摘要 鞘脂類物質(zhì)是構(gòu)成植物膜系統(tǒng)的重要組分,在不同的植物細胞和組織中鞘脂的結(jié)構(gòu)和含量分布迥異。鞘脂也是細胞內(nèi)重要的信號分子,參與調(diào)控植物的程序性細胞死亡、氣孔開閉、根的生長、花粉發(fā)育、植物形態(tài)建成和果實成熟與脫落等多個過程。鞘脂代謝紊亂會造成底物或者產(chǎn)物的積累,進而導(dǎo)致植物生長、發(fā)育異常。目前植物中鞘脂的研究主要通過突變體表型和鞘脂類物質(zhì)的含量來評估,缺乏對精密調(diào)控網(wǎng)絡(luò)的研究,因此有較大的研究空白亟待填補。綜述了目前參與植物鞘脂代謝過程的鞘脂類物質(zhì)及其相關(guān)的代謝酶類,并分類討論了鞘脂代謝基因突變體表型和造成突變體表型的潛在原因。
關(guān)鍵詞 鞘脂;鞘脂穩(wěn)態(tài);鞘氨醇;神經(jīng)酰胺;生長發(fā)育
中圖分類號:Q946 DOI:10.16152/j.cnki.xdxbzr.2024-05-005
Sphingolipids metabolism regulates plant growth and development
LUO Xiao1,CHEN Liyu2
(1.College of Life Sciences, Fujian Agriculture and Forestry University, Fuzhou 350002,China;
2.National Engineering Research Center for Sugarcane/School of Future Technology,F(xiàn)ujian Agriculture and Forestry University, Fuzhou 350002, China)
Abstract Sphingolipids are essential components of plant biomembrane system with their structure and content varying significantly across different plant cells and tissues.They also function as intracellular signaling molecules, participating in the regulation of processes such as programmed cell death, stomatal open and closure, root growth, pollen development, plant morphogenesis, and fruit ripening and shedding.Disruptions in sphingolipid metabolism result in the accumulation of substrates or products, leading to abnormal plant development.Currently, research on sphingolipids in plants primarily evaluates mutant phenotype and changes of sphingolipids content. However, there are significant research gaps due to a lack of precise regulatory networks. This review summarized sphingolipids involved in plant sphingolipid metabolism and their related enzyme. It also discussed the mutant phenotype of sphingolipid metabolism related genes and the possible causes of the mutant phenotype.
Keywords sphingolipid; sphingolipid homeostasis; sphingosine; ceramide; growth and development
鞘脂(sphingolipids)是一類結(jié)構(gòu)復(fù)雜并且功能多樣的脂質(zhì)的總稱。普遍存在于真核細胞和部分細菌的膜組分中,維持膜結(jié)構(gòu)的完整性[1-2]。在植物細胞中鞘脂約占質(zhì)膜脂質(zhì)的40%[3]。鞘脂結(jié)構(gòu)的多樣性導(dǎo)致了功能的多樣性。鞘脂參與多種生物學(xué)過程,如作為信號分子參與細胞間和細胞內(nèi)信號傳遞[4]、程序性細胞死亡(programmed cell death, PCD)[1, 5-7]、脫落酸(abscisic acid, ABA)依賴的保衛(wèi)細胞的氣孔關(guān)閉[8]等過程。此外,鞘脂也影響植物的生長發(fā)育過程,比如種子的萌發(fā)[9]、根的發(fā)育[6]、花粉的發(fā)育[10-11]、細胞類型的分化與器官的發(fā)生[12]和果實發(fā)育[13]等。鞘脂穩(wěn)態(tài)對于植物的生長發(fā)育至關(guān)重要,鞘脂的合成、轉(zhuǎn)運以及分解過程發(fā)生改變均會引起植物產(chǎn)生相應(yīng)的表型,進而影響植物的生存和繁衍。
鞘脂在動物中的研究相對廣泛,鞘脂代謝途徑被擾亂會導(dǎo)致多種疾病的產(chǎn)生[14]。由于技術(shù)手段的限制,在植物中鞘脂的研究相對滯后。近年來,隨著生物技術(shù)的發(fā)展,尤其是基因編輯技術(shù)[15]和質(zhì)譜技術(shù)[3, 16]的使用,極大地促進了植物鞘脂研究的進度,多種重要植物鞘脂代謝突變體的表型及其鞘脂含量與分布得以證明。但鞘脂在植物生長發(fā)育中的功能研究目前尚處于起步階段。結(jié)構(gòu)多樣化的鞘脂對維持植物的生長發(fā)育至關(guān)重要,那么植物中的鞘脂到底如何形成,在哪里形成,去向哪里,以及發(fā)揮怎樣的功能呢?本文就目前關(guān)于鞘脂調(diào)控植物生長發(fā)育相關(guān)的研究進展進行了總結(jié)和討論。
1 植物鞘脂的合成、分解及轉(zhuǎn)運
1884年,約翰·路德維希·圖迪庫姆(Johann Ludvig Thudichum)在動物的腦組織中發(fā)現(xiàn)了結(jié)構(gòu)獨特的脂類,因其結(jié)構(gòu)像“謎一樣”(sphinx),故將其命名為“sphingo”。1947年,赫伯特·卡特(Herbert Carter)和同事在研究和闡述鞘脂的結(jié)構(gòu)時,提出了現(xiàn)在常見的鞘脂描述詞“sphingolipid”[17]。鞘脂的結(jié)構(gòu)在不同物種之間存在一定的共性,除了鞘氨醇以外,其基礎(chǔ)骨架均由鞘氨醇和脂肪酸構(gòu)成,最主要的區(qū)別在于鏈長、修飾基團和飽和度的不同[18]。植物鞘脂主要分為四大類(見圖1),即游離的長鏈堿基(long-chain bases, LCB)、神經(jīng)酰胺(ceramides, Cer)、葡糖神經(jīng)酰胺(glucosyl-ceramides, GlcCer)和糖基肌醇磷酸神經(jīng)酰胺(glycosyl-inositol-phosphoryl-ceramides, GIPCs)。
1.1 植物鞘脂的結(jié)構(gòu)
植物中的長鏈堿基(也被稱為鞘氨醇)是鞘脂的關(guān)鍵組分,長鏈堿基通常為18碳的氨基醇,一般分為兩類。一類以二氫鞘氨醇(d18∶0, dihydrosphinganine)為基礎(chǔ)骨架,其基本的特征是在C1和C3位置處各有一個羥基〔見圖1(a)〕。另一類以4-羥基鞘氨醇〔t18∶0, hydroxyphinganine,也被稱為植物鞘氨醇(Phyto-LCB)〕為骨架,在二氫鞘氨醇的基礎(chǔ)上進一步在C4位羥基化形成〔見圖1(b)〕。兩者共有特征是在C2位置有一個氨基[19-20]。鞘氨醇結(jié)構(gòu)豐富的另一個原因是C4和C8位可以形成不飽和的雙鍵,并且C8位存在順反異構(gòu)體的形式(4E, 8E, 8Z)[1],由此形成了9種不同結(jié)構(gòu)的鞘氨醇〔見圖1(a)和(b)〕。位于C8位置的雙鍵構(gòu)型可以是順式或反式,而位于C4位置的雙鍵通常為反式構(gòu)型,并且順式和反式異構(gòu)體的比例因植物種類而異[4]。鞘氨醇被認(rèn)為是最簡單的鞘脂。
神經(jīng)酰胺是形成高級鞘脂的骨架,由鞘氨醇C2位的氨基和脂肪酸(fatty acid, FA)的脂?;纬甚0锋I連接而成〔見圖1(c)〕[1]。在植物中,神經(jīng)酰胺的脂肪酸鏈含有14-26個碳原子[1],通常碳原子數(shù)目為偶數(shù),但也存在少量含有21、23和25個碳原子的情況[21]。神經(jīng)酰胺脂肪酸鏈的C2位發(fā)生羥基化可以形成羥基神經(jīng)酰胺(hydroxyceramides, hCer)[4]。通過脂肪酸鏈長度的變化,鞘氨醇和脂肪酸的羥基化以及碳鏈的去飽和等衍生出結(jié)構(gòu)多樣化的神經(jīng)酰胺。9種不同的鞘氨醇和32種不同的脂肪酸的組合可以產(chǎn)生288種潛在的神經(jīng)酰胺[22]。神經(jīng)酰胺骨架構(gòu)成復(fù)雜鞘脂的疏水尾部,通過在神經(jīng)酰胺分子中鞘氨醇鏈的C1位添加不同的極性基團形成具有更高級結(jié)構(gòu)的鞘脂,同時也賦予鞘脂分子雙親性[23]。極性基團可以是1分子的磷酸殘基〔見圖1(d)〕,也可以是多種糖基殘基[1, 4]。當(dāng)鞘氨醇的C1位與1分子的葡萄糖殘基通過糖苷鍵連接時,形成最簡單的鞘糖脂,即葡糖神經(jīng)酰胺〔見圖1(e)〕[24]。神經(jīng)酰胺骨架與肌醇磷酸基團連接可形成肌醇磷酸神經(jīng)酰胺(inositol-phosphoryl-ceramides, IPC),該結(jié)構(gòu)可以進一步糖基化形成糖基肌醇磷酸神經(jīng)酰胺〔見圖1(e)〕[25]。鞘脂多樣性的另外一個來源是糖基肌醇磷酸神經(jīng)酰胺頭部基團的多樣性,其糖基化的糖基類型和數(shù)量多變,常見糖基的主要類型有葡萄糖、阿拉伯糖、半乳糖和甘露糖等[26-28]。糖基肌醇磷酸神經(jīng)酰胺根據(jù)極性頭部基團中所包含的糖基的數(shù)量分為7個不同的系列,依次為0系列到F系列(Series 0 ~ Series F)[29]。0系列為基礎(chǔ)的糖基肌醇磷酸神經(jīng)酰胺,A系列在0系列的基礎(chǔ)上添加了1個己糖殘基,B系列在A系列的基礎(chǔ)上添加了1個己糖殘基,依次類推,可到達F系列〔見圖1(e)〕。部分研究表明最多可以有10個糖基殘基被添加到糖基肌醇磷酸神經(jīng)酰胺的結(jié)構(gòu)中[30]。極性頭部基團的多樣性是由植物種類和所在的器官或組織類型決定的[1, 19, 28]。
1.2 植物鞘脂的合成
在真核生物中,鞘脂從頭合成和降解的主要途徑基本保守,但鞘脂結(jié)構(gòu)的多樣性在不同物種之間存在差異,有些鞘脂的結(jié)構(gòu)修飾僅發(fā)生在植物中[2, 17, 22]。植物中鞘脂合成可以分為鞘氨醇的合成、神經(jīng)酰胺的合成和復(fù)雜鞘脂的合成3個步驟,其中神經(jīng)酰胺是鞘脂代謝的中心。在植物中,鞘脂的合成起始于內(nèi)質(zhì)網(wǎng)(endoplasmic reticulum, ER),合成的神經(jīng)酰胺被轉(zhuǎn)運至高爾基體(Golgi)進一步修飾形成復(fù)雜的鞘脂。植物鞘脂合成和降解的基本途徑如圖2所示。
鞘氨醇由絲氨酸棕櫚?;D(zhuǎn)移酶(serine palmitoyl transferase, SPT)將絲氨酸和棕櫚酰輔酶A縮合形成中間體3-酮基二氫鞘氨醇(3-ketosphinganine)[31-32],該中間體通過3-酮基鞘氨醇還原酶(3-ketosphinganine reductase, KSR)還原為二氫鞘氨醇(d18∶0),擬南芥中的KSR分別由KSR1和KSR2基因編碼,二者在功能上存在冗余,KSR1占主導(dǎo)地位,具有更高的還原酶活性[33]。鞘氨醇羥化酶(sphingoid base hydroxylase, SBH)在二氫鞘氨醇的C4位添加第三個羥基,生成植物鞘氨醇(t18∶0)[32]。t18∶0和t18∶1是植物中最豐富的鞘氨醇[34]。含有不飽和鍵的鞘氨醇的形成依賴于鞘氨醇 Δ4去飽和酶(LCB Δ4 desaturase, LCB Δ4 DES)和鞘氨醇 Δ8去飽和酶(LCB Δ8 desaturase, LCB Δ8 DES)。植物中鞘氨醇Δ4的不飽和與鞘氨醇Δ8的不飽和幾乎同時發(fā)生,與LCB Δ8 DES相比,LCB Δ4 DES只在反式構(gòu)型中引入雙鍵[35]。LCB Δ8 DES僅在植物和一些絲狀或雙態(tài)性真菌中能檢測到,具有一定的物種特異性,其在擬南芥中有兩個同源基因,即SLD1(Sphingoid LCB Desaturase 1)和SLD2[36-37]。鞘氨醇羥化酶和去飽和酶的存在是鞘氨醇具有結(jié)構(gòu)多樣性的主要原因。
神經(jīng)酰胺通過神經(jīng)酰胺合成酶(ceramide synthase, CerS)將鞘氨醇和脂肪酸縮合形成,目前在植物中鑒定出3個CerS〔Longevity Assurance Gene One (LAG1) Homologue 1-3, LOH 1-3〕,其中LOH1和LOH3具有合成鞘氨醇和極長鏈脂肪酸(very-long-chain fattyacid, VLCFAs, C20~C26)的偏好性,而LOH2主要負(fù)責(zé)含有長鏈脂肪酸(long-chain fatty acid, LCFAs, C16~C18)的神經(jīng)酰胺的合成[38-40]。神經(jīng)酰胺中的脂肪酸在脂肪酸2-羥基化酶(fatty acid 2-hydroxylase, FAH)的作用下形成羥基神經(jīng)酰胺(hydroxyceramide, hCer),擬南芥有2個FAH同源基因,分別是AtFAH1和AtFAH2,AtFAH1主要羥基化VLCFAs,而AtFAH2偏好含有C16的脂肪酸[41-42]。值得注意的是,在酵母中的研究表明,鞘脂類脂肪酸的2-羥基化可能僅發(fā)生在神經(jīng)酰胺的脂?;溕希皇怯坞x的脂?;溕?sup>[43-44]。
神經(jīng)酰胺作為底物,與葡萄糖在葡糖神經(jīng)酰胺合成酶(GlcCer synthase, GCS)的催化下形成葡糖神經(jīng)酰胺[45]。GCS在擬南芥中定位于內(nèi)質(zhì)網(wǎng)[46]。合成葡糖神經(jīng)酰胺的神經(jīng)酰胺骨架主要由16個碳原子的脂肪酸和二氫鞘氨醇構(gòu)成[38]。合成糖基肌醇磷酸神經(jīng)酰胺的神經(jīng)酰胺骨架主要由極長鏈脂肪酸和植物鞘氨醇組成[23]。糖基肌醇磷酸神經(jīng)酰胺主要在高爾基體上通過一系列酶促反應(yīng)形成。首先,合成肌醇磷酸神經(jīng)酰胺,由肌醇磷酸神經(jīng)酰胺合成酶(IPC synthase, IPCS)將磷脂酰肌醇的頭部基團轉(zhuǎn)移到神經(jīng)酰胺上形成肌醇磷酸神經(jīng)酰胺,在擬南芥中IPCS 由3個基因編碼,分別是IPCS1(也被稱為enhancing RPW8-mediated HR-like cell death, ERH1)、IPCS2和IPCS3[47-48]。合成后的肌醇磷酸神經(jīng)酰胺進一步添加一些糖基殘基形成糖基肌醇磷酸神經(jīng)酰胺。一般情況下,最先通過肌醇磷酸神經(jīng)酰胺葡萄糖醛酸糖基轉(zhuǎn)移酶1(inositol-phosphoryl-ceramide glucuronosyl transferase 1, IPUT1)轉(zhuǎn)移一分子葡萄糖醛酸(glucuronic acid, GlcA)到肌醇磷酸神經(jīng)酰胺上,形成糖基肌醇磷酸神經(jīng)酰胺[49]。糖基肌醇磷酸神經(jīng)酰胺在擬南芥中有兩條潛在的途徑形成更復(fù)雜的糖基肌醇磷酸神經(jīng)酰胺[18]。第一種途徑是通過糖基肌醇磷酸神經(jīng)酰胺甘露糖轉(zhuǎn)移酶1(GIPC mannosyl transferase 1, GMT1),將甘露糖殘基轉(zhuǎn)移至糖基肌醇磷酸神經(jīng)酰胺,從而形成甘露糖糖基肌醇磷酸神經(jīng)酰胺(mannose-GIPCs)[50]。GDP-甘露糖主要由高爾基體核糖體糖基轉(zhuǎn)運體1或2(golgi nucleotide sugar transporter 1/2, GONST1/2)將GDP-甘露糖轉(zhuǎn)移到高爾基體腔,提供給GMT1[27, 51]。另一種途徑是通過葡萄糖胺肌醇磷酸神經(jīng)酰胺轉(zhuǎn)移酶1(glucosamine inositol-phosphoryl-ceramide transferase 1, GINT1)在糖基肌醇磷酸神經(jīng)酰胺中添加一個N-乙酰葡萄糖胺殘基,形成GlcN(Ac)-GIPCs[52]。與GONST 1/2相似,轉(zhuǎn)運N-乙酰葡萄糖胺殘基的轉(zhuǎn)運體1(UDP-GlcNAc transporter 1, UGNT1)也是定位在高爾基體上的糖基轉(zhuǎn)運體[53]。
1.3 植物鞘脂的分解
植物鞘脂的降解主要包括神經(jīng)酰胺、葡糖神經(jīng)酰胺和糖基肌醇磷酸神經(jīng)酰胺的降解。復(fù)雜的鞘脂最終被分解代謝為神經(jīng)酰胺、鞘氨醇或者鞘氨醇-1-磷酸[54]。鞘氨醇-1-磷酸可以進一步在二羥鞘氨醇-1-磷酸水解酶(dihydrosphingosine-1-phosphate lyase 1, DPL1)的作用下分解形成磷酸乙醇胺和含有16個碳原子的脂肪醛[55-56]。參與神經(jīng)酰胺分解的酶是神經(jīng)酰胺酶類(ceramidases, CDases),此類酶具有細胞器特異性,可能對不同形式的神經(jīng)酰胺具有專一性,從而使得復(fù)雜鞘脂生成特定的鞘氨醇[54]。一般情況下,CDases可以將神經(jīng)酰胺分解為游離的鞘氨醇和脂肪酸[57],但研究發(fā)現(xiàn)在植物(擬南芥)[7]和動物(人)[58]中還存在具有反向活性的CDase,即參與鞘氨醇與脂肪酸形成神經(jīng)酰胺的酶活性。在動物鞘脂的研究中,使用體外檢測的方法,依據(jù)酶解時最適pH值的不同,將CDase分為中性、堿性和酸性神經(jīng)酰胺酶[54],但目前在植物中僅發(fā)現(xiàn)中性神經(jīng)酰胺酶和堿性神經(jīng)酰胺酶。擬南芥有3個中性神經(jīng)酰胺酶(neutral ceramidases,NCERs)與人類的NCER同源[59],分別命名為AtNCER 1(At1g07380)、AtNCER 2 (At2g38010)和AtNCER 3 (At5g58980)。其中,AtNCER 1參與羥基神經(jīng)酰胺的水解, 而AtNCER 2可能具有神經(jīng)酰胺合成酶的功能, 即神經(jīng)酰胺酶反向活性[7]。 水稻中鑒定出一個中性神經(jīng)酰胺酶OsCDase, 定位在內(nèi)質(zhì)網(wǎng)和高爾基體上, 體外檢測其酶活的最適pH值為5.7和6.6, 相對含有d18∶0的神經(jīng)酰胺而言, 優(yōu)先水解含t18∶1(主要在Δ4發(fā)生不飽和)的神經(jīng)酰胺[57, 60],并且發(fā)現(xiàn)OsCDase可能具有反向神經(jīng)酰胺酶活性[60]。此外,在小麥中也克隆出一個中性神經(jīng)酰胺酶基因TaCDase[61]。在魔芋中克隆出來的中性神經(jīng)酰胺酶的最適pH值為6.5~8.0[62]。在擬南芥中僅發(fā)現(xiàn)了兩個堿性神經(jīng)酰胺酶(alkaline ceramidases, ACER),其中之一是AtACER,該基因突變會提高植物對鹽的敏感度和增加植物對細菌性病原體的敏感性[59]。另一個是參與花粉管生長并且與角果保衛(wèi)細胞膨壓相關(guān)的TOD1(TurgOr regulation Defect1)[63]。
葡糖神經(jīng)酰胺的分解主要依賴于葡糖神經(jīng)酰胺酶(glucosylceramidase, GCD),擬南芥有4個和人類GCD同源的基因,分別是At5g49900、At1g33700、At4g10060(AtGCD3)和At3g24180[64]。目前在擬南芥中僅鑒定出AtGCD 3具有酸性葡糖神經(jīng)酰胺酶活性,該酶主要定位在質(zhì)膜(plasma membrane, PM)上,部分定位在內(nèi)質(zhì)網(wǎng),AtGCD3在pH為4.5 ~ 6.0時酶活性較高,最適pH為6.0,優(yōu)先水解含C16和C18的葡糖神經(jīng)酰胺[64]。植物中復(fù)雜鞘脂糖基肌醇磷酸神經(jīng)酰胺的分解與磷脂酶D(phospholipase D, PLD)相關(guān),一種是糖基肌醇磷酸神經(jīng)酰胺特異性的PLD(GIPC-PLD)。催化糖基肌醇磷酸神經(jīng)酰胺中磷酸基團和肌醇之間連接的鍵,產(chǎn)生的底物為植物神經(jīng)酰胺-1-磷酸(phytoceramide-1-phosphate, Phyto-C-1-P),該酶偏好含有兩種糖基基團的糖基肌醇磷酸神經(jīng)酰胺,肌醇磷酸神經(jīng)酰胺、甘露糖肌醇磷酸神經(jīng)酰胺和3個糖基基團的糖基肌醇磷酸神經(jīng)酰胺均不能作為底物[65]。另一種是非特異性的磷脂酶C4(nonspecific phospholipase C4, NPC4)。NPC4被鑒定為在磷饑餓時負(fù)責(zé)催化糖基肌醇磷酸神經(jīng)酰胺水解的酶,該酶對糖基肌醇磷酸神經(jīng)酰胺作為底物的活性高于對普通甘油磷脂酰膽堿的活性,并且NPC4定位在質(zhì)膜上,與脂筏中糖基肌醇磷酸神經(jīng)酰胺的分解相關(guān)[66]。
1.4 植物中鞘脂的轉(zhuǎn)運
合成或分解后的鞘脂均需要一定的介質(zhì)來轉(zhuǎn)運產(chǎn)物到達目的場所,才能發(fā)揮具體的生物學(xué)功能。前人將鞘脂的運輸方式分為非囊泡運輸和囊泡運輸兩種[67]。非囊泡運輸一般為蛋白質(zhì)介導(dǎo)的運輸。ACD11(Accelerated Cell Death 11)被證明具有神經(jīng)酰胺-1-磷酸和植物神經(jīng)酰胺-1-磷酸轉(zhuǎn)移酶活性,晶體結(jié)構(gòu)揭示其通過表面定位的磷酸基團識別中心與神經(jīng)酰胺結(jié)合[68],但是ACD11不能轉(zhuǎn)運糖脂[69]。從擬南芥中鑒定出另外一種轉(zhuǎn)運植物鞘脂的蛋白為糖脂轉(zhuǎn)移蛋白(glycolipid transfer protein 1, GLTP1),由At2g33470編碼的AtGLTP1的活性位點是Arg59和Asn95,其在體外轉(zhuǎn)運葡糖神經(jīng)酰胺的速率較快,轉(zhuǎn)運半乳糖神經(jīng)酰胺和乳糖神經(jīng)酰胺的速率較慢[70],表明AtGLTP1具有葡糖神經(jīng)酰胺特異性。此外,植物細胞中反式高爾基體網(wǎng)絡(luò)結(jié)構(gòu)(trans-Golgi network, TGN)分泌富含C24和C26的羥基化鞘脂的囊泡,介導(dǎo)生長素載體PIN2的極性分選過程,在此過程中合成的鞘脂也隨囊泡進行轉(zhuǎn)移[71]。最近有研究發(fā)現(xiàn),植物脂質(zhì)翻轉(zhuǎn)酶(flippases)的氨基磷脂ATP酶(aminophospholipid ATPases, ALAs)參與鞘脂相關(guān)的跨脂質(zhì)雙分子層的選擇性運輸[72]。其中,ALA4和ALA5可能翻轉(zhuǎn)葡糖神經(jīng)酰胺到細胞質(zhì)面,隨后葡糖神經(jīng)酰胺被定位在質(zhì)膜上的GCDs降解。已有的研究證明了在缺乏ALAs的情況下,葡糖神經(jīng)酰胺會在胞質(zhì)外積累,ALA類翻轉(zhuǎn)酶促進葡糖神經(jīng)酰胺的分解代謝,并且類似的分解代謝途徑也存在于人類和一些酵母中,表明該機制具有保守性[73]。
1.5 植物中鞘脂穩(wěn)態(tài)的調(diào)控
鞘脂的合成和分解途徑涉及多個過程和多種酶的參與,每一步驟的擾亂都可能影響細胞中鞘脂的穩(wěn)態(tài),進而影響相應(yīng)的生物學(xué)過程。SPT是鞘脂從頭合成途徑中第一步反應(yīng)的催化酶,也是關(guān)鍵的限速酶[74],SPT的核心組分是LCB1和LCB2亞基組成的內(nèi)質(zhì)網(wǎng)膜相關(guān)的異二聚體[31, 74],在擬南芥中LCB2由LCB2a(At5g23670)和LCB2b(At3g48780)編碼。另外SPT還具有一個含56個氨基酸的小亞基ssSPT(small suit of SPT),擬南芥的ssSPT由兩個基因編碼,分別是ssSPTa(At1g06515)和ssSPTb(At2g30942)[75],植物中SPT的活性主要由ssSPT和類黏膜蛋白(orosomucoid, ORM)調(diào)控(見圖3A)[22],ssSPT可以與LCB1和LCB2相互作用,增強SPT的活性[75]。擬南芥中ORMs由ORM1(At1g01230)和ORM2(At5g42000)編碼,ORMs通過與ssSPTs相互作用抑制SPT活性[76]。ORM的功能喪失會激發(fā)鞘脂的從頭合成途徑,導(dǎo)致鞘脂的急劇積累,最為顯著的是鞘氨醇和神經(jīng)酰胺的積累[76]。與植物不同的是,在動物和酵母中ORM蛋白與LCB1和LCB2發(fā)生直接相互作用調(diào)節(jié)SPT的活性[77]。
磷酸化修飾也是參與鞘脂穩(wěn)態(tài)調(diào)控的一種方式。植物中鞘氨醇和神經(jīng)酰胺與其磷酸化形式的衍生物之間存在代謝平衡,植物中積累鞘氨醇和神經(jīng)酰胺會導(dǎo)致PCD,但外源施加其磷酸化形式的衍生物可以緩解PCD的表型〔見圖3(b)〕[78-79]。擬南芥中的鞘氨醇可被LCB激酶(LCB kinases, LCB-Ks)磷酸化生成磷酸化的鞘氨醇(LCB phosphates, LCB-Ps),LCBKs通常也被稱為鞘氨酸激酶(sphingosine kinases, SPHKs),由4個基因編碼,分別是LCBK1、LCBK2、SPHK1和SPHK2,植物中LCB-Ps一般分為鞘氨醇-1-磷酸(sphingosine-1-phosphate, S1P)和植物鞘氨醇-1-磷酸(phytosphingosine-1-phosphate, Phyto-S1P)[80-83]。反之,LCB-Ps可以被LCB磷酸磷酸酶(long-chain base phosphate phosphatases, LCB-PP1/2)去磷酸化形成鞘氨醇[80]。與鞘氨醇相似,神經(jīng)酰胺與其磷酸化衍生物之間也存在代謝平衡的調(diào)節(jié),神經(jīng)酰胺可以被Cer激酶(Cer kinases, Cer-K)磷酸化形成磷酸化的神經(jīng)酰胺(ceramide-1-phosphates, Cer-1-P)。與LCB-K不同,Cer-K僅以神經(jīng)酰胺為底物,對鞘氨醇沒有活性,表明Cer-1-P的形成不依賴于LCB-P的形成[9],在擬南芥中有一個編碼Cer-K的基因被命名為ACD5(accelerated cell death 5),調(diào)節(jié)擬南芥中Cer-1-P的代謝平衡[9]。但是迄今為止尚未發(fā)現(xiàn)將Cer-1-P去磷酸化形成神經(jīng)酰胺的酶。
目前關(guān)于影響植物鞘脂穩(wěn)態(tài)的因素除了植物自身表達的蛋白調(diào)控外, 還存在一些化學(xué)試劑干擾調(diào)控鞘脂的合成過程(見圖2)。伏馬菌素B1(Fumonisin B1, FB1)和AAL(Alternaria alternatalycopersici)毒素均可以干擾鞘脂代謝過程,引起游離鞘氨醇的積累[84]。 FB1是一種鞘氨醇類似物, 被認(rèn)為是探究植物鞘脂多種功能的研究工具[6, 85]。 在擬南芥中, FB1對LOH1具有較強的抑制作用[40]。 FB1處理植物會導(dǎo)致鞘氨醇(d18∶0)、植物鞘氨醇(t18∶0)和LCB-Ps的積累[5, 55, 86]。AAL也抑制鞘脂的從頭合成過程,導(dǎo)致細胞內(nèi)鞘氨醇的積累,鞘氨醇是細胞內(nèi)的PCD信號[87]。由AAL引起的PCD表型可以被多球殼菌素(myriocin)減弱[88]。由于myriocin是鞘脂合成途徑中SPT的抑制劑,myriocin處理植物會導(dǎo)致游離鞘氨醇(d18∶0)的積累被減少,合成神經(jīng)酰胺的底物減少,從而使得誘導(dǎo)PCD的信號減弱[88]。此外,PDMP(1-phenyl-2-decanoylamino-3-morpholino-1-propanol, PDMP)通過抑制GCS的活性抑制葡糖神經(jīng)酰胺的合成,使用PDMP處理煙草的葉片和擬南芥的根時觀察到高爾基體復(fù)合體的形態(tài)被破壞,并且干擾高爾基體介導(dǎo)的蛋白質(zhì)轉(zhuǎn)運、液泡的快速融合和液泡膜的內(nèi)陷[46, 89]。
1.6 植物中鞘脂的分布與富集區(qū)域
鞘脂在細胞內(nèi)的分布和富集區(qū)域可能與其功能的特異性相關(guān)。鞘脂是細胞膜的主要結(jié)構(gòu)組分,但在生物膜系統(tǒng)中的分布并不均勻,糖基肌醇磷酸神經(jīng)酰胺和葡糖神經(jīng)酰胺主要富集在細胞膜的外小葉(external leaflet)中[90]。此外,鞘脂和甾醇在細胞質(zhì)膜上富集形成脂筏(lipid rafts)結(jié)構(gòu)[91],脂筏與特定質(zhì)膜蛋白的分選和運輸有關(guān),比如ATP酶、阿拉伯半乳聚糖蛋白和糖基磷脂酰肌醇錨定蛋白等,由鞘脂介導(dǎo)的這些蛋白參與細胞中的多種活動過程,包括細胞壁合成、降解和一些信號傳導(dǎo)過程[69, 92, 93]。
有研究表明細胞中可能至少有500種不同的鞘脂分子[94]。在擬南芥中,至少有168種不同的鞘脂[16, 95]。在擬南芥葉片中,糖基肌醇磷酸神經(jīng)酰胺和葡糖神經(jīng)酰胺分別占總鞘脂含量的64%和34%,糖基肌醇磷酸神經(jīng)酰胺的含量約為葡糖神經(jīng)酰胺的兩倍,表明糖基肌醇磷酸神經(jīng)酰胺在植物葉片中含量最豐富[23]。在植物花粉中最豐富的鞘脂是葡糖神經(jīng)酰胺[96]。游離的鞘氨醇和神經(jīng)酰胺在植物組織中含量較低,不足總鞘脂含量的3%[16]。相對于葉片而言,花粉中游離鞘氨醇和神經(jīng)酰胺的含量更高[16]。對擬南芥葉片鞘脂的檢測發(fā)現(xiàn)約85%的鞘氨醇主要以C4位羥基化的形式存在, 并且大多數(shù)在C8位不飽和(t18∶1, 8E/8Z)[97]。擬南芥中C4位不飽和的鞘氨醇(d18∶2)表達模式具有特異性,僅在花粉和花中可檢測到,在其他組織中幾乎不存在[98]。但是在除了十字花科外的其他物種中,含C4位不飽和的鞘脂(如含d18∶2)在植株的不同組織中均能表達,并且在番茄和大豆等物種中葡糖神經(jīng)酰胺的鞘氨醇鏈大多以d18∶2的形式存在[23, 99]。 擬南芥LCB Δ4 DES缺失突變體花的形態(tài)和花粉活力與野生型之間沒有差異, 但花中葡糖神經(jīng)酰胺的含量減少了約25%, 表明LCB Δ4 DES催化的產(chǎn)物決定下游葡糖神經(jīng)酰胺的合成[98], 即反映出植物中鞘氨醇的結(jié)構(gòu)類型決定了下游產(chǎn)物的分配, 并最終決定了總鞘脂或特定鞘脂類的組成和含量。
2 鞘脂代謝對植物生長發(fā)育的影響
鞘脂類物質(zhì)的結(jié)構(gòu)和種類多樣,在植物細胞中的含量和分布參差不齊,進而導(dǎo)致功能各異。當(dāng)植物受到脅迫時,位于質(zhì)膜上的鞘脂響應(yīng)外界環(huán)境的變化,同時作為細胞內(nèi)信號分子的鞘脂進一步將脅迫信號傳遞給下游響應(yīng)的靶標(biāo)來調(diào)控植物對外界環(huán)境的適應(yīng)。越來越多的研究表明鞘脂可能與植物細胞中的激素之間存在密切的聯(lián)系,并以細胞或組織特異性的方式調(diào)節(jié)植物的生長和發(fā)育。此外,ROS途徑與鞘脂介導(dǎo)的PCD也具有相關(guān)性。功能多樣的鞘脂在植物生長發(fā)育的不同階段分別扮演不同的角色(見圖4)。
2.1 鞘脂誘導(dǎo)植物PCD
植物細胞中鞘脂及其磷酸化衍生物之間的動態(tài)平衡調(diào)節(jié)PCD過程[5, 79]。鞘脂代謝途徑相關(guān)的真菌毒素如FB1和AAL也會誘導(dǎo)植物產(chǎn)生PCD,該過程被認(rèn)為涉及鞘脂代謝的紊亂[100]。FB1誘導(dǎo)的PCD具有光依賴性,需要水楊酸(salicylic acid, SA)、茉莉酸(jasmonic acid, JA)和乙烯(ethylene, ET)信號通路的介導(dǎo)[101]。在擬南芥中,SPT轉(zhuǎn)錄本在葉片衰老過程中積累,SPT催化形成的鞘脂代謝產(chǎn)物參與植物離體組織或病原體感染應(yīng)激的信號轉(zhuǎn)導(dǎo),從而調(diào)節(jié)隨后的PCD過程[102]。FBR11(Fumonisin B1 Resistant 11)編碼SPT的小亞基LCB1,使用FB1處理fbr11-1突變體后鞘氨醇的形成減少,檢測發(fā)現(xiàn)fbr11-1突變體中缺乏活性氧中間產(chǎn)物(reactive oxygen intermediates, ROIs)的產(chǎn)生,使得fbr11-1突變體并未表現(xiàn)出PCD的表型,由此推測游離鞘氨醇通過調(diào)節(jié)ROIs水平誘導(dǎo)植物PCD[5]。進一步的研究發(fā)現(xiàn)磷酸化形式的二氫鞘氨醇-1-磷酸以劑量依賴的方式特異性阻斷二氫鞘氨醇誘導(dǎo)的PCD,這種互相拮抗的調(diào)節(jié)作用可能是通過調(diào)節(jié)ROIs來實現(xiàn)的,也表明游離的鞘氨醇及其磷酸化衍生物通過活性氧途徑?jīng)Q定細胞命運[5]。
擬南芥神經(jīng)酰胺激酶突變體acd5植株在發(fā)育初期生長正常,發(fā)育后期由于依賴于SA途徑的神經(jīng)酰胺積累導(dǎo)致PCD,并且JA通過調(diào)節(jié)鞘脂代謝和增加神經(jīng)酰胺水平促進acd5突變體的細胞死亡[79, 103]。Cer-1-P能夠部分挽救神經(jīng)酰胺積累引起的細胞死亡,表明神經(jīng)酰胺及其磷酸化衍生物之間的平衡調(diào)節(jié)植物的PCD[79]。此外,神經(jīng)酰胺也被證明以鈣依賴的方式誘導(dǎo)植物PCD[104]。ACD11也參與PCD和防御途徑,在acd11突變體中激活PCD和防御信號通路依賴于SA,并且需要光,但不完全依賴于ET和JA信號通路[69]。此外,擬南芥中IPCS也參與PCD和防御調(diào)控,編碼IPCS的ERH1功能缺失會導(dǎo)致SA積累,進而增強擬南芥防御相關(guān)基因RPW的表達和神經(jīng)酰胺的積累并導(dǎo)致PCD,植物的外觀表現(xiàn)為株高降低[47]。fah1fah2突變體與loh2突變體雜交產(chǎn)生的fah1fah2loh2三重突變體也會導(dǎo)致PCD,此過程依賴于SA和EDS1(enhanced disease susceptibility 1),可能是fah1fah2loh2突變體積累了鞘氨醇d18∶0、鞘氨醇t18∶0 和鞘氨醇-1-磷酸d18∶0 P引起的[42]。
2.2 鞘脂調(diào)控植物氣孔開閉
植物蒸騰作用損失的水分和光合作用吸收的二氧化碳幾乎都是通過植物葉片表面的氣孔進行的[105]。氣孔孔徑的動態(tài)調(diào)節(jié)是植物優(yōu)化水分利用和二氧化碳吸收的關(guān)鍵,氣孔的開啟或關(guān)閉伴隨著保衛(wèi)細胞膨壓的調(diào)節(jié)[106]。因此氣孔不僅對植物的生長發(fā)育至關(guān)重要,而且對全球的水循環(huán)和碳循環(huán)也具有重要意義[105, 107]。與鞘脂代謝相關(guān)的LCB-Ps已經(jīng)被驗證是參與調(diào)控氣孔孔徑的信號分子[108-109],LCB-Ps以ABA依賴的方式影響胞質(zhì)中Ca2+濃度的振蕩來調(diào)節(jié)氣孔的開關(guān)[108]。負(fù)責(zé)催化LCB-Ps生成的激酶SPHK也可被ABA激活,通過調(diào)節(jié)保衛(wèi)細胞K+通道和陰離子通道的方式參與ABA抑制的氣孔打開和促進氣孔關(guān)閉的過程[8]。參與ABA介導(dǎo)的氣孔孔徑調(diào)控的LCB-Ps一般為LCB(d18∶1)-P (Δ4磷酸化衍生物)和LCB(t18∶0)-P(植物鞘氨醇-1-磷酸)[108-109]。擬南芥中At3g58490基因編碼的AtSPP1是LCB-P去磷酸化的酶,spp1突變體的氣孔比野生型對ABA更敏感,表明AtSPP1可能通過LCB-P介導(dǎo)的ABA信號在氣孔應(yīng)答中發(fā)揮作用[110]。由磷脂酶D(phospholipase Ds, PLDs)催化產(chǎn)生的磷脂酸(phosphatidic acid, PA)可以與SPHK結(jié)合并提高其產(chǎn)生植物鞘氨醇-1-磷酸的活性[81],植物鞘氨醇-1-磷酸活性的增加激活下游的PLDα1,隨后導(dǎo)致PA水平升高。在ABA介導(dǎo)的氣孔關(guān)閉過程中,PA作為信號分子調(diào)控ABI1和NADPH氧化酶等下游蛋白[111-112]。ABA信號被傳導(dǎo)到下游通路,調(diào)控離子通道,導(dǎo)致氣孔關(guān)閉[113]。此外,LCB-Ps還需要異源三聚體G蛋白的α-亞基GPA1來調(diào)節(jié)保衛(wèi)細胞離子通道活性和氣孔開度[8]。鞘脂調(diào)控植物氣孔孔徑變化的信號通路缺乏明確的受體和轉(zhuǎn)導(dǎo)的信號分子,更多的調(diào)控機理需要進一步展開研究。
2.3 鞘脂調(diào)控植物根的生長
鞘脂代謝途徑影響非生物脅迫條件下植物根的發(fā)育過程。低溫誘導(dǎo)擬南芥快速形成植物鞘氨醇-1-磷酸,該過程由LCBK2介導(dǎo),lcbk2突變體在22 ℃和4 ℃下的生長表型與野生型植物相似,但在12 ℃下表現(xiàn)出更長的根生長表型,主要與冷響應(yīng)性DELLA基因家族的RGL3表達量變化有關(guān)[114]。RGL3在lcbk2中的表達改變可能與植物鞘氨醇-1-磷酸的代謝改變有關(guān)[114]。擬南芥acer-1突變體在鹽脅迫條件下種子萌發(fā)率顯著下降、根長變短,AtACER轉(zhuǎn)錄本的減少導(dǎo)致神經(jīng)酰胺的積累和鞘氨醇的減少,其中植物神經(jīng)酰胺和植物鞘氨醇(t18∶0)的變化最顯著,AtACER可能在鹽脅迫下通過植物神經(jīng)酰胺和植物神經(jīng)酰胺-1-磷酸代謝通路發(fā)揮作用[59]。棉花GhIPCS1基因的異源表達可以增強擬南芥對鹽的敏感性,擬南芥過表達株系種子的萌發(fā)率顯著性降低、根長變短,可能是不同鞘脂類之間的平衡被破壞導(dǎo)致上述表型的產(chǎn)生[115]。在擬南芥中,npc4突變體的主根長度在缺磷條件下比野生型短約25%,但突變體主根的細胞長度與寬度均與野生型相似,可能由于植物體內(nèi)糖基肌醇磷酸神經(jīng)酰胺代謝的紊亂導(dǎo)致根部細胞增殖過程減少,使得npc4突變體根長變短[66]。
2.4 鞘脂代謝調(diào)控植物花粉發(fā)育
SPT在調(diào)控雄配子體發(fā)育過程中起著重要作用[10, 116]。伏馬菌素耐藥突變體fbr11-2被報道是比lcb1-1功能更強的等位突變,在生殖發(fā)育過程中表現(xiàn)出更嚴(yán)重的表型。fbr11-2突變體的花粉在進行第二次有絲分裂的過程中,在雙核小孢子中發(fā)生了以細胞核DNA斷裂為特征的PCD,隨后在三核期發(fā)生細胞質(zhì)萎縮和細胞器退化,最終導(dǎo)致形成的花粉粒具有異常的內(nèi)膜系統(tǒng)[116]。編碼LCB2的AtLCB2a和AtLCB2b單個基因發(fā)生突變均沒有明顯的植物生長發(fā)育表型和鞘脂含量的變化,但是兩個基因同時突變后小孢子的發(fā)育受到影響,導(dǎo)致花粉在發(fā)育早期喪失活力,花粉在結(jié)構(gòu)上表現(xiàn)出內(nèi)膜系統(tǒng)異常,比如內(nèi)質(zhì)網(wǎng)和高爾基體膜的破壞以及花粉壁內(nèi)層的缺失等[10]。在atlcb2a-1突變背景下利用化學(xué)試劑誘導(dǎo)型基因沉默系統(tǒng)誘導(dǎo)AtLCB2b基因沉默會有致死性的表型,進一步說明了AtLCB2a和AtLCB2b在功能上冗余[10]。此外,擬南芥ssSPTa基因突變也會導(dǎo)致花粉活力喪失,并且ssSPTa和ssSPTb之間也存在功能冗余[75]。水稻中無論ORMDL基因缺失還是ORMDL基因表達降低的3種RNAi轉(zhuǎn)基因植株均雄性不育,其花粉形態(tài)和活力均異常。在水稻突變體的小穗中,鞘氨醇的整體水平?jīng)]有顯著變化,但占主導(dǎo)地位的神經(jīng)酰胺(d18∶2 c20∶1)減少了約30%,表明植物ORMDL基因通過調(diào)節(jié)鞘脂穩(wěn)態(tài)影響花粉發(fā)育,但是更確切的調(diào)控機制仍然未知[11]。
擬南芥花粉管的發(fā)育受到多種調(diào)控因素的影響。SPHK基因的敲除會降低體內(nèi)花粉管的生長速度,過表達SPHK基因則會促進擬南芥花粉管的生長,鞘氨醇-1-磷酸通過介導(dǎo)Ca2+內(nèi)流調(diào)節(jié)花粉管生長[117]。堿性神經(jīng)酰胺酶TOD1功能缺失會導(dǎo)致擬南芥體內(nèi)花粉管生長緩慢,結(jié)實率降低,TOD1基因僅在成熟花粉、花粉管和角果表皮的保衛(wèi)細胞中特異性表達,tod1突變體花粉管生長缺陷可能是由于自身膨壓調(diào)節(jié)異常所導(dǎo)致的[63],并且被子植物中TOD1基因在花粉管生長中功能保守[118]。GCS/gcs-1植株表現(xiàn)出花粉傳遞缺陷,其花粉活力和花粉萌發(fā)沒有顯著差異,花粉表型可能是由于花粉管向卵細胞生長過程缺陷所致[12]。擬南芥IPUT1在花粉發(fā)育過程中具有重要作用,無法獲得純合的iput1突變體[119]。將pLAT52∶IPUT1轉(zhuǎn)入iput1/+突變體中,通過草銨膦(Basta)篩選兩代轉(zhuǎn)基因植株后獲得純合的pLAT52∶IPUT1 iput1/iput1突變體。該突變體表現(xiàn)出嚴(yán)重的矮化特征,并且存在花粉管導(dǎo)向缺陷[49, 119],對IPUT1的進一步研究將有助于更精確地認(rèn)識糖基肌醇磷酸神經(jīng)酰胺如何參與花粉管導(dǎo)向調(diào)控。
2.5 鞘脂調(diào)控植物形態(tài)的建成和維持
鞘脂參與植物早期形態(tài)建成和后期植株形態(tài)維持。編碼SPT亞基的AtLCB1基因突變后,lcb1-1突變體表現(xiàn)出胚胎致死的表型,但通過RNAi的方法降低AtLCB1的表達水平,使得植物細胞變小,最終導(dǎo)致植株整體變小[32]。在正常的生長條件下,sbh1-1和sbh2-1突變體植株沒有明顯的生長缺陷表型,但當(dāng)二者同時突變后,sbh1-1 sbh2-1雙突變體完全缺乏植物鞘氨醇,植株嚴(yán)重矮化,無法從營養(yǎng)生長過渡到生殖生長階段,并且PCD相關(guān)基因的表達增強[97]。通過RNAi方法構(gòu)建SBH1和SBH2基因同時抑制的突變體來評估植物鞘氨醇對植物生長的影響,檢測發(fā)現(xiàn)植株的矮化程度與植物鞘氨醇的缺失程度相關(guān),當(dāng)葉片中植物鞘氨醇的相對含量小于等于總鞘氨醇含量的35%時,即植物鞘氨醇含量相對較低時,植株的生長受到抑制[97]。loh1loh3雙突變體沒有固定的植株形態(tài),在固體培養(yǎng)基上延長培養(yǎng)時間,部分loh1loh3雙突變體種子能萌發(fā)幼苗,但其形態(tài)嚴(yán)重變形,沒有根、葉片形狀變形,無法進一步生長發(fā)育,表明由LOH1和LOH3催化的含VLCFAs的鞘脂對植物幼苗發(fā)育至關(guān)重要[38]。但過表達LOH1和LOH3導(dǎo)致植物整體較野生型大,可能是由于含VLCFAs的植物神經(jīng)酰胺積累促進了細胞分裂和生長[120]。在擬南芥中過表達LOH2基因?qū)е轮仓臧?,并且積累神經(jīng)酰胺(C16-FA),進而引起SA的水平升高導(dǎo)致PCD的組成性表達[120]。雖然鞘氨醇和神經(jīng)酰胺在植物中的含量較少,但在維持植物生長發(fā)育和調(diào)節(jié)鞘脂含量,以及在相關(guān)的激素途徑或者防御途徑中具有重要的作用。
GCS基因缺失會導(dǎo)致擬南芥幼苗嚴(yán)重矮化,并且子葉畸形幾乎不能形成初葉,細胞內(nèi)高爾基體形態(tài)異常[12]。雖然gcs-1突變體幼苗能夠形成愈傷組織,并在含有蔗糖的培養(yǎng)基上增殖生長到8周左右,但將愈傷組織轉(zhuǎn)移至具有促進分化的培養(yǎng)基上時,愈傷組織細胞不能分化出器官,表明葡糖神經(jīng)酰胺對擬南芥的細胞類型分化和器官發(fā)生至關(guān)重要[12]。向高爾基體腔轉(zhuǎn)運GDP-Man促進糖基肌醇磷酸神經(jīng)酰胺合成的GONST1基因突變也會導(dǎo)致植株矮化的表型,并且伴隨結(jié)實率降低,植株SA水平升高和PCD的發(fā)生[27]。gonst1通過與SA積累能力受損的植株(如35S啟動子調(diào)控下表達細菌水楊酸羥化酶基因——NahG的擬南芥植株和SA合成酶Isochorismate Synthase功能缺失的植株——ics1)雜交可部分恢復(fù)矮化的表型,但是gonst1表型不是由脂質(zhì)代謝的改變引起的, 而是由糖基肌醇磷酸神經(jīng)酰胺頭部基團的改變引起的,對下游生長發(fā)育的影響可能是通過SA介導(dǎo)和不依賴于SA的途徑發(fā)揮作用的[27]。與gonst1表型相似,gmt1也表現(xiàn)出矮化的特征且SA和H2O2水平升高。此外gmt1的鮮重約為野生型的一半,其細胞壁的纖維素含量降低、下胚軸長度減少,雖然突變體可完成一個完整的生命周期,但只能產(chǎn)生少量可以存活的種子[50]。gmt1突變體中防御相關(guān)激素的產(chǎn)生可能是上游ROS的過量產(chǎn)生引起的,進而導(dǎo)致了細胞死亡和幼苗期衰老表型的發(fā)生。特異性參與糖基肌醇磷酸神經(jīng)酰胺糖基化的GMT1功能缺失,可能導(dǎo)致糖基肌醇磷酸神經(jīng)酰胺頭部基團的修飾發(fā)生紊亂,誘導(dǎo)了組成性防御反應(yīng),但引起誘導(dǎo)途徑的信號級聯(lián)依舊未知[50]。總體而言,目前發(fā)現(xiàn)涉及糖基肌醇磷酸神經(jīng)酰胺糖基化過程中的多種突變體都具有嚴(yán)重的生長缺陷表型,表明糖基肌醇磷酸神經(jīng)酰胺頭部基團的修飾在植物發(fā)育和信號傳導(dǎo)中具有關(guān)鍵作用,但更詳細的調(diào)控機制需要深入研究。
擬南芥脂質(zhì)翻轉(zhuǎn)酶ALA4和ALA5功能的同時缺失會導(dǎo)致植株嚴(yán)重矮化,且伴隨細胞伸長缺陷,ala4ala5雙突變體中僅葡糖神經(jīng)酰胺的含量顯著性增加,并且積累的葡糖神經(jīng)酰胺類型分布范圍較廣,包括含C16-C26脂肪酸鏈在內(nèi)的所有葡糖神經(jīng)酰胺,可能是鞘脂代謝紊亂導(dǎo)致的植物細胞營養(yǎng)生長失衡引起了ala4ala5雙突變體抑制生長的表型[121]。最近的研究表明,水稻的IPCSs功能缺失會影響株高[122]。水稻具有3個IPCSs同源基因,OsIPCSs單基因突變導(dǎo)致植株矮化,其中osipcs3突變體矮化程度較osipcs1和osipcs2嚴(yán)重,并且多個基因的突變會導(dǎo)致表型加重同時,具有疊加效應(yīng),osipcs1/2/3三突變體生長最弱,導(dǎo)致無法結(jié)實[122]。在矮化的突變體中觀察到節(jié)間表皮細胞伸長顯著減少,在osipcs2/3和osipcs1/2/3突變體中IPC含量顯著降低,總體表明OsIPCSs可能通過影響水稻細胞生長和鞘脂代謝參與了植物株高的調(diào)控[122]。鞘脂代謝調(diào)控植物形態(tài)的研究大多是通過表型進一步推測和檢測植物鞘脂類物質(zhì)含量的變化,來證明二者之間是否有緊密的聯(lián)系,但是缺乏直接的證據(jù)證明植物形態(tài)調(diào)控相關(guān)的基因與鞘脂類物質(zhì)之間的作用機制,因此需要更精確的實驗設(shè)計和方法來完善調(diào)控網(wǎng)絡(luò)。
2.6 鞘脂代謝參與植物果實發(fā)育
橄欖(Olea europaea L. cv. Picual)中鞘脂代謝動態(tài)參與果實的成熟和脫落[123-124]。在橄欖果實發(fā)育早期,鞘氨醇含量增加,鞘脂生物合成關(guān)鍵基因表達上調(diào),并且發(fā)現(xiàn)OeSPHK可能是果實生長早期鞘脂代謝的重要貢獻者[13]。此外,鞘脂與植物激素油菜素內(nèi)酯(Brassinosteroid, BR)在果實早期發(fā)育過程中可能存在相互作用[13]。OeACER是一個成熟誘導(dǎo)基因,與橄欖果實成熟的最后階段相關(guān),內(nèi)源鞘脂水平在果實發(fā)育過程中受到復(fù)雜的調(diào)控[123]。橄欖成熟果實脫落時鞘氨醇總量增加,其中以t18∶1(8E) 的增加最為顯著,大于81%的鞘氨醇是三羥基化的形式,大于87%的鞘氨醇在脫落期含有順式或反式Δ8雙鍵,并且d18∶0在脫落前不存在,但在脫落過程中富集,表明鞘氨醇的 C4羥基化和Δ8去飽和修飾可能在果實脫落過程中介導(dǎo)橄欖離層區(qū)中總鞘脂含量或特定鞘脂類物質(zhì)含量中起重要作用[124]。對桃果實采摘后低溫貯藏過程的研究發(fā)現(xiàn),NO可以增加KDSR、CerS和CERK活性,導(dǎo)致桃果實中鞘氨醇、神經(jīng)酰胺和神經(jīng)酰胺-1-磷酸的含量增加,表明NO能夠維持鞘脂代謝,緩解采摘后果實對低溫的反應(yīng)[125]。目前,關(guān)于鞘脂代謝參與植物果實發(fā)育的研究處于起步階段,主要以鞘脂含量及其相關(guān)基因表達量的變化來間接評估鞘脂在植物果實中發(fā)揮的作用,更深層次的調(diào)控機制未見報道,研究的物種傾向于具有肉質(zhì)果實的植物,相對范圍較小,因此有很大的研究空間急待探究。
3 結(jié)語
植物中結(jié)構(gòu)多樣化的鞘脂類物質(zhì)特異性的分布在不同細胞和組織中,調(diào)節(jié)植物的不同生長發(fā)育過程。以模式植物擬南芥為例,鞘脂調(diào)控擬南芥的幼苗發(fā)育、根的生長、氣孔開閉、細胞程序性死亡、花粉及花粉管的發(fā)育和植株形態(tài)(見圖4)。鞘脂代謝紊亂會導(dǎo)致植物生長發(fā)育過程異常,進而表現(xiàn)出相應(yīng)的生長發(fā)育缺陷的表型。雖然植物鞘脂代謝途徑的多種參與組分已經(jīng)被鑒定,但隨著鞘脂類物質(zhì)研究技術(shù)的不斷更迭,新的成員不斷被發(fā)現(xiàn),新的功能也不斷被揭示。現(xiàn)有鞘脂代謝的主流研究方法是鞘脂基因突變體表型分析和基于液相質(zhì)譜聯(lián)用的鞘脂組學(xué)技術(shù),這些技術(shù)的應(yīng)用可以評估鞘脂代謝相關(guān)基因突變后植物體內(nèi)不同組織中鞘脂類物質(zhì)含量的變化,進而推測出導(dǎo)致植物出現(xiàn)生長發(fā)育缺陷表型的根本原因。雖然通過已有技術(shù)手段可以確定部分鞘脂類突變體出現(xiàn)表型的原因,但對具體的調(diào)控機制仍然缺乏足夠的了解,植物細胞內(nèi)的鞘脂分子在內(nèi)膜系統(tǒng)的轉(zhuǎn)運機制亟需完善。當(dāng)外界的脅迫信號被質(zhì)膜上的受體識別后,質(zhì)膜上是否存在與鞘脂相關(guān)的膜受體,或者如何將信號傳遞給鞘脂分子,細胞內(nèi)的鞘脂信號分子如何響應(yīng)植物生長發(fā)育信號,以及在分子水平上鞘脂如何調(diào)節(jié)植物的激素水平進而影響植物生長發(fā)育等諸多鞘脂相關(guān)研究目前還未明確。鞘脂代謝調(diào)控植物生長發(fā)育的相關(guān)研究將有助于加深植物對環(huán)境適應(yīng)的認(rèn)識,為改善植物營養(yǎng)生長、結(jié)實和貯藏提供新的思路。
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