作物学报 ›› 2017, Vol. 43 ›› Issue (05): 640-647.doi: 10.3724/SP.J.1006.2017.00640
徐熠**,彭阳**,李帅,赵秋棱,张双娟,李加纳,倪郁*
XU Yi**,PENG Yang**,LI Shuai,ZHAO Qiu-Ling,ZHANG Shuang-Juan,LI Jia-Na,NI Yu*
摘要:
在植物蜡质合成途径中,中链烷烃羟化酶(mid-chain alkane hydroxylase, MAH) 催化烷烃羟基化形成二级醇,进一步氧化为酮。本研究以拟南芥P450依赖性酶CYP96A15/MAH1基因为探针,采用电子克隆与RT-PCR技术,获得2个甘蓝型油菜MAH1的全长编码区序列,分别命名为BnMAH1-1和BnMAH1-2 (GenBank登录号分别为KT795344和KT795345)。二者ORF长度均为1491 bp,无内含子,核苷酸与氨基酸序列分别有92.4%与90.9%的一致性。根据编码区预测的BnMAH1-1和BnMAH1-2前体蛋白均为包含496个氨基酸残基的多肽链,具有典型的P450蛋白家族保守结构P415xR417x、K螺旋(E359xxR362)、C末端的血红素结合域(F436xxGxRxCxG445) 以及氧结合带保守区域(A/G)G309x(D/E)T312(T/S)。NCBI BlastN、氨基酸序列多重比对与系统学分析表明, 两者与拟南芥MAH1/CYP96A15同源性最高。实时荧光定量PCR表明,BnMAH1-1与BnMAH1-2主要在甘蓝型油菜茎、叶、花、及角果中表达,其中在叶片中的表达量最高,在根系中的表达量很低,这与角质层蜡质主要沉积在植株地上部分相一致。BnMAH1-1和BnMAH1-2在无蜡粉材料茎、叶片中几乎不表达,表明蜡质的减少与MAH1的转录下调有关。BnMAH1-1与BnMAH1-2受SA、MeJA、ACC、ABA、NaCl及干旱胁迫诱导表达,其中BnMAH1-1可能在水分胁迫响应中起主要作用。
[1]Schreiber L, Skrabs M, Hartmann K D, Diamantopoulos P, Simanova E, Santrucek J. Effect of humidity on cuticular water permeability of isolated cuticular membranes and leaf disks. Planta, 2001, 214: 274–282 [2]Riederer M, Schreiber L. Protecting against water loss: analysis of the barrier properties of plant cuticles. J Exp Bot, 2001, 52: 205–208 [3]Ficke A, Gadoury D M, Godfrey D, Dry I B. Host barriers and responses to Uncinula necator in developing grape berries. Phytopathology, 2004, 94: 438–445 [4]Eigenbrode S D, Rayor L, Chow J, Latty P. Effects of wax bloom variation in Brassica oleracea on foraging by avespid wasp. Entomol Exp Appl, 2000, 97: 161–166 [5]Krauss P, Markst?dter C, Riederer M. Attenuation of UV radiation by plant cuticles from woody species. Plant Cell Environ, 1997, 20: 1079–1085 [6]Ni Y, Xia R E, Li J N. Changes of epicuticular wax induced by enhanced UV-B radiation impact on gas exchange in Brassica napus. Acta Physiol Plant, 2014, 36: 2481–2490 [7]Kunst L, Samuels A L. Biosynthesis and secretion of plant cuticular wax. Prog Lipid Res, 2003, 42: 51–80 [8]Barthhlott W, Neinhuis C. Purity of the sacred lotus, or escape from contamination in biological surface. Planta, 1997, 202: 1–8 [9]Zhou X, Jenks M, Liu J, Liu A, Zhang X, Xiang J, Zou J, Peng Y, Chen X. Overexpression of transcription factor OsWR2 regulates wax and cutin biosynthesis in rice and enhances its tolerance to water deficit. Plant Mol Biol Rep, 2014, 32: 719–731 [10]Al-Abdallat A M, Al-Debei H S, Ayad J Y, Hasan S. Over-expression of SlSHN1 gene improves drought tolerance by increasing cuticular wax accumulation in tomato. Int J Mol Med, 2014, 15: 19499–19515 [11]Lee S B, Kim H, Kim R J, Suh M C. Overexpression of Arabidopsis MYB96 confers drought resistance in Camelina sativa via cuticular wax accumulation. Plant Cell Rep, 2014, 33: 1535–1546 [12]Millar A A, Clemens S, Zachgo S, Giblin M, Taylor D C, Kunst L. CUT1, an Arabidopsis gene required for cuticular wax biosynthesis and pollen fertility, encodes a very-long- chain fatty acid condensing enzyme. Plant Cell, 1999, 11: 825–838 [13]Kolattukudy P E, Liu T Y. Direct evidence for biosynthetic relationships among hydrocarbons, secondary alcohols, and ketones in Brassica oleracea. Biochem Biophys Res Commun, 1970, 41: 1369–1374 [14]Greer S, Wen M, Bird D, Wu X, Samuels L, Kunst L, Jetter R. The cytochrome P450 enzyme CYP96A15 is the mid-chain alkane hydroxylase responsible for formation of secondary alcohols and ketones in stem cuticular wax of Arabidopsis thaliana. Plant Physiol, 2007, 145: 653–667 [15]Pu Y, Gao J, Guo Y, Liu T, Zhu L, Xu P, Yi B, Wen J, Tu J, Ma C, Fu T, Zou J, Shen J. A novel dominant glossy mutation causes suppression of wax biosynthesis pathway and deficiency of cuticular wax in Brassica napus. BMC Plant Biol, 2013, 13: 1471–2229 [16]Liu F, Xiong X, Wu L, Fu D, Hayward A, Zeng X, Cao Y, Wu Y, Li Y, Wu G. BraLTP1, a lipid transfer protein gene involved in epicuticular wax deposition, cell proliferation and flower development in Brassica napus. PloS One, 2014, 9: 1–12 [17]李帅, 赵秋棱, 彭阳, 徐熠, 李加纳, 倪郁. SA、MeJA和ACC处理对甘蓝型油菜叶角质层蜡质组分、结构及渗透性的影响. 作物学报, 2016, 42: 1827–1833 Li S, Zhao Q L, Peng Y, Xu Y, Li J N, Ni Y. Effects of SA, MeJA, and ACC on leaf cuticular wax constituents, structure and permeability in Brassica napus. Acta Agron Sin, 2016, 42: 1827–1833 (in Chinese with English abstract) [18]Durst F, Nelson D R. Diversity and evolution of plant P450 and P450-reductases. Drug Metabol Drug Interact, 1995, 12: 189–206 [19]Nelson D R, Schuler M A, Paquette S M, Werck-Reichhart D, Bak S. Comparative genomics of rice and Arabidopsis: analysis of 727 cytochrome P450 genes and pseudogenes from a monocot and a dicot. Plant Physiol, 2004, 135: 756–772 [20]Lee D S, Nioche P, Hamberg M, Raman C S. Structural insights into the evolutionary paths of oxylipin biosynthetic enzymes. Nature, 2008, 455: 363–368 [21]Schuler M A. Plant Cytochrome P450 monooxygenases. Crit Rev Plant Sci, 1996, 15: 235–284 [22]Chen S, Zhou D. Functional domains of aromatase cytochrome P450 inferred from comparative analyses of amino acid sequences and substantiated by site-directed mutagenesis experiments. J Biol Chem, 1992, 267: 22587–22594 [23]Shepherd T, Wynne Griffiths D. The effects of stress on plant cuticular waxes. New Phytol, 2006, 171: 469–499 [24]Kosma D K, Bourdenx B, Bernard A, Parsons E P, Lü S, Joubès J, Jenks M A. The impact of water deficiency on leaf cuticle lipids of Arabidopsis. Plant Physiol, 2009, 151: 1918–1929 [25]Go Y S, Kim H, Kim H J, Suh M C. Arabidopsis cuticular wax biosynthesis is negatively regulated by the DEWAX gene encoding an AP2/ERF-type transcription factor. Plant Cell, 2014, 26: 1666–1680 [26]Gauvrit C, Gaillardon P. Effect of low-temperatures on 2,4-D behaviour in maize plants. Weed Res, 1991, 31: 135–142 [27]Joubès J, Raffaele S, Bourdenx B, Garcia C, Laroche-Traineau J, Moreau P, Domergue F, Lessire R. The VLCFA elongase gene family in Arabidopsis thaliana: phylogenetic analysis, 3D modelling and expression profiling. Plant Mol Biol, 2008, 67: 547–566 [28]Macková J, Va?ková M, Macek P, Hronková M, Schreiber L, ?antr??ek J. Plant response to drought stress simulated by ABA application: changes in chemical composition of cuticular waxes. Environ Exp Bot, 2013, 86: 70–75 [29]Seo P J, Lee S B, Suh M C, Park M J, Go Y S, Park C M. The MYB96 transcription factor regulates cuticular wax biosynthesis under drought conditions in Arabidopsis. Plant Cell, 2011, 23: 1138–1152 |
[1] | 陈松余, 丁一娟, 孙峻溟, 黄登文, 杨楠, 代雨涵, 万华方, 钱伟. 甘蓝型油菜BnCNGC基因家族鉴定及其在核盘菌侵染和PEG处理下的表达特性分析[J]. 作物学报, 2022, 48(6): 1357-1371. |
[2] | 秦璐, 韩配配, 常海滨, 顾炽明, 黄威, 李银水, 廖祥生, 谢立华, 廖星. 甘蓝型油菜耐低氮种质筛选及绿肥应用潜力评价[J]. 作物学报, 2022, 48(6): 1488-1501. |
[3] | 袁大双, 邓琬玉, 王珍, 彭茜, 张晓莉, 姚梦楠, 缪文杰, 朱冬鸣, 李加纳, 梁颖. 甘蓝型油菜BnMAPK2基因的克隆及功能分析[J]. 作物学报, 2022, 48(4): 840-850. |
[4] | 黄成, 梁晓梅, 戴成, 文静, 易斌, 涂金星, 沈金雄, 傅廷栋, 马朝芝. 甘蓝型油菜BnAPs基因家族成员全基因组鉴定及分析[J]. 作物学报, 2022, 48(3): 597-607. |
[5] | 王瑞, 陈雪, 郭青青, 周蓉, 陈蕾, 李加纳. 甘蓝型油菜白花基因InDel连锁标记开发[J]. 作物学报, 2022, 48(3): 759-769. |
[6] | 王艳花, 刘景森, 李加纳. 整合GWAS和WGCNA筛选鉴定甘蓝型油菜生物产量候选基因[J]. 作物学报, 2021, 47(8): 1491-1510. |
[7] | 李杰华, 端群, 史明涛, 吴潞梅, 柳寒, 林拥军, 吴高兵, 范楚川, 周永明. 新型抗广谱性除草剂草甘膦转基因油菜的创制及其鉴定[J]. 作物学报, 2021, 47(5): 789-798. |
[8] | 唐鑫, 李圆圆, 陆俊杏, 张涛. 甘蓝型油菜温敏细胞核雄性不育系160S花药败育的形态学特征和细胞学研究[J]. 作物学报, 2021, 47(5): 983-990. |
[9] | 周新桐, 郭青青, 陈雪, 李加纳, 王瑞. GBS高密度遗传连锁图谱定位甘蓝型油菜粉色花性状[J]. 作物学报, 2021, 47(4): 587-598. |
[10] | 李书宇, 黄杨, 熊洁, 丁戈, 陈伦林, 宋来强. 甘蓝型油菜早熟性状QTL定位及候选基因筛选[J]. 作物学报, 2021, 47(4): 626-637. |
[11] | 张春, 赵小珍, 庞承珂, 彭门路, 王晓东, 陈锋, 张维, 陈松, 彭琦, 易斌, 孙程明, 张洁夫, 傅廷栋. 甘蓝型油菜千粒重全基因组关联分析[J]. 作物学报, 2021, 47(4): 650-659. |
[12] | 唐婧泉, 王南, 高界, 刘婷婷, 文静, 易斌, 涂金星, 傅廷栋, 沈金雄. 甘蓝型油菜SnRK基因家族生物信息学分析及其与种子含油量的关系[J]. 作物学报, 2021, 47(3): 416-426. |
[13] | 蒙姜宇, 梁光伟, 贺亚军, 钱伟. 甘蓝型油菜耐盐和耐旱相关性状的QTL分析[J]. 作物学报, 2021, 47(3): 462-471. |
[14] | 李倩, Nadil Shah, 周元委, 侯照科, 龚建芳, 刘珏, 尚政伟, 张磊, 战宗祥, 常海滨, 傅廷栋, 朴钟云, 张椿雨. 抗根肿病甘蓝型油菜新品种华油杂62R的选育[J]. 作物学报, 2021, 47(2): 210-223. |
[15] | 魏丽娟, 申树林, 黄小虎, 马国强, 王曦彤, 杨怡玲, 李洹东, 王书贤, 朱美晨, 唐章林, 卢坤, 李加纳, 曲存民. 锌胁迫下甘蓝型油菜发芽期下胚轴长的全基因组关联分析[J]. 作物学报, 2021, 47(2): 262-274. |
|