華中農(nóng)業(yè)大學(xué)學(xué)者揭示轉(zhuǎn)錄因子OsWRKY37調(diào)控銅吸收運輸影響水稻開花結(jié)實的分子機制

　　4月9日，華中農(nóng)業(yè)大學(xué)植物營養(yǎng)生物學(xué)團隊在Plant Physiology上發(fā)表了題為“OsWRKY37 acts as a positive regulator for flowering time and grain fertility in rice under copper deficient condition”的研究論文，揭示了缺銅條件下轉(zhuǎn)錄因子OsWRKY37調(diào)控銅吸收運輸影響水稻開花結(jié)實的分子機制。

 

　　銅是植物生長發(fā)育所必需的微量營養(yǎng)元素，也是對水稻花器官發(fā)育和產(chǎn)量形成影響最大的微量元素；缺銅易導(dǎo)致水稻開花期推遲、花藥形成受阻、花粉活性下降、結(jié)實率降低、無效分蘗增加，從而嚴重影響水稻籽粒產(chǎn)量。近年來，我國高強度、集約化土地利用方式及作物復(fù)種比例的增加，加劇了土壤養(yǎng)分的攜出量，降低了土壤肥力水平；同時，由于耕作過程中忽視微量元素肥料施用，引起土壤養(yǎng)分比例失衡，極易導(dǎo)致耕地銅缺乏，從而影響水稻正常的生殖生長及產(chǎn)量形成。隨著科學(xué)研究的不斷探索以及對水稻基因功能的逐步解析，參與水稻銅吸收和運輸過程的重要基因特別是轉(zhuǎn)運基因不斷被克隆和鑒定，在水稻中逐漸形成了一個清晰的銅運輸路徑，然而該過程的上游調(diào)控因子及其具體的調(diào)控網(wǎng)絡(luò)尚不清楚。

 

　　課題組前期對不同銅水平處理下的水稻根系進行轉(zhuǎn)錄組分析，獲得一個受缺銅誘導(dǎo)表達功能未知的WRKY轉(zhuǎn)錄因子OsWRKY37。熒光定量PCR和ProOsWRKY37:GUS轉(zhuǎn)基因植株分析結(jié)果顯示，OsWRKY37主要在根、葉片、葉枕、節(jié)的維管組織、新生小穗以及花藥中表達。缺銅條件下，超量表達OsWRKY37促進水稻根系對銅的吸收以及銅從根向地上部的轉(zhuǎn)運；而敲除突變OsWRKY37抑制銅從根向地上部的轉(zhuǎn)運以及向小穗的分配，顯著降低雄蕊中的銅含量，影響花粉發(fā)育；敲除突變體表現(xiàn)為開花期推遲、育性降低、籽粒產(chǎn)量下降。酵母單雜交、煙草瞬時表達、EMSA和ChIP-qPCR等試驗證明，OsWRKY37可與銅轉(zhuǎn)運基因OsCOPT6及OsYSL16啟動子中的W-box相互結(jié)合，并上調(diào)其表達水平?？偠灾便~誘導(dǎo)OsWRKY37表達，OsWRKY37可與OsCOPT6和OsYSL16的啟動子相互結(jié)合，正向調(diào)控其表達水平，促進水稻根系對銅的吸收以及銅從根向地上部和小穗中轉(zhuǎn)運，有利于花器官正常發(fā)育及籽粒產(chǎn)量形成。

 

　　我校資源與環(huán)境學(xué)院已畢業(yè)博士研究生姬晨晨為第一作者，蔡紅梅副教授為通訊作者，已畢業(yè)博士研究生丁靜麗和已畢業(yè)碩士研究生李海星、郁露參與了該研究，徐芳森教授、石磊教授、姜存?zhèn)}教授、王創(chuàng)教授、丁廣大教授和汪社亮副教授對該研究的試驗設(shè)計和數(shù)據(jù)分析提供了寶貴的建議。本研究得到國家重點研發(fā)計劃項目資助。

 

　　【英文摘要】

 

　　Efficient uptake, translocation, and distribution of Cu to rice （Oryza Sativa） spikelets is crucial for flowering and yield production. However, the regulatory factors involved in this process remain unidentified. In this study, we isolated a WRKY transcription factor gene induced by Cu deficiency, OsWRKY37, and characterized its regulatory role in Cu uptake and transport in rice. OsWRKY37 was highly expressed in rice roots, nodes, leaf vascular bundles, and anthers. Overexpression of OsWRKY37 promoted the uptake and root-to-shoot translocation of Cu in rice under -Cu condition but not under +Cu condition. While mutation of OsWRKY37 significantly decreased Cu concentrations in the stamen, the root-to-shoot translocation and distribution ratio in brown rice affected pollen development, delayed flowering time, decreased fertility, and reduced grain yield under -Cu condition. Y1H, transient co-expression and EMSAs, together with in situ RT-PCR and RT-qPCR analysis, showed that OsWRKY37 could directly bind to the upstream promoter region of OsCOPT6 （copper transporter） and OsYSL16 （yellow stripe-like protein） and positively activate their expression levels. Analyses of oscopt6 mutants further validated its important role in Cu uptake in rice. Our study demonstrated that OsWRKY37 acts as a positive regulator involved in the uptake, root-to-shoot translocation, and distribution of Cu through activating the expression of OsCOPT6 and OsYSL16, which is important for pollen development, flowering, fertility, and grain yield in rice under Cu deficient conditions. Our results provide a genetic strategy for improving rice yield under Cu deficient condition.