并对其进行基因亚细胞定位、染色体定位、多序列比对、基因保守基序、基因结构及系统进化等分析;利用转录组信息结合实时荧光定量PCR技术(qRT-PCR)检测AhCLE基因家族各成员在花生不同组织部位及不同的土壤紧实及氮素胁迫条件下的表达情况。研究结果表明:AhCLE基因家族由9个成员组成,均含有12个氨基酸组成的CLE保守结构域序列;亚细胞定位预测结果显示该基因家族成员均位于细胞外,为分泌型蛋白;启动子区顺式作用元件分析发现,该家族成员具有多个逆境胁迫及植物激素响应元件,表明AhCLE家族可能与多种植物激素协同参与调控植物逆境胁迫;聚类分析发现,该家族成员分布于4个不同的分支中,分别与拟南芥的AtCLE家族不同成员聚在一起,表明AhCLE成员间可能具有不同的生物学功能。组织表达分析显示,AhCLE7,AhCLE8主要在茎、叶中表达,AhCLE6主要在根和根瘤中表达,而AhCLE9主要在花中表达,其余成员在上述组织中也呈现不同的组织特异性,体现了不同成员可能在特定的组织中发挥特定的功能。在土壤氮素含量正常条件下,土壤紧实会抑制AhCLE1,AhCLE2和AhCLE3的表达。在正常土壤紧实度条件下,AhCLE1和AhCLE2在低氮条件下抑制表达,而AhCLE9则在低氮条件下被诱导表达。AhCLE1与AhCLE2能够同时响应土壤紧实胁迫及氮素胁迫,推测在花生根系发育和氮素吸收过程中可能发挥重要作用。本研究为进一步发掘抗逆型CLE多肽及其功能研究提供了理论基础。

Peanut is an important oilseed crop in China,and its growth process is exposed to a variety of soil environmental adversity stresses.CLE(CLAVATA3/Embryo Surrounding Region)peptide is the most intensively studied plant peptide(peptide hormone),which has regulatory functions similar to those of traditional plant hormones and is widely involved in plant growth and development and stress resistance processes.Studies on the identification of CLEfamily genes and stress toleran in peanut have not yet been reported.In this study,the members of the peanut CLE(AhCLE)family were systematically identified based on the annotation information of the peanut genome,and the gene subcellular localization,chromosomal localization,multiple sequence alignment,conserved gene motifs,gene structure and phylogeny of AhCLEfamily members were analyzed.The expression patterns of AhCLEfamily members in different peanut tissues,and peanut roots under different soil compacting and nitrogen stress conditions were detected using transcriptome information combined with quantitative real-time fluorescence PCR(qRT-PCR).The results showed that the AhCLEgene family consists of nine members,all of which contain a conserved structural domain sequence of CLE consisting of twelve amino acids.Subcellular localization prediction showed that all members of AhCLEs are located outside the cell and are secreted proteins.The analysis results of cis-acting elements in the promoter region revealed that the members of AhCLEshave multiple response elements to stress and phytohormone,suggesting that the peanut AhCLEfamily may coordinate with various plant hormones to regulate plant stresses.Members of AhCLEs are distributed in four different branches of the phylogenetic tree and clustered with different members of the AtCLEs,indicating that members of AhCLEs may have different biological functions.Expression analysis showed that AhCLE7and AhCLE8were mainly expressed in stems and leaves,AhCLE6was mainly expressed in roots and root nodules,while AhCLE9was mainly expressed in flowers,and the remaining members also showed different tissue specificity in the above tissues,reflecting that different members may perform specific functions in specific tissues.Under normal soil nitrogen content conditions,soil compaction suppressed the expressions of AhCLE1,AhCLE2,and AhCLE3.Under normal soil compactness conditions,the expressions of AhCLE1and AhCLE2were repressed under low nitrogen stress,while the expression of AhCLE9was induced under low nitrogen stress.AhCLE1and AhCLE2were able to respond to both soil compaction and low-nitrogen stresses,and presumably may play an important role in peanut root development and nitrogen uptake.This study provides a theoretical basis for further exploration of resistance-type CLE peptides and their functional studies.