4月9日，華中農(nóng)業(yè)大學(xué)植物營(yíng)養(yǎng)生物學(xué)團(tuán)隊(duì)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(yùn)輸影響水稻開花結(jié)實(shí)的分子機(jī)制。
銅是植物生長(zhǎng)發(fā)育所必需的微量營(yíng)養(yǎng)元素，也是對(duì)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(shí)率降低、無(wú)效分蘗增加，從而嚴(yán)重影響水稻籽粒產(chǎn)量。近年來(lái)，我國(guó)高強(qiáng)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(shí)，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(zhǎng)及產(chǎn)量形成。隨著科學(xué)研究的不斷探索以及對(duì)水稻基因功能的逐步解析，參與水稻銅吸收和運(yùn)輸過程的重要基因特別是轉(zhuǎn)運(yùn)基因不斷被克隆和鑒定，在水稻中逐漸形成了一個(gè)清晰的銅運(yùn)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(duì)不同銅水平處理下的水稻根系進(jìn)行轉(zhuǎn)錄組分析，獲得一個(gè)受缺銅誘導(dǎo)表達(dá)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(dá)。缺銅條件下，超量表達(dá)OsWRKY37促進(jìn)水稻根系對(duì)銅的吸收以及銅從根向地上部的轉(zhuǎn)運(yùn)；而敲除突變OsWRKY37抑制銅從根向地上部的轉(zhuǎn)運(yùn)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(shí)表達(dá)、EMSA和ChIP-qPCR等試驗(yàn)證明，OsWRKY37可與銅轉(zhuǎn)運(yùn)基因OsCOPT6及OsYSL16啟動(dòng)子中的W-box相互結(jié)合，并上調(diào)其表達(dá)水平。總而言之，缺銅誘導(dǎo)OsWRKY37表達(dá)，OsWRKY37可與OsCOPT6和OsYSL16的啟動(dòng)子相互結(jié)合，正向調(diào)控其表達(dá)水平，促進(jìn)水稻根系對(duì)銅的吸收以及銅從根向地上部和小穗中轉(zhuǎn)運(yùn)，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(duì)該研究的試驗(yàn)設(shè)計(jì)和數(shù)據(jù)分析提供了寶貴的建議。本研究得到國(guó)家重點(diǎn)研發(fā)計(jì)劃項(xiàng)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.
日期：2024-05-08
銅是植物生長(zhǎng)發(fā)育所必需的微量營(yíng)養(yǎng)元素，也是對(duì)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(shí)率降低、無(wú)效分蘗增加，從而嚴(yán)重影響水稻籽粒產(chǎn)量。近年來(lái)，我國(guó)高強(qiáng)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(shí)，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(zhǎng)及產(chǎn)量形成。隨著科學(xué)研究的不斷探索以及對(duì)水稻基因功能的逐步解析，參與水稻銅吸收和運(yùn)輸過程的重要基因特別是轉(zhuǎn)運(yùn)基因不斷被克隆和鑒定，在水稻中逐漸形成了一個(gè)清晰的銅運(yùn)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(duì)不同銅水平處理下的水稻根系進(jìn)行轉(zhuǎn)錄組分析，獲得一個(gè)受缺銅誘導(dǎo)表達(dá)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(dá)。缺銅條件下，超量表達(dá)OsWRKY37促進(jìn)水稻根系對(duì)銅的吸收以及銅從根向地上部的轉(zhuǎn)運(yùn)；而敲除突變OsWRKY37抑制銅從根向地上部的轉(zhuǎn)運(yùn)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(shí)表達(dá)、EMSA和ChIP-qPCR等試驗(yàn)證明，OsWRKY37可與銅轉(zhuǎn)運(yùn)基因OsCOPT6及OsYSL16啟動(dòng)子中的W-box相互結(jié)合，并上調(diào)其表達(dá)水平。總而言之，缺銅誘導(dǎo)OsWRKY37表達(dá)，OsWRKY37可與OsCOPT6和OsYSL16的啟動(dòng)子相互結(jié)合，正向調(diào)控其表達(dá)水平，促進(jìn)水稻根系對(duì)銅的吸收以及銅從根向地上部和小穗中轉(zhuǎn)運(yùn)，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(duì)該研究的試驗(yàn)設(shè)計(jì)和數(shù)據(jù)分析提供了寶貴的建議。本研究得到國(guó)家重點(diǎn)研發(fā)計(jì)劃項(xiàng)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.
日期：2024-05-08
