Chilling is a major abiotic stress harming rice development and productivity. The C-REPEAT BINDING FACTOR (CBF)-dependent transcriptional regulatory pathway plays a central role in cold stress and acclimation in Arabidopsis. In rice, several genes have been reported in conferring chilling tolerance, however, the chilling signaling in rice remains largely unknown. Here, we report the chilling-induced OSMOTIC STRESS/ABA-ACTIVATED PROTEIN KINASE 6 (OsSAPK6)-IDEAL PLANT ARCHITECTURE 1 (IPA1)-OsCBF3 signal pathway in rice. Under chilling stress, OsSAPK6 could phosphorylate IPA1 and increase its stability. In turn, IPA1 could directly bind to the GTAC motif on the OsCBF3 promoter to elevate its expression. Genetic evidence showed that OsSAPK6, IPA1 and OsCBF3 were all positive regulators of rice chilling tolerance. The function of OsSAPK6 in chilling tolerance depended on IPA1, and overexpression of OsCBF3 could rescue the chilling-sensitive phenotype of ipa1 loss-of-function mutant. Moreover, the natural gain-of-function allele ipa1-2D could simultaneously enhance seedling chilling tolerance and increase grain yield. Taken together, our results revealed a chilling-induced OsSAPK6-IPA1-OsCBF signal cascade in rice, which shed new lights on chilling stress-tolerant rice breeding.
水稻苗期遇到低溫天氣后，稻苗會(huì)發(fā)生生長遲緩、黃化、發(fā)育不良、枯萎甚至死亡，嚴(yán)重影響水稻生長發(fā)育與產(chǎn)量。提高水稻苗期的冷脅迫抗性，對(duì)于減少水稻苗期冷害損失、推廣水稻直播種植等均具有重要意義，然而目前對(duì)于水稻冷脅迫后的轉(zhuǎn)錄調(diào)控響應(yīng)機(jī)制仍不清楚。
中國科學(xué)院遺傳與發(fā)育生物學(xué)研究所李家洋團(tuán)隊(duì)在水稻冷脅迫響應(yīng)調(diào)控機(jī)制上取得了新進(jìn)展。該團(tuán)隊(duì)在此前構(gòu)建的水稻突變體庫中發(fā)現(xiàn)一個(gè)苗期冷敏感的突變體，并鑒定到是由于OSMOTIC STRESS/ABA-ACTIVATED PROTEIN KINASE 6（OsSAPK6）的提前終止導(dǎo)致水稻冷敏感的表型，同時(shí)OsSAPK6的過表達(dá)材料表現(xiàn)出了苗期抗冷的表型。為鑒定OsSAPK6的底物，利用GFP-OsSAPK6進(jìn)行了免疫沉淀-質(zhì)譜聯(lián)用(IP-MS)實(shí)驗(yàn)，發(fā)現(xiàn)IPA1（Ideal Plant Architecture 1）在候選蛋白中多次出現(xiàn)。IPA1是此前李家洋團(tuán)隊(duì)鑒定到的一個(gè)水稻株型調(diào)控的主效基因（Jiao et al., 2010）。該研究發(fā)現(xiàn)OsSAPK6能夠與IPA1發(fā)生蛋白互作，并在S201和S213位點(diǎn)對(duì)IPA1進(jìn)行磷酸化修飾，進(jìn)而穩(wěn)定IPA1蛋白。隨后的研究表明，IPA1的S213位點(diǎn)的磷酸化對(duì)于其抗冷的表型具有關(guān)鍵作用，將213位點(diǎn)的絲氨酸突變成無法被磷酸化的天冬酰胺后，ipa1S213N突變體表現(xiàn)出了冷敏感的表型。通過RNA-seq發(fā)現(xiàn)，OsCBF3基因在ipa1-3D功能獲得突變體中的冷誘導(dǎo)表達(dá)明顯增強(qiáng)，而在ipa1-10功能喪失突變體中冷誘導(dǎo)表達(dá)明顯減弱，且IPA1能夠直接結(jié)合OsCBF3啟動(dòng)子區(qū)的GTAC motif激活其基因表達(dá)。遺傳與生化實(shí)驗(yàn)的結(jié)果表明，OsSAPK6、IPA1與OsCBF3在同一途徑中調(diào)控水稻的冷響應(yīng)，冷脅迫后OsSAPK6能夠磷酸化IPA1蛋白，使得IPA1蛋白積累并激活下游OsCBF3的表達(dá)，增強(qiáng)水稻冷脅迫抗性。ipa1-2D是此前鑒定到的一個(gè)IPA1優(yōu)異等位基因型，在高產(chǎn)水稻品種培育中被廣泛使用（Zhang et al., 2017）。研究表明，攜帶ipa1-2D的近等基因系與對(duì)照相比，具有更強(qiáng)的苗期耐冷性，兼具高產(chǎn)與抗冷。綜上，該項(xiàng)成果闡明了OsSAPK6-IPA1-OsCBF調(diào)控水稻冷脅迫響應(yīng)的分子機(jī)制，為創(chuàng)制高產(chǎn)耐冷水稻品種提供了新的分子機(jī)制與遺傳資源。