宋松泉 劉 軍 徐恒恒 張 琪 黃 薈 伍賢進

?

乙烯的生物合成與信號及其對種子萌發(fā)和休眠的調控

宋松泉1,3,*劉 軍2徐恒恒2張 琪2黃 薈3伍賢進3

1中國科學院植物研究所, 北京 100093;2廣東省農(nóng)業(yè)科學院農(nóng)業(yè)生物基因研究中心, 廣東廣州 510640;3懷化學院民族藥用植物資源研究與利用湖南省重點實驗室/ 生物與食品工程學院, 湖南懷化 418008

種子萌發(fā)是一種關鍵的生態(tài)和農(nóng)業(yè)性狀, 由調控種子休眠狀態(tài)和萌發(fā)潛勢的內在和外部信息所決定, 在植物隨后的生長發(fā)育和產(chǎn)量中起著極其重要的作用。休眠是指種子在合適的條件下暫時不能萌發(fā)。乙烯是一種簡單的具有多種功能的氣體植物激素, 在分子、細胞和整體植物水平調節(jié)植物的代謝。在適宜和逆境條件下, 乙烯通過與其他信號分子的相互作用影響植物的行為。本文主要綜述乙烯的生物合成與信號、乙烯在種子萌發(fā)和休眠釋放中的作用以及乙烯與植物激素脫落酸和赤霉素的相互作用; 并提出了需要進一步研究的科學問題, 試圖為解釋乙烯調控種子萌發(fā)與休眠的分子機制提供新的研究思想。

脫落酸; 生物合成與信號; 交叉反應; 乙烯; 赤霉素; 種子萌發(fā)和休眠

種子是可持續(xù)農(nóng)業(yè)和植物生物多樣性必需的重要遺傳傳遞系統(tǒng), 種子的成功萌發(fā)和幼苗建成在農(nóng)業(yè)生產(chǎn)和自然生態(tài)系統(tǒng)中起決定性作用[1-2]。休眠(dormancy)是指在合適的條件下種子暫時不萌發(fā)[3]。在許多種子植物中, 種子休眠是一種適應性特征, 使植物能夠在逆境條件下存活[4]。種子休眠對植物特別是一年生植物的存活是非常重要的, 因為它能確保種子僅僅在環(huán)境條件合適時萌發(fā)[5-6]。與野生物種比較, 大多數(shù)農(nóng)作物品種表現(xiàn)出休眠水平降低, 以及播種后高的出苗率[7-8]。種子休眠特性的不適當喪失引起新鮮成熟種子的迅速萌發(fā), 或者甚至在收獲前萌發(fā)(pre-harvest sprouting), 也稱為胎萌(vivipary), 導致農(nóng)業(yè)生產(chǎn)中產(chǎn)量和質量的巨大損失, 嚴重影響收獲后的種子管理和隨后的產(chǎn)業(yè)利用[9]。

種子萌發(fā)與休眠在對環(huán)境信號的反應中被脫落酸(abscisic acid, ABA)和赤霉素(gibberellin, GA)之間的平衡所調控; 高水平的ABA和低水平的GA引起種子深休眠和出苗率降低, 而低水平的ABA和高水平的GA誘導種子胎萌[1,6,10-12]。此外, 其他植物激素(乙烯、茉莉酸和生長素)也在種子萌發(fā)控制中起作用[13-15], 特別是乙烯通過復雜的信號網(wǎng)絡調節(jié)許多物種的萌發(fā)與休眠[13,16-17]。

乙烯是一種簡單的具有多種功能的氣體植物激素, 在分子、細胞和整體植物水平調節(jié)植物的代謝[18-20]。在適宜和逆境條件下, 乙烯通過與其他信號分子的相互作用影響植物的行為[21-22]。本文主要綜述乙烯的生物合成與信號, 乙烯在種子萌發(fā)和休眠釋放中的作用, 以及乙烯與植物激素ABA和GA的相互作用; 試圖為解釋乙烯調控種子萌發(fā)與休眠釋放的分子機制提供新的研究思想。

1 乙烯的生物合成與信號途徑

1.1 乙烯的生物合成

Arc等[13]提出, 萌發(fā)種子中乙烯的生物合成途徑與植物其他器官相同, 即甲硫氨酸→S-腺苷甲硫氨酸(S-adenosyl-methionine, S-AdoMet)→1-氨基環(huán)丙烷-1-羧酸(1-aminocyclopropane-1-carboxylic acid, ACC)→乙烯。乙烯的作用主要取決于它在細胞中的濃度以及植物組織對它的敏感性[23-24]。Lieberman等[25]最初在一個化學模式系統(tǒng)中發(fā)現(xiàn)甲硫氨酸是乙烯的前體, 乙烯來自甲硫氨酸的C3和C4; 標記的甲硫氨酸能被蘋果()果實組織有效地轉化成為乙烯[26]。這些發(fā)現(xiàn)隨后被其他研究人員用蘋果和其他植物組織證實[27], 然而, 更重要的進展是S-AdoMet和ACC被確定為植物中乙烯合成的前體[27]。甲硫氨酸通過3個關鍵的酶促反應產(chǎn)生乙烯: (1)甲硫氨酸被S-AdoMet合成酶轉化成為S-AdoMet; (2) ACC合酶(ACC synthase, ACS)轉化S-AdoMet成為ACC; (3) ACC氧化酶(ACC oxidase, ACO)分解ACC釋放乙烯(圖1)。ACC的形成通常被認為是乙烯生物合成途徑中的限速步驟[19]。除了ACC以外, ACS也產(chǎn)生5’-甲硫腺苷(5’-methylthioadenosine, MTA), 它被用于新的甲硫氨酸的合成, 確保即使當甲硫氨酸庫變小時, 高速率的乙烯生物合成也能被維持(圖1)。ACO催化ACC轉化成為乙烯是氧依賴的, 在厭氧條件下, 乙烯的形成被完全抑制; 在這個反應中, 還需要Fe2+和抗壞血酸(ascorbic acid, AsA)作為輔因子和共同底物。ACC能被轉化成為丙二酰ACC(malonyl-ACC, MACC), 從而被失活; 從ACC分解形成的有毒氣體氰化氫(HCN)被β-氰丙氨酸合酶(β-cyanoalanine synthase)去毒(圖1)[19]。在N2下, ACC在蘋果組織中積累[27]。

1.1.1 ACC合酶 在種子萌發(fā)過程中, 乙烯的增加與和轉錄本的逐漸積累有關[14,28]。ACS定位于細胞質, 是依賴吡哆醛-5’-磷酸(pyridoxal-5’- phosphate, PLP)酶的成員之一, 它利用維生素b6作為酶功能的輔助因子[19]。在擬南芥()中, ACS由12個成員組成的多基因家族編碼, 其中8個編碼功能性ACC合酶(ACS2、ACS4~ACS9、ACS11),是一個失活的異構體,是一個假基因,和編碼氨基轉移酶[29]。三維結構測定表明, ACS形成功能二聚體; 異源二聚體的形成增加了ACS蛋白家族的結構和功能復雜性[30]。在擬南芥中, 大的基因家族表現(xiàn)出一種組織專一的和差異的表達模式; 利用單個和多個敲除突變體, 證明基因家族的個別成員具有特定的發(fā)育和生理作用, 而且它們之間也存在著復雜的組合相互作用[30]。在許多物種中, 不同的內外信號調節(jié)乙烯生物合成的水平, 在基因表達的水平起作用; 這些誘導因子包括生長素、細胞分裂素、油菜素甾體、乙烯、銅、機械刺激、臭氧、病原體和傷害[19,31]。

根據(jù)C端結構, ACS蛋白分成三種類型。類型I ACS蛋白在它們的C端結構域含有一個假定的鈣依賴蛋白激酶(calcium-dependent protein kinase, CDPK)磷酸化靶位點和3個促分裂原激活的蛋白激酶(mitogen-activated protein kinase, MAPK), 類型II ACS蛋白僅僅含有MAPK磷酸化位點, 而類型Ⅲ ACS蛋白不含任何磷酸化位點[32]。研究表明, 在擬南芥[33-34]和番茄()[35]中, 一些ACS成員的差異磷酸化引起蛋白質通過蛋白酶體(proteasome)降解; 一些ACS成員的蛋白穩(wěn)定性進一步被蛋白磷酸酶2A(protein phosphatase 2A, PP2A)和PP2C所調節(jié)[36-37]; 這些結果表明磷酸化和去磷酸化之間的復雜平衡確保蛋白質的活性和穩(wěn)定性。

圖1 乙烯生物合成途徑

S-腺苷甲硫氨酸(S-AdoMet)合成酶催化從甲硫氨酸形成S-AdoMet, 合成1分子的S-AdoMet消耗1分子的ATP(1)。ACC合酶催化S-AdoMet轉化成為ACC是乙烯合成的限速步驟(2)。隨著ACC的合成, 甲硫腺苷(MTA)是ACC合酶產(chǎn)生的副產(chǎn)物。MTA回到甲硫氨酸的再循環(huán)保存了甲硫基, 能夠維持細胞中恒定的甲硫氨酸濃度。ACC丙二?；饔贸蔀楸?ACC 使ACC庫枯竭并減少乙烯的產(chǎn)生。ACC氧化酶利用ACC作為底物, 催化乙烯合成的最后步驟, 同時產(chǎn)生二氧化碳和氰化物(3)。氰化物被β-氰丙氨酸合酶代謝產(chǎn)生無毒的物質。ACC合酶和ACC氧化酶被同源異構蛋白、發(fā)育和環(huán)境信息的轉錄調節(jié)用虛線箭頭表示。引自Lin等[19]。

The formation of S-adenosyl methionine (S-AdoMet) from methionine is catalysed by S-AdoMet synthetase at the expense of one molecule of ATP per molecule of S-AdoMet synthesized (1). A rate-limiting step of ethylene synthesis is the conversion of S-AdoMet to ACC by ACC synthase (2). Methylthioadenosine (MTA) is the by-product generated, along with ACC, by ACC synthase. Recycling of MTA back to methionine conserves the methylthio group and is able to maintain a constant concentration of methionine in cells. Malonylation of ACC to malonyl-ACC depletes the ACC pool and reduces ethylene production. ACC oxidase catalyses the final step of ethylene synthesis using ACC as substrate and generates carbon dioxide and cyanide (3). Cyanide is metabolized by β-cyanoalanine synthase to produce non-toxic substances. Transcriptional regulation of both ACC synthase and ACC oxidase by homeotic proteins and developmental and environmental cues is indicated by dashed arrows. From Lin et al.[19]

1.1.2 ACC氧化酶 盡管ACS被認為是大多數(shù)植物對非生物和生物脅迫反應中產(chǎn)生乙烯的一個關鍵調節(jié)酶[38], 但是ACO活性已經(jīng)被證明在種子萌發(fā)過程中起重要作用[14,39]。有趣的是, 分離ACO的關鍵環(huán)節(jié)是在提取介質中加入AsA[40]。雖然AsA對蛋白質穩(wěn)定性/活性的確切作用還不清楚, 但已經(jīng)證實AsA通過向活性位點提供一個單電子參與ACC環(huán)的打開[41]。這一催化反應釋放乙烯和氰甲酸根離子(NCCO2)?, 后者被分解成為CO2和氰化物(CN?)[41]。ACO屬于雙加氧酶(dioxygenase)超家族, 需要Fe2+作為輔因子, 重碳酸氫鹽作為激活劑[42-44]。ACO的亞細胞定位目前還不清楚, 一些研究將ACO定位于細胞質; 而另一些研究則將ACO定位于質膜[45-46]。盡管ACO蛋白序列不包含任何預測的跨膜結構域, 但該蛋白仍有可能通過直接(或者間接)相互作用與質膜結合[31]。在擬南芥中, ACO也由一個含有5個成員的多基因家族編碼[ACO1、ACO2、ACO4、At1g12010 (ACO3)和At1g77330 (ACO5)]。擬南芥中的3個基因也是由乙烯自動調控的[47]。Van de Poel 等[48]通過數(shù)學模型研究推測, ACO存在轉錄后和/或者翻譯后調節(jié)機制。

1.2 乙烯信號

在曝露于乙烯氣體下, 暗生長的擬南芥幼苗的“三重反應(triple response)”表型能夠使我們容易識別乙烯不敏感的(ethylene-insensitive)和組成性反應(constitutive-response)的突變體, 這些突變體的克隆和鑒定導致了乙烯信號轉導途徑的線性模型的提出[49-50], 即植物中存在一條復雜的乙烯信號途徑包括正、負反饋調控環(huán), 并特別強調植物如何精細調控的機制(圖2)。

圖2 擬南芥中乙烯信號途徑的最近模型

乙烯由受體蛋白ETR1、ERS1、ETR2、ERS2和EIN4 (綠色表示)感受, 受體是乙烯信號的負調控因子。受體通過它們的GAF結構域(在受體的細胞質區(qū)域用五邊形表示)與其他的受體相互作用, 并在ER膜中形成更高層次的復合物。銅(一種乙烯結合的輔因子, 紅色圓圈)由銅轉運體RAN1 (橙色表示)傳遞給受體。RTE1 (粉紅色)與ETR1相聯(lián)系, 介導受體信號輸出。(A)在乙烯缺乏時, 受體激活CTR1 (黃色)。CTR1通過直接磷酸化EIN2的C-末端(藍色圓圈)使其失活(紫色)。EIN2能夠直接與受體的激酶結構域(在受體的細胞質區(qū)域在五邊形下較大的橢圓)相互作用。EIN2的水平通過26S蛋白酶體(灰色)被F-box蛋白ETP1和ETP2(綠色星狀物)負調控。在細胞核中, 轉錄因子EIN3/EIL1 (紅色)通過蛋白酶體被另外2個F-box蛋白EBF1/2(藍色星狀物)降解。在EIN3/EIL1缺乏時, 乙烯反應基因的轉錄被關閉。(B)在乙烯存在時, 受體與激素結合并失去活性, 依次關閉CTR1。這種失活阻止正調控因子EIN2的磷酸化。EIN2的C-末端被一種未知的機制所剪切, 并移動到細胞核, 在細胞核中使EIN3/EIL1穩(wěn)定和誘導EBF1/2的降解。轉錄因子EIN3/EIL1形成二聚體, 激活乙烯靶基因的表達, 包括F-box基因(深藍色卷曲線, 它產(chǎn)生抑制乙烯途徑活性的負反饋環(huán))或者轉錄因子基因(淡藍色線, 它依次始啟一個轉錄級聯(lián), 導致數(shù)百個乙烯調控基因的活化和抑制)。在乙烯反應基因中有受體基因(綠色線), 它的mRNA被乙烯上調, 以及被翻譯成為一批新的沒有與乙烯結合的受體分子; 這些受體分子然后活化負調控因子CTR1, 從而提供了在不添加乙烯的情況下向下調節(jié)乙烯信號的手段。途徑中的其他調控節(jié)點是核糖核酸外切酶EIN5 (淡橙色, 它控制的mRNA水平)以及F-box蛋白ETP1和ETP2 (綠色星狀物, 在乙烯存在時, 它們被降解, 導致EIN2的穩(wěn)定)。正箭頭和負箭頭分別表示激活和下調這個過程。顏色變淺表示的分子(在‘沒有乙烯’中的EIN3/EIL1, 或者在‘乙烯’中的ETP1/2和EBF1/2)相應于蛋白酶體介導降解的被標記的不穩(wěn)定蛋白。卷曲線表示特定的mRNA, 它們的顏色與相應的蛋白質顏色相一致。引自Merchante等[51]。

Ethylene is perceived by the receptor proteins ETR1, ERS1, ETR2, ERS2, and EIN4 (represented in green), the receptors are negative regulators of ethylene signaling. The receptors interact with other receptors and form higher order complexes in the ER membrane through their GAF domains (represented as pentagons in the receptors’ cytosolic domain). Copper (a cofactor for ethylene binding, red circles) is delivered to the receptors by the copper transporter RAN1 (represented in orange). RTE1 (in pink) is associated with ETR1 and mediates the receptor signal output. (A) In the absence of ethylene, the receptors activate CTR1 (in yellow). CTR1 inactivates EIN2 (in purple) by directly phosphorylating (blue circles) its C-terminal end. EIN2 can directly interact with the kinase domain of the receptors (represented as the larger ovals under the pentagons in the cytosolic domain of the receptors). The levels of EIN2 are negatively regulated by the F-box proteins ETP1 and ETP2 (green star)the 26S proteasome (gray). In the nucleus, the transcription factors EIN3/EIL1 (in red) are being degraded by two other F-box proteins, EBF1/2 (blue star), through the proteasome. In the absence of EIN3/EIL1, transcription of the ethylene response genes is shut off. (B) In the presence of ethylene, the receptors bind the hormone and become inactivated, which in turn, switches off CTR1. This inactivation prevents the phosphorylation of the positive regulator EIN2. The C-terminal end of EIN2 is cleaved off by an unknown mechanism and moves to the nucleus where it stabilizes EIN3/EIL1 and induces degradation of EBF1/2. The transcription factors EIN3/EIL1 dimerize and activate the expression of ethylene target genes, including the F-box gene(dark blue curly line) [which generates a negative feedback loop dampening the activity of the ethylene pathway] or the transcription factor gene(light blue line) [which, in turn, initiates a transcriptional cascade resulting in the activation and repression of hundreds of ethylene-regulated genes]. Among the ethylene responsive genes the receptor gene is(green line), whose mRNA is up-regulated by ethylene and translated into the new batch of ethylene-free receptor molecules which then activate the negative regulator CTR1, thus providing the means of tuning down ethylene signaling in the absence of additional ethylene. Other regulatory nodes in the pathway are the exoribonuclease EIN5 (light orange), which controls the levels ofmRNA, and the F-box proteins ETP1 and ETP2 (green star) that are degraded in the presence of ethylene leading to the stabilization of EIN2. Positive and negative arrows represent activation and down-regulation processes, respectively. Molecules shown in fading colors (EIN3/EIL1 in ‘no ethylene’, or ETP1/2 and EBF1/2 in ‘ethylene’) correspond to unstable proteins targeted to proteasome-mediated degradation. Curly lines indicate specific mRNAs, with their colors matching that of the corresponding proteins. From Merchante et al.[51]

1.2.1 乙烯信號途徑 乙烯信號級聯(lián)從乙烯與受體結合開始, 到轉錄調節(jié)結束。乙烯受體是一個多成員家族, 在擬南芥中由ETR1 (ethylene resistant 1)、ERS1 (ethylene response sensor 1)、ETR2、ERS2和EIN4 (ethylene insensitive 4)組成, 它們與乙烯的結合都具有高的親和力。根據(jù)接受區(qū)域的存在(ETR1、ETR2和EIN4)或者缺乏(ERS1和ERS2), 受體可分為2種類型[51]。這些受體以同源二聚體的形式起作用, 是信號途徑的負調控因子, 在乙烯缺乏時主動抑制乙烯反應[21,51-52]。已經(jīng)證明, 這些受體在乙烯反應的控制中是大量冗余的, 但不同異構體之間具有一些功能特異性[51]。受體主要存在于內質網(wǎng)(endoplasmic reticulum, ER)膜中, 由于乙烯能夠在細胞的水環(huán)境和脂質環(huán)境中自由擴散, 受體的ER定位可能促進與其他細胞成分的相互作用和/或者使信號能夠與其他途徑整合[53]。

根據(jù)系統(tǒng)發(fā)育分析和共有的結構特征, 所有的乙烯受體都有一個模塊化結構, 包括一個負責與乙烯結合的N端跨膜結構域, 一個不同受體類型之間與蛋白質-蛋白質相互作用有關的GAF結構域, 以及一個與途徑下游組分相互作用所需的C端結構域[49,54]。盡管受體的C端具有細菌雙組分組氨酸激酶(two- component histidine kinases)的結構相似性, 但受體的自體激酶活性(autokinase activity)在乙烯反應中僅僅起較小的作用[53]。乙烯受體的基本功能單元是能夠與乙烯結合的同源二聚體。在通過GAF結構域相互作用的同源二聚體中, 能夠發(fā)生更高層級的聯(lián)系, 從而在膜中產(chǎn)生受體簇[55-56]。

由細胞內銅轉運體RAN1 (copper transporter RAN1)提供的銅是乙烯結合和受體功能都需要的[57]。功能喪失突變的植株缺乏乙烯結合活性, 表現(xiàn)出類似于受體功能喪失的突變體的表型; 此外, 用銅螯合劑處理的弱等位基因表現(xiàn)出類似于乙烯處理的野生型植株的表型[58], 以及添加Cu2+到這些植株能部分抑制的表型[57]。這些結果提出, RAN1在乙烯受體的生物發(fā)生中起必不可少的作用。

乙烯敏感性逆轉1 (reversion-to-ethylene sensitivity 1, RTE1)是乙烯反應的一種負調控因子[59], 與受體一起共定位在ER, 但在高爾基體(Golgi apparatus)的膜中也被檢測到[60]。RTE1通過促進ETR1從失活(在乙烯存在下)信號狀態(tài)轉變成為活化(無乙烯)信號狀態(tài)專一地激活ETR1[61]。

盡管受體的確切輸出功能仍然不清楚, 但遺傳學研究表明, 在乙烯缺乏時, 受體激活了途徑中的一個負調控因子CTR1 (constitutive triple response 1)。CTR1是一種絲氨酸/蘇氨酸(Ser/Thr)蛋白激酶, 當它被活化時形成同源二聚體?；罨腃TR1激酶二聚體參與乙烯受體簇之間的交叉相互作用[62]。CTR1的下游是EIN2, 乙烯信號級聯(lián)中的一個關鍵分子。EIN2蛋白包含一個由12個預測的跨膜結構域組成的N端疏水區(qū)域和一個含有保守的核定位序列的親水C端區(qū)域[63-64]。疏水區(qū)域與金屬離子轉運體的NRAMP家族類似, 但EIN2沒有表現(xiàn)出轉運體的活性[65]。EIN2存在于ER膜中, 與乙烯受體的激酶結構域相互作用[66]。當用乙烯處理時, EIN2積累, 以及對于下游途徑組分EIN3的穩(wěn)定是絕對需要的[67]。EIN2作為關鍵組分在乙烯信號中起作用, 但是花了13年多的時間才確定這個有趣的分子怎樣從ER中的受體把乙烯信號傳遞到核內轉錄因子EIN3/EIL1, 從而調控下游基因的表達。已經(jīng)表明, EIN2的C端從ER膜物理運動到細胞核, 允許乙烯信號到達下游組分EIN3和EILs[64,68-69]。Chen等[70]的研究表明, 在乙烯存在下, EIN2在多個絲氨酸和蘇氨酸殘基上缺乏磷酸化。Ju等[68]隨后證明, EIN2與CTR1之間存在物理相互作用, 在乙烯缺乏時CTR1直接磷酸化EIN2 C端的蛋白激酶, 從而阻止C端向下游組分EIN3及其同系物EILs傳遞信號。然而, 去磷酸化是否直接促進EIN2的裂解或者增強EIN2 C端的穩(wěn)定性目前還不清楚[71]。Ju等[68]的研究結果顯示, 對于CTR1和EIN2之間的信號轉導不需要MAPKK或者MAPK活性。EIN2的C端一旦進入細胞核將使EIN3穩(wěn)定和引起EIN3/EILs依賴的轉錄級聯(lián)的活化[64,68-69]。

EIN3和EILs (擬南芥中的EIL1)是短壽命蛋白, 它們作為乙烯信號途徑的正調控因子起作用。EIN3和EIL1是產(chǎn)生乙烯反應的主要輸出的2個關鍵轉錄因子, 對于乙烯反應基因的表達是必需的和足夠的。EIN3/EILs以二聚體的形式起作用, 至少在番茄EIL1中一個保守的磷酸化位點的突變擾亂了煙草()雙分子熒光互補(Bimolecular fluorescence complementation, BiFC)系統(tǒng)中的熒光信號, 以及消除了番茄植株中相應的轉基因活性[72]。當被EIN3/EILs轉錄激活時, 乙烯靶基因介導了植物對乙烯的一系列反應[50]。使用染色質免疫沉淀測序(chromatin immunoprecipitation sequence, ChIP-seq), Chang等[73]發(fā)現(xiàn)EIN3以四波的方式(four-wave manner)調控下游基因的轉錄, 每波包含一組唯一的EIN3靶子, 它們逐漸增加地調節(jié)許多下游的轉錄級聯(lián)。重要的是, 一些下游的EIN3靶子相當于其他激素信號途徑的關鍵組分, 從而強化了不同植物激素之間存在復雜的相互作用網(wǎng)絡的思想。在擬南芥中鑒定的上述所有乙烯信號組分在進化上距離較遠的物種中都是保守的, 表明植物中的乙烯信號機制是普遍的[51]。

1.2.2 信號組分的轉換與反饋調節(jié) 隨著研究進展, 已經(jīng)發(fā)現(xiàn)乙烯線性信號途徑實際上是一條更為復雜的路線, 包括反饋調控的轉錄網(wǎng)絡, 以及mRNA和蛋白質轉換調控模塊[50]。蛋白酶體介導的蛋白質降解在乙烯信號級聯(lián)的調控中起主要作用。在受體水平, 乙烯通過26S蛋白酶體誘導ETR2降解; 同時, 乙烯轉錄激活、和[74]。

EIN2和EIN3/EIL1的蛋白水平也被專一的F-box蛋白嚴格調控, 在乙烯缺乏時, F-box蛋白使它們發(fā)生蛋白酶體介導的降解[75-76]。在對乙烯信號反應中, ETP1 (EIN1-targeting protein 1)和ETP2控制EIN2的水平[76], 而EBF1 (EIN3 binding F-box 1)特別是EBF2調控EIN3的水平[75,77]。為了進一步增加這個調控模塊的復雜性, EBF1/2和ETP1/2的蛋白水平被乙烯下調, 至少在EBF1/2中這一過程被蛋白酶體介導[67,76]。(被乙烯轉錄誘導)本身就是EIN3的一個靶子, 這能夠解釋乙烯反應中每個EBF的不同作用[78], 從而建立一個復雜的反饋調控機制。作為這些調控回路的最終輸出, 細胞核中EIN3/EIL1的蛋白水平被精細地調控, 以協(xié)調一組乙烯反應的活化。換句話說,轉錄的乙烯依賴性的增加與EBF1和EBF2蛋白穩(wěn)定性的減少之間的平衡被認為是調節(jié)EIN3/EIL1的轉換, 為調節(jié)植物對乙烯的反應提供了一種動態(tài)機制。5'–3'核糖核酸外切酶XRN4/EIN5提供了另一個層次的調控, 該酶通過一個未知的機制下調和mRNA的水平。由于EIN5的分子性質, 提出了一種控制乙烯反應的RNA降解模塊[50]。與上述其他調控環(huán)不同, EIN2和ETPs都不受乙烯的轉錄調控[79]。

2 乙烯在種子萌發(fā)與休眠釋放中的作用

2.1 乙烯對種子萌發(fā)的促進作用

乙烯的產(chǎn)生在種子吸脹開始后立即發(fā)生, 并隨著萌發(fā)時間的延長而增加; 乙烯釋放高峰與胚根突破種皮一致[2,80-81]。種子中乙烯的產(chǎn)生是物種依賴的, 但在吸脹過程中乙烯的釋放量常常低于用氣相色譜可檢測到的水平。利用高靈敏度的激光光聲光譜(laser photoacoustic spectroscopy), El-Maarouf- bouteau等[2]已經(jīng)證實在向日葵()種子萌發(fā)結束時乙烯出現(xiàn)高峰。乙烯對種子萌發(fā)的促進作用是劑量依賴的, 當應用的濃度為0.1~200 μL L-1時是有效的, 這取決于物種、休眠深度和環(huán)境條件。盡管乙烯促進許多光敏種子的萌發(fā), 但是它不能克服反枝莧()、芹菜()、萵苣()和大爪草()種子萌發(fā)對光的需要[16]。

2.2 外源乙烯打破種子休眠

外源乙烯或者乙烯利(一種釋放乙烯的化合物)能打破一些種子的初生和次生休眠[16,82](表1)。在表現(xiàn)出種皮強制休眠的一些物種中, 乙烯也能打破休眠和促進萌發(fā), 例如蒼耳()、地三葉()、皺葉酸模()和擬南芥[16]。特別是在萵苣、向日葵、尾穗莧()和繁穗莧()種子中, 乙烯也能打破由高溫誘導的次生休眠[16,83]。ACC能促進松果菊屬()植物種子的萌發(fā)[84], 也能促進寄生植物例如獨腳金()[16]種子的萌發(fā)。乙烯增加非休眠種子在非最適環(huán)境條件例如高溫、滲透脅迫、缺氧和鹽脅迫下的萌發(fā)[16,85-86]。

表1 乙烯、乙烯利或者1-氨基環(huán)丙烷-1-羧酸打破種子休眠的物種(引自Corbuneau et al.[16])

在蘋果種子冷處理[87], 或者向日葵[88]、反枝莧[82]和柱花草()[89]種子干藏過程中, 休眠的打破都與乙烯敏感性的增加有關。在剛收獲時, 休眠的向日葵種子在15℃下需要50 μL L–1乙烯才能發(fā)芽; 但在5℃下分別干藏8周和15周后, 僅僅需要10 μL L-1和3 μL L-1乙烯[88]。相反, 在誘導次生休眠的環(huán)境條件下, 乙烯的反應性在種子萌發(fā)過程中逐漸下降[88]。

2.3 乙烯生物合成與信號對種子萌發(fā)與休眠的調控

許多研究表明, 萌發(fā)能力與乙烯的產(chǎn)生有關, 表明乙烯調控種子萌發(fā)與休眠[13,39,90]。例如, 鷹嘴豆()、向日葵和萵苣種子在高溫下熱休眠的誘導與乙烯產(chǎn)生的降低有關[16]。乙烯產(chǎn)生的下降可能導致ACC-丙二酰轉移酶活性的增加, ACC含量下降; ACO活性被抑制, 或者和表達降低[16,91]。相反, 一些處理(如低溫、GA、一氧化氮、HCN)打破種子休眠, 導致乙烯產(chǎn)生增加[13]。

利用乙烯生物合成途徑的抑制劑或者改變乙烯生物合成和信號途徑的突變體獲得的數(shù)據(jù)表明, 內源乙烯在種子萌發(fā)和休眠的調控中起關鍵作用。種子在氨基乙氧基乙烯甘氨酸(aminoethoxyvinyl glycine)和氨基氧乙酸(aminooxyacetic acid, AOA)(ACS活性抑制劑), CoCl2和α-氨基異丁酸(α-aminoisobutyric acid)(ACO活性抑制劑)或者2,5-降冰片二烯(2,5- norbornadiene)和硫代硫酸銀(silver thiosulfate)(乙烯作用抑制劑)中的萌發(fā)表明, 內源乙烯參與萌發(fā)和打破休眠[16,92]。相反, 乙烯的直接前體ACC促進許多物種的種子萌發(fā), 例如萵苣、向日葵、蒼耳、莧屬植物(sp.)、鷹嘴豆和甜菜()[16]。值得注意的是, ACC氧化的一種副產(chǎn)物HCN也能打破蘋果[93]、向日葵[94]和反枝莧[82]種子的休眠。

利用在乙烯生物合成和信號中發(fā)生改變的擬南芥品系允許表征乙烯對休眠的調控作用。與野生型比較,和突變體的種子表現(xiàn)出初生休眠增加, 可能是由于ABA敏感性提高; 而突變體輕微地提高萌發(fā)速率[95-96]。EIN2的功能喪失導致擬南芥種子在萌發(fā)和早期幼苗發(fā)育過程中對鹽和滲透脅迫過敏[97]。在休眠的水青岡()胚中()的表達極少, 但在濕冷過程中增加, 從而打破休眠[98]。在向日葵中,在非休眠胚中的表達是休眠胚的5倍, 以及的表達被HCN顯著地促進[94]。在番茄萌發(fā)種子中轉錄本的豐度比非萌發(fā)種子高, 在轉基因系中的過表達導致種子成熟前萌發(fā)[99]。

休眠和后熟的擬南芥種子的轉錄組數(shù)據(jù)表明, 在休眠狀態(tài)下基因的表達上調, 在萌發(fā)狀態(tài)下的表達上調[100]。在萵苣中, Argyris等[91]表明乙烯反應基因被熱抑制調控; 在高溫下,和的基因表達下降, 而、和的表達增加。這些結果提示激素代謝與信號調控在基因表達水平上存在差距。在小麥()種子中, 注釋為乙烯代謝和信號基因的78個探針組在休眠和后熟種子之間被差異表達。果膠裂解酶1、擴展蛋白A2、β-1,3-葡聚糖酶和幾丁質酶β被認為是假定的乙烯反應的下游基因, 這些基因在獨行菜()種子萌發(fā)過程中在胚乳弱化和/或者胚根生長中起重要的作用[101]。

3 乙烯與植物激素脫落酸和赤霉素的相互作用

種子植物中種子休眠的激素調控可能是一種高度保守的機制。在許多物種中觀察到種子休眠被ABA誘導和維持, 被GA釋放[102]。利用ABA和GA生物合成和信號突變體的大量遺傳研究表明, 這2種激素在種子休眠和萌發(fā)中具有重要的作用和互相拮抗的作用[10,103]。在萵苣種子萌發(fā)中, ABA抑制種子萌發(fā), GA促進種子萌發(fā)且具有拮抗ABA的作用[104]。下面主要討論乙烯通過抑制ABA的代謝與信號和增強GA的代謝與信號調控種子的萌發(fā)與休眠。

3.1 乙烯和ABA之間的交叉作用

3.1.1 ABA對乙烯代謝的影響 在種子萌發(fā)過程中, ABA和乙烯之間的拮抗作用已經(jīng)在許多物種中被闡明[13-14,105]。在擬南芥和家獨行菜()中, 乙烯拮抗ABA對胚乳帽弱化和胚乳破裂的抑制作用[101]。ABA也增加打破初生和次生休眠的乙烯需要量; ABA對萌發(fā)的抑制與乙烯產(chǎn)生的減少有關; ABA明顯抑制體內ACO的活性, 這種抑制作用與減少的轉錄本的積累有關[13,16,101](圖3)。在擬南芥中, ABA通過ABI4介導的和的轉錄抑制拮抗乙烯的產(chǎn)生[105]。在擬南芥種子萌發(fā)過程中, 胚和胚乳中轉錄本的積累被ABA抑制; ABA不敏感突變體中高水平的轉錄本表明,的表達被ABA調節(jié)[101,106]; 在胚中,轉錄本的積累也被ABA抑制[106]。在家獨行菜中, ABA對和的抑制作用被限于胚乳帽[101]。同樣, 擬南芥突變體的芯片分析發(fā)現(xiàn)了轉錄本積累的上調[107]。

圖3 乙烯、脫落酸和赤霉素在種子萌發(fā)和休眠調控中的相互作用

該方案是基于正文中引證的種子對乙烯、脫落酸或者GA響應的遺傳分析、芯片數(shù)據(jù)和生理研究。乙烯通過抑制ABA的合成和促進它的失活或者分解代謝下調ABA的積累, 也負調控ABA信號。ABA通過ACS和ACO的活性抑制乙烯的生物合成。乙烯也增強GA的代謝和信號, 反過來也一樣。“→”和“┤”分別表示信號級聯(lián)的不同元素之間的正、負相互作用。根據(jù)Corbineau等[16]重繪。

This scheme is based on genetic analyses, microarray data, and physiological studies on seed responsiveness to ethylene, ABA or GA cited in the text. Ethylene down-regulates ABA accumulation by both inhibiting its synthesis and promoting its inactivation or catabolism, and also negatively regulates ABA signaling. ABA inhibits ethylene biosynthesis through ACS and ACO activities. Ethylene also improves the GA metabolism and signaling, and. “→” and“ ┤” indicate positive and negative interactions between the different elements of the signaling cascade, respectively. Redrew from Corbineau et al.[16].

3.1.2 乙烯對ABA代謝和信號的影響和的種子表現(xiàn)出比野生型更高的ABA含量, 以及萌發(fā)更緩慢[95,97]。ABA-葡萄糖酯(ABA-glucose ester)的水平在種子中減少, 因此, 增加的ABA積累可能是由于ABA結合的減少[95]。然而, 乙烯也可能調節(jié)其他酶促步驟, 芯片分析表明在中上調, 在中下調[107]。中的高水平ABA也與的上調有關[97]。

一些研究表明, 在種子萌發(fā)過程中乙烯不僅對ABA代謝起作用以降低ABA水平, 而且負調節(jié)ABA信號(圖3)。實際上, 降低乙烯敏感性的突變(如、和)導致ABA敏感性的增加, 而在和(ethylene overproduction 1)中增加的乙烯敏感性降低了ABA的敏感性[96,101,108]。例如,的突變增強了種子的ABA不敏感性, 乙烯不敏感突變體如降低了ABA不敏感性[109]。然而, 在和中, 沒有觀察到ABA敏感性的顯著性差異[96]。

此外, 水青岡種子酪氨酸磷酸酶在擬南芥種子中的過表達通過ABA信號下調和上調減少休眠, 提出在ABA信號中的負作用可能是由乙烯信號調控的結果[110]。盡管ABA和乙烯信號途徑之間存在相互作用, 但遺傳證據(jù)表明它們可能平行地起作用, 因為通過乙烯突變體(、、和)與突變體雜交獲得的雙突變體表現(xiàn)出ABA缺乏和乙烯敏感性改變所引起的表型[107]。

3.2 乙烯和GA之間的相互作用

在許多物種中, GA促進休眠種子的萌發(fā), 這種休眠也被乙烯、乙烯利或者ACC打破(表1)。在擬南芥中, 乙烯恢復GA缺乏突變體的萌發(fā), 而對番茄GA缺乏突變體的萌發(fā)沒有促進效應; 但GA3促進突變體的萌發(fā)。這些數(shù)據(jù)表明乙烯和GA途徑相互作用[15,39]。

在水青岡種子中, 胚在GA3中培養(yǎng)導致ACC的積累、ACC氧化酶活性和乙烯產(chǎn)生增加, 與的表達增加一致[111]。同樣, GA4對擬南芥突變體種子萌發(fā)的促進作用與的增加有關[112]。在GA生物合成抑制劑多效唑存在下,的表達減少證實GA激活乙烯生物合成途徑[111,113]。然而, 在鉆果大蒜芥()中, Iglesias- Fernandez等[28]表明,和在萌發(fā)過程中的表達被多效唑抑制, 但不受乙烯利或者GA4+7的影響。此外, 在GA4存在時擬南芥中乙烯反應感受器1 (,)(編碼一個乙烯受體家族成員)[112], 以及在GA3存在時水青岡中類[114]的上調表明了GA對乙烯反應的影響。

許多數(shù)據(jù)表明, 乙烯通過影響GA的生物合成或者信號途徑來促進種子萌發(fā)。與野生型比較, GA1、GA4和GA7在擬南芥突變體的干燥成熟種子中大量地積累, 在吸脹的前2 d, GA4和GA7的含量依然比野生型高[95]。萌發(fā)過程中GA含量的變化表明, ETR1即乙烯信號途徑的缺陷導致(1)GA生物合成途徑的改變; (2)促進萌發(fā)需要比野生型更高水平的GA[95]。在水青岡種子中, 與活性GA合成有關的的表達在層積種子(即非休眠種子)和用GA3或者乙烯利處理的種子中仍然較低, 但由AOA所引起的乙烯生物合成的抑制導致這個轉錄本的增加, 表明乙烯參與了GA生物合成的調控[113]。在GA4+7、乙烯以及GA合成或者乙烯合成與信號抑制劑存在時, 鉆果大蒜芥種子吸脹過程中與GA合成(和)和降解()有關的基因表達研究表明, GA生物合成被GA和乙烯顯著地調控[28]。

赤霉素信號途徑依賴于DELLA蛋白, 包括GA不敏感(GAI)、抑制因子(RGA)、類RGA1 (RGL1)、RGL2和RGL3[115]。GA使DELLA蛋白不穩(wěn)定, DELLA蛋白通過與GA結合發(fā)生泛素化和降解, 作為生長抑制因子起作用[116]。Achard等[117]報道, 在擬南芥中乙烯對下胚軸生長和花轉變(floral transition)的一部分作用是通過它對DELLA蛋白的影響介導的。在控制種子萌發(fā)中也可能是這樣, 因為DELLA蛋白似乎在種子萌發(fā)的調控中起關鍵作用[118-119]。因此, 種子中GA的含量和響應性可能是由乙烯對DELLA蛋白積累的調節(jié)所引起的。

4 結語

乙烯促進種子萌發(fā)和休眠釋放, 通過影響ABA和GA的生物合成與信號起重要作用。在ACC的代謝中, 除了被ACO氧化成為乙烯外, 也能被ACC- N-丙二酰轉移酶轉化成為它的主要衍生物1-丙二酰-ACC (1-malonyl-ACC); ACC的第二個衍生物是由γ-谷氨酰轉肽酶催化形成的γ-谷酰基-ACC (γ-glu-tamyl-ACC); 第三個衍生物是茉莉酯-ACC (jasmonyl- ACC); ACC也能由ACC脫氨酶代謝成為銨和α-酮戊二酸[31]。這些衍生物都是精細的生化分流器, 能夠調節(jié)可用于產(chǎn)生乙烯的ACC池, 但這些ACC衍生物的確切生物學作用以及它們之間怎樣維持平衡的機制還不清楚。

Zhang等[120]表明在缺乏的突變體中乙烯受體的N端部分可以有條件地介導受體信號的輸出, 提出了一條不涉及CTR1的交替乙烯受體信號途徑(alternate route of ethylene receptor signaling); 但交替信號轉導途徑所涉及的組分, 以及與線性信號轉導途徑(圖2)在負調控乙烯信號中的動態(tài)協(xié)調是不清楚的。在對寬范圍乙烯濃度的反應中, 這2條途徑可能促進乙烯信號的動態(tài)微調。

乙烯與ABA和GA相互作用, 后2種激素都是種子萌發(fā)與休眠的重要調節(jié)因子[15,102,121]。因此, 乙烯促進種子萌發(fā)的作用可能是通過參與C2H4-GA- ABA的交互作用而產(chǎn)生, 但其作用是直接的還是間接的需要證明; ABA和GA對種子中乙烯生物合成和信號途徑的影響也需要進一步的研究?；钚匝?reactive oxygen species)也通過激素網(wǎng)絡特別是與ABA和GA一起調控種子萌發(fā)[2,13,16], 因此, 區(qū)分不同信號途徑的等級及其作為環(huán)境信號感受器的作用將是重要的。

組學(-omics)技術已應用于種子萌發(fā)與休眠釋放的研究[122-124], 結合種子乙烯生物合成和信號突變體, 以及利用相應的抑制劑實驗, 建立新的乙烯對種子萌發(fā)與休眠釋放的組學研究體系, 包括轉錄組、翻譯組、蛋白質組、代謝組和環(huán)境組等將有助于探明種子萌發(fā)與休眠釋放過程的調控網(wǎng)絡。

[1] 徐恒恒, 黎妮, 劉樹君, 王偉青, 王偉平, 張紅, 程紅焱, 宋松泉. 種子萌發(fā)及其調控的研究進展. 作物學報, 2014,40: 1141–1156. Xu H H, Li N, Liu S J, Wang W Q, Wang W P, Zhang H, Cheng H Y, Song S Q. Research progress in seed germination and its control., 2014, 40: 1141–1156 (in Chinese with English abstract).

[2] El-Maarouf-Bouteau H, Sajjad Y, Bazin J, Langlade N, Cristescu S M, Balzergue S, Baudouin E, Bailly C. Reactive oxygen species, abscisic acid and ethylene interact to regulate sunflower seed germination., 2015, 38: 364–374.

[3] Bewley J D, Bradford K J, Hilhorst H W M, Nonogaki H. Physiology of Development, Germination and Dormancy, 3rd edn. New York: Springer, 2013. pp 247–297.

[4] Finkelstein R, Reeves W, Ariizumi T, Sreber C. Molecular aspects of seed dormancy., 2008, 59: 387–415.

[5] 宋松泉. 種子休眠. 見: “10000個科學難題”農(nóng)業(yè)科學編委會. 10000個科學難題. 北京: 科學出版社, 2011. pp 31–35. Song S Q. Seed dormancy. In: The Editorial Board of Agricultural Science for 10000 Selected Problems in Sciences, eds. 10000 Selected Problems in Sciences. Beijing: Science Press, 2011. pp 31–35 (in Chinese).

[6] Shu K, Liu X D, Xie Q, He Z H. Two faces of one seed: hormonal regulation of dormancy and germination., 2016, 9: 34–45.

[7] Lenser T, Theissen G. Molecular mechanisms involved in convergent crop domestication., 2013, 18: 704–714.

[8] Meyer R S, Purugganan M D. Evolution of crop species: genetics of domestication and diversification., 2013, 14: 840–852.

[9] Simsek S, Ohm J B, Lu H, Rugg M, Berzonsky W, Alamri M S, Mergoum M. Effect of pre-harvest sprouting on physicochemical changes of proteins in wheat., 2014, 94: 205–212.

[10] Graeber K, Nakabayashi K, Miatton E, Leubner-Metzger G, Soppe W J J. Molecular mechanisms of seed dormancy., 2012, 35: 1769–1786.

[11] Hoang H H, Sechet J, Bailly C, Leymarie J, Corbineau F. Inhibition of germination of dormant barley (L.) grains by blue light as related to oxygen and hormonal regulation., 2014, 37: 1393–1403.

[12] Lee H G, Lee K, Seo P J. The Arabidopsis MYB96 transcription factors play a role in seed dormancy., 2015, 87: 371–381.

[13] Arc E, Sechet J, Corbineau F, Rajjou L, Marion-Poll A. ABA crosstalk with ethylene and nitric oxide in seed dormancy and germination., 2013, 4: 63. doi: 10.3389/fpls.2013. 00063.

[14] Linkies A, Leubner-Metzger G. Beyond gibberellins and abscisic acid: how ethylene and jasmonates control seed germination., 2012, 31: 253–270.

[15] Miransari M, Smith D L. Plant hormones and seed germination., 2014, 99: 110–121.

[16] Corbineau F, Xia Q, Bailly C, EI-Maarouf-Bouteau H. Ethylene, a key factor in the regulation of seed dormancy., 2014, 5: 539. doi:10.3389/fpls.2014.00539.

[17] Khan N A, Khan M I R, Ferrante A, Poor P. Editorial: Ethylene: a key regulatory molecule in plants., 2017, 8: 1782. doi:10.3389/fpls.2017.01782.

[18] Khan N A, Khan M I R. The Ethylene: from senescence hormone to key player in plant metabolism., 2014, 2: e124. doi:10.4172/2329-9029.1000e124.

[19] Lin Z, Zhong S, Grierson D. Recent advances in ethylene research., 2009, 60: 3311–3336.

[20] Schaller G E. Ethylene and the regulation of plant development., 2012, 10: 9. doi:10.1186/1741-7007-10-9.

[21] Müller M, Munné-Bosch S. Ethylene response factors: a key regulatory hub in hormone and stress signaling., 2015, 169: 32–41.

[22] Thao N P, Khan M I R, Thu N B A, Hoang X L T, Asgher M, Khan N A, Tran L S P. Role of ethylene and its cross talk with other signaling molecules in plant responses to heavy metal stress., 2015, 169: 73–84.

[23] Arraes F B M, Beneventi M A, de Sa M E L, Paixao J F R, Albuquerque E V S, Marin S R R, Purgatto E, Nepomuceno A L, Grossi-de-Sa M F. Implications of ethylene biosynthesis and signaling in soybean drought stress tolerance., 2015, 15: 213. doi:10.1186/s12870-015-0597-z.

[24] Habben J E, Bao X, Bate N J, DeBruin J L, Dolan D, Hasegawa D, Helentjaris T G, Lafitte R H, Lovan N, Mo H, Reimann K, Schussler J R. Transgenic alteration of ethylene biosynthesis increases grain yield in maize under field drought-stress conditions., 2014, 12: 685–693.

[25] Lieberman M, Mapson L W. Genesis and biogenesis of ethylene., 1964, 204: 343–345.

[26] Lieberman M, Kunishi A, Mapson L W, Wardale D A. Stimulation of ethylene production in apple tissue slices by methionine., 1966, 41: 76–82.

[27] Yang S F, Hoffman N E. Ethylene biosynthesis and its regulation in higher plants., 1984, 35: 155–189.

[28] Iglesias-Fernandez R, Matilla A. Genes involved in ethylene and gibberellins metabolism are required for endosperm-limited germination ofL. seeds., 2010, 231: 653–664.

[29] Yamagami T, Tsuchisaka A, Yamada K, Haddon W F, Harden L A, Theologis A. Biochemical diversity among the 1-aminocyclopropane-1-carboxylate synthase isozymes encoded by the Arabidopsis gene family., 2003, 278: 49102–49112.

[30] Tsuchisaka A, Yu G, Jin H, Alonso J M, Ecker J R, Zhang X, Gao S, Theologis A. A combinatorial interplay among the 1-aminocy-clo-propane-1-carboxylate isoforms regulates ethylene biosynthesis in., 2009, 183: 979–1003.

[31] Van de Poel B, Van Der Straeten D. 1-aminocyclopropane-1- carboxylic acid (ACC) in plants: more than just the precursor of ethylene!, 2014, 5: 640. doi: 10.3389/fpls.2014. 00640.

[32] Yoon G M, Kieber J J. 1-Aminocyclopropane-1-carboxylic acid as a signalling molecule in plants., 2013, 5: plt017. doi:10.1093/aobpla/plt017.

[33] Christians M J, Gingerich D J, Hansen M, Binder B M, Kieber J J, Vierstra R D. The BTB ubiquitin ligases ETO1, EOL1, and EOL2 act collectively to regulate ethylene biosynthesis in Arabidopsis by controlling type-2 ACC synthase levels., 2009, 57: 332–345.

[34] Lyzenga W J, Booth J K, Stone S L. The Arabidopsis RING- type E3 ligase XBAT32 mediates the proteasomal degradation of the ethylene biosynthetic enzyme, 1-aminocyclopropane- 1-carboxylate synthase 7., 2012, 71: 23–34.

[35] Kamiyoshihara Y, Iwata M, Fukaya T, Tatsuki M, Mori H. Turnover of LeACS2, a wound-inducible 1-aminocyclopropane- 1-carboxylic acid synthase in tomato, is regulated by phosphorylation/dephosphorylation., 2010, 64: 140–150.

[36] Ludwikow A, Ciesla A, Kasprowicz-Maluski A, Mitula F, Tajdel M, Galganski L, Ziolkowski P A, Kubiak P, Maleck A, Piechalak A, Szabat M, Gorska A, Dabrowski M, Ibragimow I, Sadoqski J. Arabidopsis protein phosphatase 2C ABI1 interacts with type I ACC synthases and is involved in the regulation of ozone-induced ethylene biosynthesis., 2014, 7: 960–967.

[37] Skottke K R, Yoon G M, Kieber J J, DeLong A. Protein phosphatase 2A controls ethylene biosynthesis by differentially regulating the turnover of ACC synthase isoforms., 2011, 7: e1001370. doi: 10.1371/journal.pgen.10 01370.

[38] Van de Poel B, Bulens I, Markoula A, Hertog M L A T M, Dreesen R, Wirtz M, Vandoninck S, Oppermann Y, Keulemans J, Hell R, Waelkens E, De Proft M P, Sauter M, Nicolai B M, Geeraerd A H. Targeted systems biology profiling of tomato fruit reveals coordination of the Yang cycle and a distinct regulation of ethylene biosynthesis during postclimacteric ripening., 2012, 160: 1498–1514.

[39] Matilla A J, Matilla-Vazquez M A. Involvement of ethylene in seed physiology., 2008, 175: 87–97.

[40] Ververidis P, John P. Complete recovery in vitro of ethylene- forming enzyme-activity., 1991, 30: 725–727.

[41] Murphy L J, Robertson K N, Harroun S G, Brosseau C L, Werner-Zwanziger U, Moilanen J, Tuononen H M, Clyburne J A C. A simple complex on the verge of breakdown: isolation of the elusive cyanoformate ion., 2014, 344: 75–78.

[42] 葉永健, 宋松泉. Fe2+和CO２對番木瓜ACC氧化酶活性的影響. 中山大學學報, 1997, 36(2): 18–21. Ye Y J, Song S Q. Effect of Fe2+and CO2on 1-aminocyc-lo-propane-1-carboxylate oxidase from papaya (L.) fruit., 1997, 36 (2): 18–21.

[43] Dong J G, Fernandezmaculet J C, Yang S F. Purification and characterization of 1-aminocyclopropane-1-carboxylate oxidase from apple fruit., 1992, 89: 9789–9793.

[44] Song S Q, Ye Y J. Effect of O2, ACC and CO2concentration on the activity of partially purified ACC oxidase from papaya., 2001, 27: 387–392.

[45] Hudgins J W, Ralph S G, Franceschi V R, Bohlmann J. Ethylene in induced conifer defense: cDNA cloning, protein expression, and cellular and subcellular localization of 1-aminocyclopropane- 1-carboxylate oxidase in resin duct and phenolic parenchyma cells., 2006, 224: 865–877.

[46] Ramassamy S, Olmos E, Bouzayen M, Pech J C, Latche A. 1-aminocyclopropane-1-carboxylate oxidase of apple fruitis periplasmic., 1998, 49: 1909–1915.

[47] De Paepe A, Vuylsteke M, van Hummelen P, Zabeau M, van Der Staeten D. Transcriptional profiling by cDNA-AFLP and microarray analysis reveals novel insights into the early response to ethylene in Arabidopsis., 2004, 39: 537–559.

[48] Van de Poel B, Bulens I, Hertog M L A T M, Nicolai B, Geeraerd A. A transcriptomics-based kinetic model for ethylene biosynthesis in tomato () fruit: development, validation and exploration of novel regulatory mechanisms., 2014, 202: 952–963.

[49] Binder B M, Chang C, Schaller G E. Perception of ethylene by plants-ethylene receptors., 2012, 44: 117–145.

[50] Stepanova A N, Alonso J M. Ethylene signaling and response: where different regulatory modules meet., 2009, 12: 548–555.

[51] Merchante C, Alonso J, Stepanova A N. Ethylene signaling: simple ligand, complex regulation., 2013, 16: 554?560.

[52] Bakshi A, Piya S, Fernandez J C, Chervin C, Hewezi T, Bindera B M. Ethylene receptors signal via a noncanonical pathway to regulate abscisic acid responses., 2018, 176: 910–929.

[53] Ju C, Chang C. Advances in ethylene signalling: protein complexes at the endoplasmic reticulum membrane., 2012, 2012: pls031. doi: 10.1093/aobpla/pls031.

[54] Shakeel S N, Wang X, Binder B M, Schaller G E. Mechanisms of signal transduction by ethylene: overlapping and non-overlapping signalling roles in a receptor family., 2013, 5: plt010. doi:10.1093/aobpla/plt010.

[55] Chen Y F, Gao Z, Kerris R J, Wang W, Binder B M, Schaller G E. Ethylene receptors function as components of high-molecular- mass protein complexes in Arabidopsis., 2010, 5: e8640. doi: 10.1371/journal.pone.0008640.

[56] Gao Z, Wen C K, Binder B M, Chen Y F, Chang J, Chiang Y H, Kerris R J, Chang C, Schaller G E. Heteromeric interactions among ethylene receptors mediate signaling in Arabidopsis., 2008, 283: 23801–23810.

[57] Hirayama T, Kieber J J, Hirayama N, Kogan M, Guzman P, Nourizadeh S, Alonso J M, Dailey W P, Dancis A, Ecker J R. RESPONSIVE-TO-ANTAGONIST1, a Menkes/Wilson disease related copper transporter, is required for ethylene signaling in Arabidopsis., 1999, 97: 383–393.

[58] Binder B M, Rodriguez F I, Bleecker A B. The copper transporter RAN1 is essential for biogenesis of ethylene receptors in Arabidopsis., 2010, 285: 37263–37270.

[59] Resnick J S, Wen C K, Shockey J A, Chang C. REVERSION-TO ETHYLENE SENSITIVITY 1, a conserved gene that regulates ethylene receptor function in Arabidopsis., 2006, 103: 7917–7922.

[60] Dong C H, Rivarola M, Resnick J S, Maggin B D, Chang C. Subcellular colocalization of Arabidopsis RTE1 and ETR1 supports a regulatory role for RTE1 in ETR1 ethylene signaling., 2008, 53: 275–286.

[61] Resnick J S, Rivarola M, Chang C. Involvement of RTE1 in conformational changes promoting ETR1 ethylene receptor signaling in Arabidopsis., 2008, 56: 423–431.

[62] Mayerhofer H, Panneerselvam S, Mueller-Dieckmann J. Protein kinase domain of CTR1 frompromotes ethylene receptor cross talk., 2012, 415: 768–779.

[63] Alonso J M, Hirayama T, Roman G, Nourizadeh S, Ecker J R. EIN2, a bifunctional transducer of ethylene and stress responses in Arabidopsis., 1999, 284: 2148–2152.

[64] Wen X, Zhang C, Ji Y, Zhao Q, He W, An F, Jiang L, Guo H. Activation of ethylene signaling is mediated by nuclear translocation of the cleaved EIN2 carboxyl terminus., 2012, 22: 1613–1616.

[65] Cho Y H, Lee S, Yoo S D. EIN2 and EIN3 in ethylene signaling., 2012, 44: 169–187.

[66] Bisson M M A, Groth G. New insight in ethylene signaling: autokinase activity of ETR1 modulates the interaction of receptors and EIN2., 2010, 3: 882–889.

[67] An F, Zhao Q, Ji Y, Li W, Jiang Z, Yu X, Zhang C, Han Y, He W, Liu Y, Zhang S, Ecker J R, Guo H. Ethylene-induced stabilization of ETHYLENE INSENSITIVE 3 and EIN3-LIKE1 is mediated by proteasomal degradation of EIN3 binding F-box 1 and 2 that requires EIN2 in Arabidopsis., 2010, 22: 2384–2401.

[68] Ju C, Yoon G M, Shemansky J M, Lin D Y, Ying Z I, Chang J, Garrett W M, Kessenbrock M, Groth G, Tucker M L, Cooper B, Kieber J J, Chang C. CTR1 phosphorylates the central regulator EIN2 to control ethylene hormone signaling from the ER membrane to the nucleus in Arabidopsis., 2012, 109: 19486–19491.

[69] Qiao H, Shen Z, Huang S C, Schmitz R J, Urich M A, Briggs S P, Ecker J R. Processing and subcellular trafficking of ER-tethered EIN2 control response to ethylene gas., 2012, 338: 390–393.

[70] Chen R, Binder B M, Garrett W M, Tucker M L, Chang C, Cooper B. Proteomic responses inseedlings treated with ethylene., 2011, 7: 2637–2650.

[71] Ji Y, Guo H. From endoplasmic reticulum (ER) to nucleus: EIN2 bridges the gap in ethylene signaling., 2013, 6: 11–14.

[72] Li J, Li Z, Tang L, Yang Y, Zouine M, Bouzayen M. A conserved phosphorylation site regulates the transcriptional function of ETHYLENE-INSENSITIVE3-like1 in tomato., 2012, 63: 427–439.

[73] Chang K N, Zhong S, Weirauch M T, Hon G, Pelizzola M, Li H, Huang S S C, Schmitz R J, Urich M A, Kuo D, Nery J R, Qiao H, Yang A, Jamali A, Chen H, Ideker T, Ren B, Bar-Joseph Z, Hughes T R, Ecker J R. Temporal transcriptional response to ethylene gas drives growth hormone cross-regulation in Arabidopsis., 2013, 2: e00675. doi: 10.7554/eLife.00675.

[74] Chen Y F, Shakeel S N, Bowers J, Zhao X C, Etheridge N, Schaller G E. Ligand-induced degradation of the ethylene receptor ETR2 through a proteasome-dependent pathway in Arabidopsis., 2007, 282: 24752–24758.

[75] Gagne J M, Smalle J, Gingerich D J, Walker J M, Yoo S D, Yanagisawa S, Vierstra R D. Arabidopsis EIN3-binding F-box 1 and 2 form ubiquitin-protein ligases that repress ethylene action and promote growth by directing EIN3 degradation., 2004, 101: 6803–6808.

[76] Qiao H, Chang K N, Yazaki J, Ecker J R. Interplay between ethylene, ETP1/ETP2 F-box proteins, and degradation of EIN2 triggers ethylene responses in Arabidopsis., 2009, 23: 512–521.

[77] Potuschak T, Lechner E, Parmentier Y, Yanagisawa S, Grava S, Koncz C, Genschik P. EIN3-dependent regulation of plant ethylene hormone signaling by two Arabidopsis F box proteins: EBF1 and EBF2., 2003, 115: 679–689.

[78] Konishi M, Yanagisawa S. Ethylene signaling in Arabidopsis involves feedback regulation via the elaborate control of EBF2 expression by EIN3., 2008, 55: 821–831.

[79] Kevany B M, Tieman D M, Taylor M G, Cin V D, Klee H J. Ethylene receptor degradation controls the timing of ripening in tomato fruit., 2007, 51: 458–467.

[80] Fu J R, Yang S F. Release of heat pretreatment-induced dormancy in lettuce seeds by ethylene or cytokinin in relation to the production of ethylene and the synthesis of 1-aminocyclopropane-1- carboxylic acid during germination., 1983, 2: 185–192.

[81] Siriwitayawan G, Geneve R L, Downie A B. Seed germination of ethylene perception mutants of tomato and Arabidopsis., 2003, 13: 303–314.

[82] K?pczyński J, Sznigir P. Participation of GA3, ethylene, NO and HCN in germination ofL. seeds with various dormancy levels., 2014, 36: 1463–1472.

[83] Van de Poel, Smet D, Van Der Straeten D. Ethylene and hormonal cross talk in vegetative growth and development., 2015, 169: 61–72.

[84] Wood L A, Kester S T, Geneve R L. The physiological basis for ethylene-induced dormancy release in threespecies with special reference to the influence of the integumentary tapetum., 2013: 156: 63–72.

[85] Lin Y, Yang L, Paul M, Zu Y, Tang Z. Ethylene promotes germination of Arabidopsis seed under salinity by decreasing reactive oxygen species: evidence for the involvement of nitric oxide simulated by sodium niroprusside., 2013, 73: 211–218.

[86] Silva P O, Medina E F, Barros R S, Ribeiro D M. Germination of salt-stressed seeds as related to ethylene biosynthesis ability in threespecies., 2014, 171: 14–22.

[87] Sinska I. Interaction of ethephon with cytokinin and gibberellin during the removal of apple seed dormancy and germination of embryos., 1989, 64: 39–44.

[88] Corbineau F, C?me D. Germination of sunflower seeds as related to ethylene synthesis and sensitivity: an overview. In: Vendrell M, Klee H, Pech J C, Romojaro F, eds. Biology and Biotechnology of the Plant Hormone Ethylene III. Amsterdam: IOS Press, 2003. pp 216–221.

[89] Ribeiro D M, Barros R S. Sensitivity to ethylene as a major component in the germination of seeds of., 2006, 16: 37–45.

[90] Gniazdowska A, Krasuska U, Bogatek R. Dormancy removal in apple embryos by nitric oxide or cyanide involves modifications in ethylene biosynthetic pathway., 2010, 232: 1397–1407.

[91] Argyris J, Dahal P, Hayashi E, Still D W, Bradford K J. Genetic variation for lettuce seed thermoinhibition is associated with temperature-sensitive expression of abscisic acid, gibberellin, and ethylene biosynthesis, metabolism, and response genes., 2008, 148: 926–947.

[92] Hermann K, Meinhard J, Dobrev P, Linkies A, Pesek B, Hess B, Machá?ková I M, Fischer U, Leubner-Metzger G. 1-Amynocyclopropane-1-carboxylic acid and abscisic acid during the germination of sugar beet (L.): a comparative study of fruits and seeds., 2007, 58: 3047–3060.

[93] Krasuska U, Ciacka K, Debska K, Bogatek R, Gniazdowska A. Dormancy alleviation by NO or HCN leading to decline of protein carbonylation levels in apple (Borkh.) embryos., 2014, 171: 1132–1141.

[94] Oracz K, El-Maarouf-Bouteau H, Bogatek R, Corbineau F, Bailly C. Release of sunflower seed dormancy by cyanide: cross-talk with ethylene signaling pathway., 2008, 59: 2241–2251.

[95] Chiwocha S D S, Cutler A J, Abrams S R, Ambrose S J, Yang J, Kermode A R. The ert1-2 mutation inaffects the abscisic acid, auxin, cytokinin and gibberellin metabolic pathways during main tenance of seed dormancy, moist-chilling and germination., 2005, 42: 35–48.

[96] Subbiah V, Reddy K J. Interactions between ethylene, abscisisc acid and cytokinin during germination and seedling establishment in Arabidopsis., 2010, 35: 451–458.

[97] Wang Y, Liu C, Li K, Sun F, Hu H, Li X, Zhao Y, Han C, Zhang W, Duan Y, Liu M, Li X. Arabidopsis EIN2 modulates stress response through abscisic acid response pathway., 2007, 64: 633–644.

[98] Jimenez J A, Rodriguez D, Calvo A P, Mortensen L C, Nicolas G, Nicolas C. Expression of a transcription factor (FsERF1) involved in ethylene signaling during the breaking of dormancy inseeds., 2005, 125: 373–380.

[99] Pirrello J, Jaimes-Miranda F, Sanchez-Ballesta M T, Tournier B, Khalil-Ahmad Q, Regad F, Latché A, Pech J C, Bouzayen M. Sl-ERF2, a tomato ethylene response factor involved in ethylene response and seed germination., 2006, 47: 1195–1205.

[100] Cadman C S C, Toorop P E, Hilhorst H W M, Finch-Savage W E. Gene expression profiles of Arabidopsis Cvi seeds during dormancy cycling indicate a common underlying dormancy control mechanism., 2006, 46: 805–822.

[101] Linkies A, Müller K, Morris K, Ture?ková V, Wenk M, Cadman C S C, Corbineau F, Strnad M, Lynn J R, Finch-Savage W E, Leubner-Metzger G. Ethylene interacts with abscisic acid to regulate endosperm rupture during germination: a comparative approach usingand., 2009, 21: 3803–3822.

[102] Nonogaki H. Seed dormancy and germination-emerging mechanism and new hypotheses., 2014, e5: 233. doi:10.3389/fpls.2014.00233.

[103] Liu X, Hou X L. Antagonistic regulation of ABA and GA in metabolism and signaling pathways., 2018, 9: 251. doi:10.3389/fpls.2018.00251.

[104] Dong T T, Tong J H, Xiao L T, Cheng H Y, Song S Q. Nitrate, abscisic acid and gibberellin interactions on the thermoinhibition of lettuce seed germination., 2012, 66: 191– 202.

[105] Dong Z, Yu Y, Li S, Wang J, Tang S, Huang R. Abscisic acid antagonizes ethylene production through the ABI4-mediated transcriptional repression ofandin Arabidopsis., 2016, 9: 126–135.

[106] Penfield S, Li Y, Gilday A D, Graham S, Graham I A. Arabidopsis ABA INSENSITIVE 4 regulates lipid mobilization in the embryo and reveals repression of seed germination by the endosperm., 2006, 18: 1887–1899.

[107] Cheng W H, Chiang M H, Hwang S G, Lin P C. Antagonism between abscisic acid and ethylene in Arabidopsis acts inparallel with the reciprocal regulation of their metabolism and signaling pathways., 2009, 71: 61–80.

[108] Kucera B, Cohn M A, Leubner-Metzger G. Plant hormone interactions during seed dormancy release and germination., 2005, 15: 281–307.

[109] Beaudoin N, Serizet C, Gosti F, Giraudat J. Interactions between abscisic acid and ethylene signaling cascades., 2000, 12: 1103–1115.

[110] Alonso-Ramirez A, Rodriguez D, Reyes D, Jimenez J A, Nicolas G, Nicolas C. Functional analysis inof FsPTP1, a tyrosine phosphatase from beechnuts, reveals its role as a negative regulator of ABA signaling and seed dormancy and suggests its involvement in ethylene signaling modulation., 2011, 234: 589–597.

[111] Calvo A P, Nicolas C, Lorenzo O, Nicolas G, Rodriguez D. Evidence for positive regulation by gibberellins and ethylene of ACC oxidase expression and activity during transition from dormancy to germination inL. seeds., 2004, 23: 44–53.

[112] Ogawa M, Hanada A, Yamauchi Y, Kuwahara A, Kamiya Y, Yamaguchi S. Gibberellin biosynthesis and response during Arabidopsis seed germination., 2003, 15: 1591–1604.

[113] Calvo A P, Nicolas C, Nicolas G, Rodriguez D. Evidence of a crosstalk regulation of a GA 20-oxidase (FsGA20ox1) by gibberellins and ethylene during the breaking of dormancy inseeds., 2004, 120: 623–630.

[114] Lorenzo O, Rodriguez D, Nicolas C, Nicolas G. Characterization and expression of two protein kinase and an EIN3-like genes, which are regulated by ABA and GA3in dormantseeds. In: Black M, Bradford K J, Vazquez-Ramos J, eds. Seed Biology: Advances and Applications. Wallingford: CAB International, 2000. pp 329–340.

[115] Davière J M, Achard P. Gibberellin signaling in plants., 2013, 140: 1147–1151.

[116] Dill A, Thomas S G, Steber C M, Sun T P. The Arabidopsis F-box protein SLEEPY1 targets gibberellin signaling repressors for gibberellin-induced degradation., 2004, 16: 1392–1405.

[117] Achard P, Baghour M, Chapple A, Hedden P, Van Der Straeten D, Genschik P, Moritz T, Harberd N P. The plant stress hormone ethylene controls floral transition via DELLA-dependent regulation of floral meristem-identity genes., 2007, 104: 6484–6489.

[118] Piskurewicz U, Jikumaru Y, Kinoshita N, Nambara E, Kamiya Y, Lopez-Molina L. The gibberellic acid signaling repressor RGL2 inhibits Arabidopsis seed germination by stimulating abscisic acid synthesis and ABI5 activity., 2008, 20: 2729–2745.

[119] Schwechheimer C. Understanding gibberellic acid signaling: are we there yet?, 2008, 11: 9–15.

[120]Zhang J, Yu J, Wen C-K. An alternate route of ethylene receptor signaling., 2014, 5: 648. doi:10.3389/ fpls.2014.00648.

[121] Nambara E, Okamoto M, Tatematsu K, Yano R, Seo M, Kamiya Y. Abscisic acid and the control of seed dormancy and germination., 2010, 20: 55–67.

[122]Liu S J, Song S H, Wang W Q, Song S Q. De novo assembly and characterization of germinating lettuce seed transcriptome using Illumina paired-end sequencing., 2015, 96: 154?162.

[123] Wang W Q, Song B Y, Deng Z J, Wang Y, Liu S J, M?ller I M, Song S Q. Proteomic analysis ofseed germination and thermoinhibition by sampling of individual seeds at germination and removal of storage proteins by PEG fractionation., 2015, 167: 1332–1350.

[124] Xu H H, Liu S J, Song S H, Wang W Q, M?ller I M, Song S Q. Proteome changes associated with dormancy release of Dongxiang wild rice seeds., 2016, 206: 68–86.

Biosynthesis and signaling of ethylene and their regulation on seed germination and dormancy

SONG Song-Quan1,3,*, LIU Jun2, XU Heng-Heng2, ZHANG Qi2, HUANG Hui3, and WU Xian-Jin3

1Institute of Botany, Chinese Academy of Sciences, Beijing 100093, China;2Agro-Biological Gene Research Center, Guangdong Academy of Agricultural Sciences, Guangzhou 510640, Guangdong, China;3Key Laboratory of Research and Utilization of Ethnomedicinal Plant Resources of Hunan Province / College of Biological and Food Engineering, Huaihua University, Huaihua 418008, Hunan, China

Seed germination, a key ecological and agronomic trait, is determined by both internal and external cues that regulate the dormancy status and the potential for germination in seeds, and plays a critical role during the subsequent growth, development and production of plants. Dormancy is the temporary failure of seed germination under favorable conditions. Ethylene is a simple gaseous phytohormone with multiple roles in regulation of metabolism at molecular, cellular, and whole plant levels. It influences performance of plants under optimal and stressful environments by interacting with other signaling molecules. In the present paper, we mainly summarize ethylene biosynthesis and signaling, the role of ethylene in seed germination and dormancy release, and the interaction of ethylene with phytohormone abscisic acid and gibberellin, and propose some scientific problems to be required to investigate further in order to provide an idea for explaining the molecular mechanism of seed germination and dormancy regulated by ethylene.

abscisic acid; biosynthesis and signaling; crosstalk; ethylene; gibberellin; seed germination and dormancy

2018-11-22;

2019-01-19;

2019-04-09.

10.3724/SP.J.1006.2019.84175

宋松泉, E-mail: sqsong@ibcas.ac.cn

本研究由國家科技支撐計劃項目(2012BAC01B05), 國家自然科學基金項目(31371715, 31640059)和廣東省科技計劃項目(2016B030303007)資助。

This study was supported by the National Science and Technology Support Program (2012BAC01B05), the National Natural Science Foundation of China (31371715, 31640059), and the Guangdong Science and Technology Program (2016B0303007).

URL: http://kns.cnki.net/kcms/detail/11.1809.S.20190404.1439.002.html