植物克隆CCD亚基因家族研究综述 本文关键词：克隆，基因，综述，植物，家族
植物克隆CCD亚基因家族研究综述 本文简介：摘要： 类胡萝卜素是植物中一类重要的色素群,经类胡萝卜素裂解双加氧酶(carotenoidcleavagedixoygenases,CCDs)或非酶作用合成的阿朴类胡萝卜素及其衍生物在植物中可以作为着色剂、植物激素、芳香物质和信号物质。CCD基因家族包括CCD和NCED两个亚家族。到目前为止,植物中
植物克隆CCD亚基因家族研究综述 本文内容：
摘 要： 类胡萝卜素是植物中一类重要的色素群, 经类胡萝卜素裂解双加氧酶 (carotenoid cleavage dixoygenases, CCDs) 或非酶作用合成的阿朴类胡萝卜素及其衍生物在植物中可以作为着色剂、植物激素、芳香物质和信号物质。CCD基因家族包括CCD和NCED两个亚家族。到目前为止, 植物中克隆得到的CCD亚基因家族成员有5个, 包括CCD1、CCD2、CCD4、CCD7和CCD8;其中CCD1和CCD4参与了多种植物花、果实的着色及其香气物质 (如α-紫罗酮、β-紫罗酮) 的形成;CCD2仅在藏红花属 (Crocus) 植物中发现, 参与藏红花属植物的花香和花色物质 (如藏花酸) 的形成;CCD7和CCD8参与激素独脚金内酯的合成。本文概述了高等植物中各类CCD亚家族基因成员的结构、表达特性和功能等, 并展望了进一步的研究方向和重点, 以期为该基因家族功能研究和今后的应用提供参考。
关键词： 类胡萝卜素裂解双加氧酶 (CCD) ; 类胡萝卜素降解; 植物激素; 阿朴类胡萝卜素;
Abstract： Carotenoids are one of the most important pigments in plants, apocarotenoids from carotenoids via carotenoids cleavage dioxygenases (CCDs) or non-enzyme pathway and their derivatives play an important role in plants as pigment, plant hormone, volatiles and signals. CCD family consists of two subfamilies: CCD and NCED. To date, five members in CCD subfamily including CCD1, CCD2, CCD4, CCD7 and CCD8 have been cloned from higher plants. It has been proved that CCD1 and CCD4 play important role in color and volatiles (eg, α-ionone and β-ionone) formation of flowers and fruits in plants. CCD2 was only found in Crocus plants, which was involved in the formation of aroma and pigments (eg, crocetin) . CCD7 and CCD8 were involved in formation of strigolactone. In this paper, in order to provide references for the functional research and future application of CCD subfamily genes, the structure, expression patterns and functions of various members of CCD subfamily genes in higher plants are summarized, and further research directions and keynote are prospected.
Keyword： Carotenoid cleavage dixoygenases (CCD) ; Carotenoid degradation; Plant hormone; Apocarotenoid;
类胡萝卜素是广泛分布于自然界中的一类色素, 迄今已发现近800种天然类胡萝卜素 (Johnson, 2009) , 是众多植物果实如:枸杞 (Lycium chinense) (Tian et al., 2015) 、温州蜜柑 (Citrus unshiu) (Kato et al., 2006) 、番茄 (Lycopersicon esculentum) (Simkin et al., 2004a) 和花如:杜鹃 (Rhododendron japonicum) (Ureshino et al., 2016) 、菊花 (Chrysanthemum morifolium) (Yoshioka et al., 2012) 、桂花 (Osmanthus fragrans) (Han et al., 2014) 的重要呈色物质。同时, 类胡萝卜素可以作为合成多种具有生物活性物质的前体, 类胡萝卜素经过氧合酶或非酶裂解作用可以形成阿朴类胡萝卜素 (Hou et al., 2016) ;而阿朴类胡萝卜素及其衍生物可以作为着色剂、香气物质以及调节因子和信号物质等, 在吸引传播媒介 (昆虫、鸟类等) 传播花粉和种子以及调控植物生长发育等方面起重要作用 (Cazzonelli, Pogson, 2010) 。其中, 类胡萝卜素裂解双加氧酶 (carotenoid cleavage dixoygenases, CCDs) 是这些类胡萝卜素裂解氧合酶 (carotenoid cleavage oxygenases, CCOs) 中重要的一支, 参与了植物中多种类胡萝卜素的降解形成β-紫罗酮、α-紫罗酮和独脚金内酯 (strigolactone, SL) (Auldridge et al., 2006b) 。
CCD家族是植物中相对较小的基因家族, 包括CCD和9-顺式-环氧类胡萝卜素双加氧酶基因 (nineepoxycarotenoid dioxygenase, NCED) 两个亚家族 (Ohmiya, 2009) 。第一个被发现的CCD基因家族成员是Schwartz等 (1997) 在玉米 (Zea mays) 中分离获得的属于NCED亚家族的VP14。目前在植物中发现的CCD家族成员共有12个, 其中CCD亚家族有5个 (CCD1, CCD2, CCD4, CCD7和CCD8) ;NCED亚家族7个 (NCED1, NCED2, NCED3, NCED4, NCED5, NCED6和NCED9) 。在拟南芥 (Arabidopsis thaliana) 中, CCD家族共有9个成员, CCD亚家族4个 (CCD1, CCD4, CCD7和CCD8) , NCED亚家族5个 (NCED2, NCED3, NCED5, NCED6, NCED9) (Auldridge et al., 2006a) 。在其他植物中发现的类似基因则根据该基因与拟南芥中CCD基因的亲缘关系和同源性来命名。Walter和Strack (2011) 根据CCD基因家族成员的裂解位点和作用底物的差异性将其分为5个亚家族:CCD1、CCD4、CCD7、CCD8和NCED。但基于基因的同源性、底物特异性以及酶活性差异 (Ohmiya, 2009) , 本文将其分为两个亚家族, 并着重介绍CCD亚家族的研究进展。
1 CCD1基因
1.1 CCD1基因结构
根据起始密码子位置差异, 部分物种中CCD1基因分为两类:CCD1a和CCD1b, 如玉米 (Sun et al., 2008) 、番茄 (Wei et al., 2015) 、藏红花 (Crocus sativus) (Rubio et al., 2008) , 而其他的物种则只有一种CCD1, 如草莓 (Fragaria×ananassa) (Carmen et al., 2008) 。CCD1基因ORF为1 620 bp左右, 其翻译的氨基酸序列长度为540 aa左右 (表1) 。葡萄 (Vitis vinifera) 中VvCCD1基因ORF长度为1629 bp (Mathieu et al., 2005) , 玉米中ZmCCD1基因ORF长度为1 623 bp (Sun et al., 2008) 。从其内含子数目看, 水稻 (Oryza sativa) 和拟南芥CCD1基因分别含有11和13个内含子 (Auldridge et al., 2006b) 。
1.2 CCD1蛋白的作用位点和裂解底物
CCD1在不同植物中裂解的底物、作用位点都不尽相同, 较为普遍的的裂解位点是9, 10 (9'～10') 碳双键。例如青蒿 (Artemisia annua) (Liu et al., 2011) 、玉米 (Sun et al., 2008) 、桂花 (Baldermann et al., 2010) 和温州蜜柑 (Kato et al., 2006) 的CCD1重组蛋白都能裂解β-隐黄质、玉米黄质、以及全反式紫黄质的9, 10和9', 10'位置, 以及9-顺式紫黄质的9, 10位置从而形成β-紫罗兰酮 (β-ionone) 、β-环柠檬醛 (β-cyclocitral) 、香叶基丙酮 (geranylacetone) 和假紫罗兰酮 (pseudoionone) 等芳香类物质。而突厥蔷薇 (Rosa damascena) 的RdCCD1 (Huang et al., 2009a) 、厚皮香瓜 (Cucumis melo) 的CmCCD1 (Ibdah et al., 2006) 、番茄的SlCCD1B (Ilg et al., 2014) 、月桂 (Laurus nobilis) 的LnCCD1 (Yahyaa et al., 2015) 可裂解多种顺式和全反式类胡萝卜素以及不同的阿朴类胡萝卜素生成α-紫罗酮、β-紫罗酮等多种芳香性物质以及6-甲基-5-庚烯-2-酮和香叶醛 (Ilg et al., 2009) 。6-甲基-5-庚烯-2-酮是玉米中的重要挥发性物质, 在番茄红素的裂解位点是5, 6 (5', 6') 碳双键。体外分析中, ZmCCD1裂解线性和环化的类胡萝卜素的效率是相当的。根据阿朴类胡萝卜素挥发性物质的合成模式分析, 有学者认为CCD1是根据碳7和8 (7', 8') 、碳11和12 (11', 12') 之间的饱和状态或在碳5、9和13 (5', 9', 13') 上的甲基集团来识别它的裂解位点 (Vogel et al., 2008) 。有的物种, 如胡萝卜 (Daucus carota) 的DcCCD1, 不能裂解线性胡萝卜素 (Yahyaa et al., 2013) 。例如在水稻中的体内分析表明OsCCD1的作用底物是阿朴类胡萝卜素 (Ilg et al., 2010) 。因为CCD1编码的蛋白并不定位于质体上 (Auldridge et al., 2006a) , 故而在植物体内只能利用由CCD其他家族成员裂解的阿朴类胡萝卜素为底物 (Hou et al., 2016) (表2) 。
表1 植物CCD亚家族基因结构
-:相关信息参考文献中未提及
-:Related information was not mentioned in reference
1.3 CCD1表达特性
CCD1表达的部位主要有花瓣 (Baldermann et al., 2012) 、叶片 (Lashbrooke et al., 2013) 、果实 (Tian et al., 2015) 和柱头 (Rubio et al., 2008) , 在柑橘属 (Citrus) 的物种中, CitCCD1则主要在橘皮和汁囊中表达 (Kato et al., 2006) 。有些物种CCD1则是组成型表达, 如藏红花 (Crocus sativus) 的CsCCD1a (Rubio et al., 2008) 。从表达时期上看, 葡萄的VvCCD1在转色期之前表达量开始升高, 成熟过程中则趋于稳定 (Mathieu et al., 2005) 。类似的, 胡萝卜的DcCCD1 (Yahyaa et al., 2013) 、欧洲越橘 (Vaccinium myrtillus) 的VmCCD1 (Karppinen et al., 2016) 、香瓜的CmCCD1 (Ibdah et al., 2006) 和草莓的FaCCD1 (Carmen et al., 2008) 在根和果实成熟过程中其表达都会上调 (表3) 。
目前的结果表明CCD1的表达主要受光照、昼夜节律、不同光质、激素、NaCl以及高温处理等物理因素和真菌侵染等生物因素的影响。桂花OfCCD1 (Baldermann et al., 2010) 的表达受光周期影响, 白天表达量升高而夜间降低;矮牵牛 (Petunia hybrida) PhCCD1 (Simkin et al., 2004b) 的表达会受到光照的影响, 而越桔 (Vaccinium myrtillus) 的VmCCD1 (Karppinen et al., 2016) 在红光/远红光下表达上调。另外, NaCl、干旱、高温和脱落酸 (abscisic acid, ABA) 处理后碱蓬 (Suaeda salsa) SsCCD1的表达量会升高 (Cao et al., 2005) 。在大豆 (Glycine max) 中, GmCCD1对ABA处理有较强的响应 (Wang et al., 2013) 。另外, 玉米的ZmCCD1在真菌侵染的菌根中表达量会升高 (Sun et al., 2008) 。
1.4 CCD1的功能
CCD1酶活蛋白主要作用是裂解类胡萝卜素生成香气物质 (Floss, Walter, 2009) 。例如在酿酒酵母 (Saccharomyces cerevisiae) 中同时转入了crtYB和PhCCD1后, β-紫罗酮的合成增加了8.5倍 (López et al., 2015) 。相反的, 降低矮牵牛PhCCD1的转录水平导致β-紫罗酮的合成下降了58%～76% (Simkin et al., 2004b) 。
2 CCD2基因
2.1 CCD2基因结构
CCD2是最近新发现的一个CCD家族的成员, 目前只在藏红花属 (Crocus) 的植物中发现 (Ahrazem et al., 2015a) 。最近的研究发现了三类CsCCD2基因, 分别为CsCCD2a、CsCCD2b和CsCCD2-t。其中CsCCD2a含有9个内含子、10个外显子, CsCCD2b含有8个内含子和9个外显子, 而CsCCD2-t没有内含子, 缺少第8外显子 (Ahrazem et al., 2016) 。CaCCD2的cDNA长2238 bp, 编码622个氨基酸 (Ahrazem et al., 2015a) (表1) 。
2.2 CCD2蛋白的作用位点和裂解底物
CCD2的过表达会使藏红花的花被片、柱头呈现黄色、橙色以及红色 (Ahrazem et al., 2016) 。在藏红花柱头中, CsCCD2先后在7-8, 7'-8'碳双键裂解玉米黄质, 经中间产物3'-OH-β-阿朴-8'-胡萝卜醛最终生成藏花酸二醛 (Frusciante et al., 2014) 。
2.3 CCD2表达特性
CsCCD2在藏红花花柱中表达, 在花柱橙色期达到最大值 (Frusciante et al., 2014) 。CaCCD2则主要在花被和柱头中表达 (Ahrazem et al., 2015a) 。CsCCD2启动子中含有光响应以及温度响应元件, 进一步研究发现黑暗以及低温处理会诱导CsCCD2表达;光照和高温抑制CsCCD2的表达 (Ahrazem et al., 2016) (表3) 。
2.4 CCD2的功能
CCD2酶活蛋白是藏红花中藏花酸合成的关键酶, 并进一步糖基化生成西红花苷, 西红花苷是令藏红花花柱中的橙色物质。同时也有研究表明, CsCCD2L和CaCCD2与CCD4一样都定位于质体上的 (Ahrazem et al., 2015a) 。但是, 近来有研究表明在栀子 (Gardenia jasminoides) 中, 参与藏花素合成的最有可能的是CCD4而不是CCD2, 因为在栀子的转录组数据中并未发现CCD2 (Ji et al., 2017) (表2) 。
3 CCD4基因
3.1 CCD4的基因结构
柑橘属 (Citrus) 中根据CCD4保守结构域的差异可以分5个亚支:CitCCD4a、b、c、d和e (Zheng et al., 2015) 。而在菊花中CmCCD4a又可以分为CmCCD4a-1至CmCCD4a-4 (Yoshioka et al., 2012) 。按照有无内含子来分, CCD4亚家族可以分为3类:无内含子、1个内含子和2内含子, 分别以拟南芥的AtCCD4、菊花的CmCCD4和桂花的OfCCD4为代表 (Huang et al., 2009b) 。CCD4的ORF框长度差异性较大, 短的如红木 (Bixa orellana) 的BoCCD4, 只有1053 bp, 编码501个氨基酸 (Sankari et al., 2016) , 长的如OfCCD4, 1827 bp, 可编码609个氨基酸 (Huang et al., 2009b) (表1) 。
表2 CCD亚家族基因功能
3.2 CCD4蛋白的作用位点和裂解底物
菊花CmCCD4和苹果 (Malus×domestica) MdCCD4在大肠杆菌 (Escherichia coli) 中可以裂解β-胡萝卜素生成β-紫罗酮 (Huang et al., 2009b) 。在土豆 (Solanum tuberosum) 中StCCD4也可以在9', 10'位置裂解全反式-β胡萝卜素, 生成β-紫罗酮 (Mark et al., 2015) 。此外, 红木的BaCCD4a与藏红花的CsCCD4a亲缘关系很近, 这表明该基因也可能参与了香气形成, 吸引昆虫以及传粉等过程 (Sankari et al., 2016) 。在蜜柑 (Citrus unshiu) 的果皮中, CitCCD4能在7, 8/7', 8'碳双键裂解β-隐黄质和玉米黄质, 合成β-橙色素, 但不能裂解其他的类胡萝卜素 (Ma et al., 2013) (表2) 。
3.3 CCD4基因的表达特性
与CCD1的表达特点类似, CCD4主要在雄蕊 (Wei et al., 2015) 、柱头 (Rubio et al., 2008) 、花 (Tuan et al., 2013) 、叶片 (Lashbrooke et al., 2013) 和果实 (Ma et al., 2013) 等器官和组织中表达, 如葡萄的VvCCD4a和VvCCD4b分别在叶和果实表达量最高 (Lashbrooke.et al., 2013) 。有些几乎在所有的器官中都表达, 但不同的器官中表达量不同, 如McCCD4, 在花中最高, 嫩叶、茎平稳, 老叶、根中最低 (Tuan et al., 2013) 。有的随植物发育阶段的变化而变化, 如野百合 (Lilium brownii) 的LbCCD4, 花开后12 h表达量达到最高, 并保持高表达量 (Hai et al., 2012) 。有的则是组成成型表达的, 如柑橘CitCCD4a (Zheng et al., 2015) 。CCD4的表达同时也会受到胁迫、光照以及表观修饰等因素的影响, 不过不同物种有所不同。如乙烯和红光处理上调CitCCD4的表达 (Ma et al., 2013) , 而伤害、高温、冷以及渗透胁迫会使藏红花CsCCD4c表达量升高 (Angela et al., 2014) (表3) 。NaCl、聚乙二醇 (polyethylene glycol, PEG) 、高温和低温处理后, 大豆CCD4表达量都呈现先升高后降低的趋势 (Wang et al., 2013) 。在桂花中, '橙红丹桂' (Of.'Chenghong Dangui') OfCCD4启动子CG岛甲基化会下调OfCCD4的表达 (Han et al., 2014) 。
3.4 CCD4基因的功能
除CCD1外, 目前经过报道的CCD家族的酶都是定位于质体上的。如CCD4, 可以利用质体中的类胡萝卜素 (Rottet et al., 2016) 。CCD4对类胡萝卜素的裂解与果肉、花器官的着色有关, 也与香气物质的合成有关。CCD4a/b四个酶可能都与藏红花中β-紫罗酮的形成有关 (Rubio et al., 2008) 。近来有研究表明, 过表达拟南芥AtCCD4基因的水稻中β-胡萝卜素和叶黄素明显降低74%和72%, β-紫罗酮增加了2倍 (Song et al., 2016) 。CCD4功能的正常行使和功能缺失会造成果实、花器官颜色的改变。杜鹃黄色花瓣褪色 (Ureshino et al., 2016) 、桃 (Prunus persica) 果肉呈现白色 (Adami et al., 2013) 、洋桔梗 (Eustoma grandiflorum) (Liu et al., 2013) 浅黄色和白色花的形成以及百合 (Hai et al., 2012) 花被片由黄变白等表型的改变和形成是由CCD4裂解类胡萝卜素造成的。同时, CCD4表达的差异也是造成不同桂花品种花色的关键因素 (Wang et al., 2018) 。相反的, 桃子 (Fukamatsu et al., 2013) 果肉呈现黄色、油菜 (Brassica napus) (Zhang et al., 2015) 和菊花 (Jo et al., 2016) 花色由白变黄, 则是CCD4失活进而导致类胡萝卜素的含量增加造成的 (Adami et al., 2013;Bai et al., 2015) 。CCD4是拟南芥种子类胡萝卜素积累的负调控因子, CCD4功能缺失可以使其β-胡萝卜素的含量增加了8.4倍 (Sabrina et al., 2013) 。在西葫芦 (Cucurbita pepo) 中与类胡萝卜素降解关系最为密切的同样是CCD4 (GonzalezVeldejo et al., 2015) 。这些证据表明, 在桂花、杜鹃和百合等植物中, CCD4是控制它们花色呈现的关键基因。并且已经有报道通过病毒介导沉默桃白色果肉桃中的CCD4后, 基因沉默株系的果肉变黄 (Bai et al., 2015) 。这说明通过基因沉默等技术控制CCD4的表达进而创造不同花色的观赏植物是可行的。
4 CCD7和CCD8基因
4.1 CCD7和CCD8基因的结构特点
CCD7按其外显子数目可以分为两类, 第一类含有6个外显子, 如藏红花CsCCD7 (Rubiomoraga et al., 2014) , 第二类含有7个外显子, 如番茄的SlCCD7 (Vogel et al., 2009) 。CCD8根据外显子数量可分为三类, 第一类只有3个外显子, 如玉米的ZmCCD8;第二类有5个外显子, 如水稻的OsCCD8 (Rubiomoraga et al., 2014) ;第三类有6个外显子, 如土豆的StCCD8 (Pasare et al., 2013) (表1) 。
4.2 CCD7和CCD8蛋白的作用位点和裂解底物
独脚金内酯能抑制植物侧枝萌发, 并且主要在根内合成 (Mikihisa et al., 2008) 。首先, 全反式β-胡萝卜素在β-胡萝卜素顺-反异构酶Dwarf27 (β-carotene cis-trans isomerase Dwarf27, D27) 的作用下生成9-顺式-β-胡萝卜素, 随后CCD7裂解9-顺式-β-胡萝卜素的9'～10'位碳双键生成9-顺式-β-阿朴-10'-胡萝卜醛 (Alder et al., 2012) 。在CCD8的作用下将9-顺式-β-阿朴-10'-胡萝卜醛转变成己内酯 (carlactone) (Bruno et al., 2017) 。己内酯转变为独脚金内酯的机理尚不完全清楚, 在水稻 (Zhang et al., 2014) 和拟南芥 (Booker et al., 2005) 中的研究表明由MAX1编码的细胞色素酶P450与该过程有关。而CCD7和CCD8还参与了另一条代谢通路, 在积累全反式-β-胡萝卜素的大肠杆菌中, AtCCD7和AtCCD8能先后裂解全反式-β-胡萝卜素的9, 10和13, 14碳双键, 生成C18的酮 (β-apo-13-carotenone) (Schwartz et al., 2004) 。不过该酮的结构与独脚金内酯相差甚远, 并且用其处理水稻ccd8突变体后, 并未形成与野生型类似的表型 (Alder et al., 2012) (表2) 。
表3 植物CCD亚家族基因表达特点
4.3 CCD7和CCD8的表达特性
CCD7在不同物种中表达特点类似, 在番茄中SlCCD7在所有组织中均有表达, 在黄色果实中表达量最高 (Wei et al., 2015) , 同样的, 藏红花CsCCD7在所有的组织中也均有表达 (Rubiomoraga et al., 2014) 。玉米的ZmCCD7在根、茎、叶、穗中都表达, 根中最高 (Pan et al., 2016) 。番茄SlCCD8 (Wei et al., 2015) 在所有的组织器官中都有表达, 在根中表达量最高, 其次是茎和老叶。猕猴桃 (Actinidia chinensis) AcCCD8 (Ledger et al., 2010) 则主要在根中表达, 在果实发育早期以及种子中表达量相对较低。类似的, 甘蔗 (Saccharum officinarum) ScCCD8在根尖、分蘖芽、腋芽、茎尖等幼嫩组织以及根中高表达, 而在老叶、节和叶鞘中表达量较低 (吴转娣等, 2016) (表3) 。同时CCD7和CCD8的表达也会受到不同胁迫的影响。如ZmCCD7 (Pan et al., 2016) 和NtCCD8 (Gao et al., 2018) 在缺磷时会上调表达。有研究表明ABA处理下大豆中CCD7和CCD8的表达量都上调 (Wang et al., 2013) 。
4.4 CCD7和CCD8基因的功能
CCD7和CCD8是独脚金内酯合成途径中的两个关键基因 (Mikihisa et al., 2008) 。CCD7和CCD8某个基因或两个基因的表达差异会通过影响独脚金内酯的合成进而影响植物的发育。在百脉根 (Lotus japonicus) 中沉默了LjCCD7的表达后, 与对照组相比, 基因沉默株系有更多侧枝和侧根, 其总生物量也增加了。同时该株系拥有更长的初级根, 并且其衰老速度被明显延缓了 (Liu et al., 2013) 。类似的, 在矮牵牛 (Snowden et al., 2005) 、猕猴桃 (Ledger et al., 2010) 、土豆 (Pasare et al., 2013) 、番茄 (Vogel et al., 2009) 和水稻 (Kulkarni et al., 2014) 等多种植物中CCD7和 (或) CCD8参与调控衰老、根的生长、分枝分蘖和花的发育等多种生命活动。近来, 利用CRISPR/Cas9敲除烟草的CCD8后, 结果发现CRISPR/Cas9介导的ccd8突变体烟草有更多的侧根和侧枝 (Gao et al., 2018) , 这与利用RNAi技术产生的基因沉默植株所产生的表型类似。
5 总结与展望
在类胡萝卜素代谢过程中, CCD亚家族各成员编码的酶发挥了重要的作用。其主要功能如下: (1) 参与香气物质的形成和色彩呈现, 在富含类胡萝卜素的花、块茎和果肉等组织中, 类胡萝卜素含量的多少与颜色的深浅有直接关系, 而CCD4和CCD1参与了类胡萝卜素的降解, 会造成类胡萝卜素含量变化, 进而早成颜色深浅的改变, 同时裂解产物如α-紫罗酮和β-紫罗酮等使该组织呈现独特的风味, 如桂花等花卉; (2) 参与特殊成分的合成, CCD2为藏红花属植物中所独有, 并参与藏花酸、藏花醛等物质的合成, 是造成藏红花属植物特殊风味与花色的关键酶; (3) 参与植物激素的合成, CCD7和CCD8是独脚金内酯合成通路中的关键酶, CCD7和CCD8通过调控独脚金内酯合成进而调控植物侧枝与侧根的发育。
现阶段CCD亚家族基因的研究主要集中在基因表达模式、酶的作用底物、裂解位点以及裂解产物等。同时, 关于CCD亚家族的研究也有不甚清楚的地方。例如, 近来研究中发现的参与调控植物生长发育并响应各种胁迫的阿朴类胡萝卜素信号物质 (apocarotenoid signals, ACSs) , ACS1 (Avenda?o-Vázquez et al., 2014) 。但并未成功分离这些ACSs, 其合成途径也不清楚。并且, 关于CCD基因亚家族启动子和转录因子的研究仍较少。在今后, 关于CCD亚家族的研究可能会包括: (1) CCD亚家族基因的克隆、功能验证; (2) CCD亚家族基因启动子研究; (3) CCD家族基因表达密切相关的转录因子的研究; (4) CCD参与的新型植物生长调节物质合成途径的研究。这些研究的开展有利于更加了解CCD基因家族的作用和分子机理; (5) 利用CRISPR/Cas9和RNAi等技术通过控制CCD家族基因表达, 进而改变植物花色、株型等性状。
参考文献：
吴转娣, 刘新龙, 刘家勇, 等.2016.甘蔗独脚金内酯生物合成基因ScCCD8的克隆与表达分析[J].中国农业科学, 49:2662-2674. (Wu Z T, Liu X L, Liu J Y, et al.2016.Cloing and expression analysis of strigolactones biosynthesis-related gene ScCCD8 in sugarcane[J].Scientia Agriculture Sincia, 49:2662-2674.)
Adami M, Franceshi P D, Brandi F, et al.2013.Identifying a carotenoid cleavage dioxygenase (ccd4) gene controlling yellow/white fruit flesh color of peach[J].Plant Molecular Biology Reporter, 31:1166-1175 (1110) .
Ahrazem O, Rubio-Moraga A, Argando A-Picazo J, et al.2016.Intron retention and rhythmic diel pattern regulation of carotenoid cleavage dioxygenase 2 during crocetin biosynthesis in saffron[J].Plant Molecular Biology, 91:1-20.
[]Ahrazem O, Rubio-Moraga A, Berman J, et al.2015a.The carotenoid cleavage dioxygenase CCD2 catalysing the synthesis of crocetin in spring crocuses and saffron is a plastidial enzyme[J].New Phytologist, 209:650-663.
Ahrazem O, Rubio-Moraga A, Jimeno M L, et al.2015b.Structural characterization of highly glucosylated crocins and regulation of their biosynthesis during flower development in Crocus[J].Frontiers in Plant Science, 6:53-85.
Alder A, Jamil M, Marzorati M, et al.2012.The path fromβ-carotene to carlactone, a strigolactone-like plant hormone[J].Science, 335:1348.
[]Angela R M A, Rambla J L, Fern N D C A, et al.2014.New target carotenoids for CCD4 enzymes are revealed with the characterization of a novel stress-induced carotenoid cleavage dioxygenase gene from Crocus sativus[J].Plant Molecular Biology, 86:555-569.
[]Auldridge M E, Block A, Vogel J T, et al.2006a.Characterization of three members of the Arabidopsis carotenoid cleavage dioxygenase family demonstrates the divergent roles of this multifunctional enzyme family[J].Plant Journal, 45:982-993.
[]Auldridge M E, Mccarty D R, Klee H J.2006b.Plant carotenoid cleavage oxygenases and their apocarotenoid products[J].Current Opinion in Plant Biology, 9:315-321.
Avenda?o-Vázquez O V, Cordoba E, Llamas, E.2014.An uncharacterized apocarotenoid-derived signal generated inζ-carotene desaturase mutants regulates leaf development and the expression of chloroplast and nuclear genes in Arabidopsis[J].Plant Cell, 26:2524-2537.
Bai S, Tuan P A, Tatsuki M, et al.2015.Knockdown of carotenoid cleavage dioxygenase 4 (CCD4) via virus-induced gene silencing confers yellow coloration in peach fruit:Evaluation of gene function related to fruit traits[J].Plant Molecular Biology Reporter:1-8.
Baldermann S, Kato M, Fleischmann P, et al.2012.Biosynthesis ofα-andβ-ionone, prominent scent compounds, in flowers of Osmanthus fragrans[J].Acta Biochimica Polonica, 59:79-81.
Baldermann S, Kato M, Kurosawa M, et al.2010.Functional characterization of a carotenoid cleavage dioxygenase 1and its relation to the carotenoid accumulation and volatile emission during the floral development of Osman?thus fragrans Lour[J].Journal of Experimental Botany, 61:2967-2977.
[]Booker J, Sieberer T, Wright W, et al.2005.MAX1 encodes a cytochrome P450 family member that acts downstream of MAX3/4 to produce a carotenoid-derived branch-inhibiting hormone[J].Developmental Cell, 8:443-449.
Bruno M, Vermathen M, Alder A, et al.2017.Insights into the formation of carlactone from in-depth analysis of the CCD8-catalyzed reactions[J].Febs Letters, 591:792-800.
[]Cao Y, Guo X L, Zhang Q, et al.2005.Isolation and characterization of carotenoid cleavage dioxygenase genein halophyte Suaeda salsa[J].Plant Growth Regulation, 46:61-67.
Carmen G I, Kerstin S, Rosario B, et al.2008.Functional characterization of FaCCD1:A carotenoid cleavage dioxygenase from strawberry involved in lutein degradation during fruit ripening[J].Journal of Agriculture&Food Chemistry, 56:9277-9285.
[]Cazzonelli C I, Pogson B J.2010.Source to sink:Regulation of carotenoid biosynthesis in plants[J].Trends Plant Science, 15:266-274.
[]Floss D S, Walter M H.2009.Role of carotenoid cleavage dioxygenase 1 (CCD1) in apoarotenoid biogenesis revisited[J].Plant Signaling&Behavior, 4:172-175.
Frusciante S, Diretto G, Bruno M, et al.2014.Novel carotenoid cleavage dioxygenase catalyzes the first dedicated step in saffron crocin biosynthesis[J].Proceedings of the National Academy of Sciences of the USA, 111:12246-12251.
[]Fukamatsu Y, Tamura T, Hihara S, et al.2013.Mutations in the CCD4 carotenoid cleavage dioxygenase gene of yellow-flesh peaches[J].Bioscience Biotechnology&Biochemistry, 77:2514-2516.
Gao J, Zhang T, Xu B, et al.2018.CRISPR/Cas9-mediated mutagenesis of carotenoid cleavage dioxygenase 8 (CCD8) in tobacco affects shoot and root architecture[J].International Journal of Molecular Sciences, 19:1062.
Gonzalez-Verdejo C I, Obrero A, Roman B, et al.2015.Expression profile of carotenoid cleavage dioxygenase genes in summer squash (Cucurbita pepo L.) [J].Plant Foods for Human Nutrition, 70:200-206.
Hai N T L, Ji M, Ikuo M, et al.2012.Involvement of carotenoid cleavage dioxygenase 4 gene in tepal color change in Lilium brownii var.colchesteri[J].Journal-Japanese Society for Horticultural Science, 81:366-373.
[]Han Y, Wang X, Chen W, et al.2014.Differential expression of carotenoid-related genes determines diversified carotenoid coloration in flower petal of Osmanthus fragrans[J].Tree Genetics&Genomes, 10:329-338.
Hou X, Rivers J, Le N P, et al.2016.Synthesis and function of apocarotenoid signals in plants[J].Trends in Plant Science, 21:792-803.
[]Huang F C, Horvath G, Molnar P, et al.2009a.Substrate promiscuity of RdCCD1, a carotenoid cleavage oxygenase from Rosa damascena[J].Phytochemistry, 70:457-464.
Huang F C, Molnar P, Schwab W.2009b.Cloning and functional characterization of carotenoid cleavage dioxygenase 4 genes[J].Journal of Experimental Botony, 60:3011-3022.
[]Ibdah M, Azulay Y, Portnoy V, et al.2006.Functional characterization of CmCCD1, a carotenoid cleavage dioxygenase from melon[J].Phytochemistry, 67:1579-1589.
[]Ilg A, Beter P, Al-Babili S.2009.Characterization of the rice carotenoid cleavage dioxygenase 1 reveals a novel route for geranial biosynthesis[J].FEBS Journal, 276:736-747.
[]Ilg A, Bruno M, Beyer P, et al.2014.Tomato carotenoid cleavage dioxygenases 1A and 1B:Relaxed double bond specificity leads to a plenitude of dialdehydes, mono-apocarotenoids and isoprenoid volatiles[J].FEBS Open Bio, 4:584-593.
[]Ilg A, Yu Q, Schaub P, et al.2010.Overexpression of the rice carotenoid cleavage dioxygenase 1 gene in Golden Rice endosperm suggests apocarotenoids as substrates in planta[J].Planta, 232:691-699.
Ji A, Jia J, Xu Z, et al.2017.Transcriptome-guided mining of genes involved in crocin biosynthesis[J].Frontiers in Plant Science, 8:518.
Jo Y D, Kim Y S, Ryu J, et al.2016.Deletion of carotenoid cleavage dioxygenase 4a (CmCCD4a) and global up-regulation of plastid protein-coding genes in a mutant chrysanthemum cultivar producing yellow petals[J].Scientia Horticulturae, 212:49-59.
[]Johnson J D.2009.Do carotenoids serve as transmembrane radical channels?[J].Free Radical Biology&Medicine, 47:321-323.
Karppinen K, Zoratti L, Sarala M, et al.2016.Carotenoid metabolism during bilberry (Vaccinium myrtillus L.) fruit development under different light conditions is regulated by biosynthesis and degradation[J].BMC Plant Biology, 16:1-16.
Kato M, Matsumoto H, Ikoma Y, et al.2006.The role of carotenoid cleavage dioxygenases in the regulation of carotenoid profiles during maturation in citrus fruit[J].Journal of Experimental Botony, 57:2153-2164.
[]Kulkarni K P, Vishwakarma C, Sahoo S P, et al.2014.A substitution mutation in OsCCD7 cosegregates with dwarf and increased tillering phenotype in rice[J].Journal of Genetics, 93:389-401.
Lashbrooke J G, Young P R, Dockrall S J, et al.2013.Functional characterisation of three members of the Vitis vinifera L.carotenoid cleavage dioxygenase gene family[J].BMC Plant Biology, 13:156-173.
[]Ledger S E, Janssen B J, Karunairetnam S, et al.2010.Modified CAROTENOID CLEAVAGE DIOXYGENASE8expression correlates with altered branching in kiwifruit (Actinidia chinensis) [J].New Phytologist, 188:803-813.
[]Liu H, Kishimoto S, Yamamizo C, et al.2013.Carotenoid accumulations and carotenogenic gene expressions in the petals of Eustoma grandiflorum[J].Plant Breeding, 132:417-422.
Liu J, Novero M, Charnikhova T, et al.2013.CAROTENOIDCLEAVAGE DIOXYGENASE 7 modulates plant growth, reproduction, senescence, and determinate nodulation in the model legume Lotus japonicus[J].Journal of Experimental Botony, 64:1967-1981.
[]Liu S Q, Tian N, Liu Z H, et al.2011.Cloning and characterization of a carotenoid cleavage dioxygenase from Arte?misia annua L[J].Applied Mechanics and Materials, 108:274-281.
Lpez J, Essus K, Kim I K, et al.2015.Production ofβ-ionone by combined expression of carotenogenic and plant CCD1 genes in Saccharomyces cerevisiae[J].Microbial Cell Factories, 14:84-97.
[]Ma G, Zhang L, Matsuta A, et al.2013.Enzymatic formation ofβ-citraurin fromβ-cryptoxanthin and Zeaxanthin by carotenoid cleavage dioxygenase 4 in the flavedo of citrus fruit[J].Plant Physiology, 163:682-695.
[]Mark B, Peter B, Salim A B.2015.The potato carotenoid cleavage dioxygenase 4 catalyzes a single cleavage ofβ-ionone ring-containing carotenes and non-epoxidated xanthophylls[J].Archives of Biochemistry and Biophysics, 572:126-133.
Mathieu S, Terrier N, Procureur J, et al.2005.A carotenoid cleavage dioxygenase from Vitis vinifera L.:Functional characterization and expression during grape berry development in relation to C13-norisoprenoid accumulaton[J].Journal of Experimental Botony[J], 56:2721-2731.
Mikihisa U, Atsushi H, Satoko Y, et al.2008.Inhibition of shoot branching by new terpenoid plant hormones[J].Nature, 455:195-200.
Ohmiya A.2009.Carotenoid cleavage dioxygenases and their apocarotenoid products in plants[J].Plant Biotechnology, 26:351-358.
Pan X, Zheng H, Zhao J, et al.2016.ZmCCD7/ZpCCD7 encodes a carotenoid cleavage dioxygenase mediating shoot branching[J].Planta, 243:1407-1418.
[]Pasare S A, Ducreux L J M, Morris W L, et al.2013.The role of the potato (Solanum tuberosum) CCD8 gene in stolon and tuber development[J].New Phytologist, 198:1108-1120
[]Rodrigo M A J, Alqu Z B A E, Vctor M, et al.2013.A novel carotenoid cleavage activity involved in the biosynthesis of Citrus fruit-specific apocarotenoid pigments[J]Journal of Experimental Botony, 64:4461.
Rottet S, Devillers J, Glauser G, et al.2016.Identification of plastoglobules as a site of carotenoid cleavage[J].Frontiers in Plant Science, 7:1855.
Rubio A, Rambla J L, Santaella M, et al.2008.Cytosolic and plastoglobule-targeted carotenoid dioxygenases from Crocus sativus are both involved in beta-ionone release[J].Journal of Biological Chemistry, 283:24816-24825.
Rubiomoraga A, Ahrazem O, Pérez-Clemente R M, et al.2014.Apical dominance in saffron and the involvement of the branching enzymes CCD7 and CCD8 in the control of bud sprouting[J].BMC Plant Biology, 14:171-186.
Sabrina G J, Sun-Hwa H, Maria M L, et al.2013.Carotenoid cleavage dioxygenase4 is a negative regulator ofβ-carotene content in Arabidopsis seeds[J].Plant Cell, 25:4812-4826
Sankari M, Hemachandram H, Anantharaman A, et al.2016.Identifying a carotenoid cleavage dioxygenase 4a gene and its efficient Agrobacterium-mediated genetic transformation in Bixa orellana L[J].Applied Biochemistry&Biotechnology, 179:1-18.
Schmidt H, Kurtzer R, Eisenreich W, et al.2006.The carotenase AtCCD1 from Arabidopsis thaliana is a dioxygenase[J].Journal of Biological Chemistry, 281:9845-9851.
Schwartz S H, Qin X, Loewen M C.2004.The biochemical characterization of two carotenoid cleavage enzymes from Arabidopsis indicates that a carotenoid-derived compound inhibits lateral branching[J].Journal of Biological Chemistry, 279:46940-46945.
Schwartz S H, Tan B C, Gage D A, et al.1997.Specific oxidative cleavage of carotenoids by VP14 of maize[J].Science, 276:1872-1874.
[]Simkin A J, Schwartz S H, Auldridge M, et al.2004a.The tomato carotenoid cleavage dioxygenase 1 genes contribute to the formation of the flavor volatiles beta-ionone, pseudoionone, and geranylacetone[J].Plant Journal, 40:882-892.
Simkin A J, Underwood B A, Auldridge M, et al.2004b.Circadian regulation of the PhCCD1 carotenoid cleavage dioxygenase controls emission of beta-ionone, a fragrance volatile of petunia flowers[J].Plant Physiology, 136:3504-3514.
Snowden K C, Simkin A J, Janssen B J, et al.2005.The decreased apical dominance1/petunia hybrida CAROT-ENOID CLEAVAGE DIOXYGENASE8 gene affects branch production and plays a role in leaf senescence, root growth, and flower development[J].Plant Cell, 17:746-759.
Song M H, Lim S H, Kim J K, et al.2016.In planta cleavage of carotenoids by Arabidopsis carotenoid cleavage dioxygenase 4 in transgenic rice plants[J].Plant Biotechnology Report, 10:291-300.
[]Sun Z, Hans J, Walter M H, et al.2008.Cloning and characterisation of a maize carotenoid cleavage dioxygenase (ZmCCD1) and its involvement in the biosynthesis of apocarotenoids with various roles in mutualistic and parasitic interactions[J].Planta, 228:789-801.
Tian X, Ji J, Gang W, et al.2015.Molecular cloning and characterization of a novel carotenoid cleavage dioxygenase1 from Lycium chinense[J].Biotechnology and Applied Biochemistry, 62:772-779.
[]Tuan P A, Park S U.2013.Molecular cloning and characterization of c DNAs encoding carotenoid cleavage dioxygenase in bitter melon (Momordica charantia) [J].Journal of Plant Physiology, 170:115-120.
Ureshino K, Nakayama M, Miyajima I.2016.Contribution made by the carotenoid cleavage dioxygenase 4 gene to yellow colour fade in azalea petals[J].Euphytica, 207:401-417.
Vogel J T, Tan B C, Mccrty D R, et al.2008.The Carotenoid cleavage dioxygenase 1 enzyme has broad substrate specificity, cleaving multiple carotenoids at two different bond positions[J].Journal of Biological Chemistry, 283:11364-11373.
[]Vogel J T, Walter M H, Giavalisco P, et al.2009.SlCCD7 controls strigolactone biosynthesis, shoot branching and mycorrhiza-induced apocarotenoid formation in tomato[J].The Plant Journal, 61:300-311.
Walter M H, Strack D 2011.Carotenoids and their cleavage products:Biosynthesis and functions[J].Natural Product Reports, 28:663-692.
[]Wang R K, Wang C E, Fei Y Y, et al.2013.Genome-wide identification and transcription analysis of soybean carotenoid oxygenase genes during abiotic stress treatments[J].Molecular Biology Reports, 40:4737-4745.
Wang Y, Zhang C, Dong B, et al.2018.Carotenoid accumulation and its contribution to flower coloration of Osman?thus fragrans[J].Frontiers in Plant Science, 9:1499.
Wei Y, Wan H, Wu Z, et al.2015.A comprehensive analysis of carotenoid cleavage dioxygenases genes in Solanum Ly?copersicum[J].Plant Molecular Biology Reporter, 34:512-523.
Yahyaa M, Bar E, Dubey N K, et al.2013.Formation of norisoprenoid flavor compounds in carrot (Daucus carota L.) roots:Characterization of a cyclic-specific carotenoid cleavage dioxygenase 1 gene[J].Journal of Agricultural&Food Chemistry, 61:12244-12252.
Yahyaa M, Berim A, Isaacson T, et al.2015.Isolation and functional characterization of carotenoid cleavage dioxygenase-1 from Laurus nobilis L. (Bay Laurel) fruits[J].Journal of Agricultural&Food Chemistry, 63:8275-8282.
[]Yoshioka S, Aida R, Yamamizo C, et al.2012.The carotenoid cleavage dioxygenase 4 (CmCCD4a) gene family encodes a key regulator of petal color mutation in chrysanthemum[J].Euphytica, 184:377-387.
[]Zhang B, Liu C, Wang Y, et al.2015.Disruption of a CA-ROTENOID CLEAVAGE DIOXYGENASE 4 gene converts flower colour from white to yellow in Brassica species[J].New Phytologist, 206 (4) :1513-1526.
Zhang Y, Dijk A D J V, Scaffidi A, et al.2014.Rice cytochrome P450 MAX1 homologs catalyze distinct steps in strigolactone biosynthesis[J].Nature Chemical Biology, 10:1028-1033.
Zheng X, Xie Z, Zhu K, et al.2015.Isolation and characterization of carotenoid cleavage dioxygenase 4 genes from different Citrus species[J].Molecular Genetics and Genomics, 290:1-15.
植物克隆CCD亚基因家族研究综述 来源：网络整理
免责声明：本文仅限学习分享，如产生版权问题，请联系我们及时删除。
网站内容来源：网络整理 免责声明：本文仅限学习分享，如产生版权问题，请联系我们及时删除。
Copyright © www.yuan0.cn Corporation, All Rights Reserved 共享时代 共享你我他 版权所有