OsMAX1a and OsMAX1e, Involved in the Biosynthesis of Strigolactones,Regulate Rice Tillering

doi:10．3969/j．issn．1001G7216．2015．03．001

Abstract

Abstract:

Strigolactones (SLs) are a class of phytohormone which play essential roles in regulating rice tillering, and ATMAX1 in Arabidopsis encodind P450 is an important gene involved in the synthesis of SLs.Our study found five MAX1 homologous genes through BLAST, namely OsMAX1a, OsMAX1b, OsMAX1c, OsMAX1d and OsMAX1e. Among them, OsMAX1a and OsMAX1e have stable and high expression in rice tissues. Our experiments verified that the overexpression of OsMAX1aand OsMAX1e in Arabidopsis rescued the max1 branched mutant phenotype, and the interference of OsMAX1a and OsMAX1e expression in rice increased rice tillering number. Furthermore, enogenous GR24 application inhibited the expression of OsMAX1a and OsMAX1e, and P-starvation enhanced the expression of OsMAX1a and OsMAX1e,thus promoted the biosynthesis of SLs. In addition, IAA and Kinetin(a cytokinin:KT) treatment apparently had remarkable effect on the expression of OsMAX1a and OsMAX1e. The results demonstrated that the expression of OsMAX1a and OsMAX1e responded to the SLs level and they were involved in the synthesis of SLs and regulated rice tillering.

Key words: strigolactones, OsMAX1a, OsMAX1e, tillering

摘要：

独角金内酯是调控水稻分蘖的重要激素,在拟南芥中AtMAX1编码细胞色素P450蛋白,是独角金内酯合成途径的重要基因。通过同源序列比对,发现在水稻中有5个MAX1同源基因,包括OsMAX1a、OsMAX1b、OsMAX1c、OsMAX1d和OsMAX1e。分析5个OsMAX1基因的转录水平,发现OsMAX1a和OsMAX1e基因能够在水稻组织中稳定高效表达。对OsMAX1a和OsMAX1e的基因功能进行了验证,在拟南芥中过量表达OsMAX1a和OsMAX1e基因能够使max1突变体多分支表型恢复正常;在水稻中干涉OsMAX1a和OsMAX1e基因的表达能够增加水稻分蘖数。进一步的研究表明,外施GR24能够抑制OsMAX1a和OsMAX1e基因的表达,磷饥饿条件能够提高两个基因的表达水平从而促进独角金内酯的合成。另外,外施生长素和激动素能够显著影响OsMAX1a和OsMAX1e的表达。因此,OsMAX1a和OsMAX1e基因参与了独角金内酯的合成,并且在水稻生长发育中响应植物激素信号的诱导,是调控水稻分蘖的重要基因。

关键词: 独角金内酯, OsMAX1a, OsMAX1e, 分蘖

CLC Number:

Q946
S511.01

Xiu-mei WANG, Yue-yang LIANG, Ling LI, Chang-wei GONG, Hai-peng WANG, Xiao-xi HUANG, Shuang-cheng LI, Qi-ming DENG, Jun ZHU, Ai-ping ZHENG, Ping LI, Shi-quan WANG. OsMAX1a and OsMAX1e, Involved in the Biosynthesis of Strigolactones,Regulate Rice Tillering[J]. Chinese Journal OF Rice Science, 2015, 29(3): 223-231.

王秀梅, 梁越洋, 李玲, 贡常委, 王海鹏, 黄晓西, 李双成, 邓其明, 朱军, 郑爱萍, 李平, 王世全. OsMAX1a, OsMAX1e通过参与独角金内酯的合成调控水稻分蘖[J]. 中国水稻科学, 2015, 29(3): 223-231.

Figures/Tables 9

Fig. 1. Chromosomal location and gene structure of rice OsMAX1 candidate genes.（In Fig. B, black boxes indicate exon and the connecting lines indicate intron.）

Table 1 The OsMAX1 candidate genes in rice.

基因 Gene	登录号 ID	染色体 Chromosome	核苷酸大小 Nucleotide size / aa
OsMAX1a	LOC_Os01g50530	1	412
	LOC_Os01g50520	1
OsMAX1b	LOC_Os06g36920	6	549
OsMAX1c	LOC_Os01g50590	1	517
OsMAX1d	LOC_Os01g50580	1	385
OsMAX1e	LOC_Os02g12890	2	548

Table 2 Sequence of RNAi markers used in the study.

引物 Marker	上游引物 Sence primer	下游引物 Anti-sence primer
RNAi-OsMAX1CON-F	5'-GCGTCGACGTTCTCAAGAGGCTTCGCTG-3'	5'-CGAGCTCGTTCTCAAGAGGCTTCGCTG-3'
RNAi-OsMAX1CON-R	5'-CCCAAGCTTCAAGGTAGGGGAATTTGGTCT-3'	5'-CGGAATTCCAAGGTAGGGGAATTTGGTCT-3'
RNAi-OsMAX1a-F	5'-GCGTCGAC TCCTGAAGGAAGCGATGAGA-3'	5'-CGAGCTC TCCTGAAGGAAGCGATGAGA-3'
RNAi-OsMAX1a-R	5'-CCCAAGCTTCACCTCTGGCTCTGGGAAAT-3'	5'-CGGAATTCCACCTCTGGCTCTGGGAAAT-3'
RNAi-OsMAX1e-F	5'-GCGTCGACGTCCAGCTCGCTGTCCACCA-3'	5'-TCCCCCGGGGTCCAGCTCGCTGTCCACCA-3'
RNAi-OsMAX1e-R	5'-CCCAAGCTTGAGACGAGGTGGAAGCGCATG-3'	5'-CGGAATTCGAGACGAGGTGGAAGCGCATG-3'

Fig. 2. Transcriptional analysis of OsMAX1.

Fig.3. Overexpression of OsMAX1a and OsMAX1e gene in Arabidopsis max1. （A and D, Over-expression of OsMAX1a in Arabidopsis max1; B and E, Over-expression of OsMAX1e in Arabidopsis max1; C and F, Arabidopsis max1.）

Fig. 4. PCR analysis of transgenic rice plants.（+, Positive control; -, Negative control; 1-12, Transgenic rice plants.）

Fig. 5. Tiller number and height comparison of Nipponbare, OsMAX1a and OsMAX1e gene interference in rice.（OsMAX1CON-RNAi, The conserved region interference of OsMAX1 gene.）

Fig. 6. Expression levels of OsMAX1a and OsMAX1e as affected by GR24 application and P-deficiency.

Fig. 7. Expression levels of OsMAX1a and OsMAX1e after IAA, KT, NPA treatment.

References 35

[1]	Ruyter-Spira C, Al-Babili S, van der Krol S, et al. The biology of strigolactones.Trends Plant Sci, 2013, 18: 72-83 .
[2]	Wang Y H, Li J Y.Branching in rice.Plant Biol, 2011, 14: 94-99.
[3]	Domagalska M A, Leyser O.Signal integration in the control of shoot branching.Cell Biol, 2011(2): 211-221.
[4]	Cardoso C, Ruyter-Spira C, Bouwmeester H J, et al.Strigolactones and root infestation by plant-parasitic Striga, Orobanche and Phelipanche spp.Plant Sci, 2011, 180(3): 414-420.
[5]	Dun E A, Brewer P B, Beveridge C A, et al.Strigolactones: Discovery of the elusive shoot branching hormone.Trends Plant Sci, 2009, 14: 364-372.
[6]	Stirnberg P, van De Sande K, Leyser H M. MAX1 and MAX2 control shoot lateral branching in Arabidopsis.Development, 2002, 129: 1131-1141.
[7]	Umehara M, Hanada A, Yoshida S, et al.Inhibition of shoot branching by new terpenoid plant hormones.Nature, 2008, 455: 195-200.
[8]	Ruyter-Spira C, Al-Babili S, van der Krol, et al. The biology of strigolactones.Trends Plant Sci, 2013, 18: 72-83.
[9]	Woo H R, Chung K M, Park J H, et al.ORE9, an F-box protein that regulates leaf senescence in Arabidopsis.Plant Cell, 2001, 13: 1779-1790.
[10]	Gomez-Roldan, Fermas V S, Brewer P B, et al.Strigolactone inhibition of shoot branching.Nature, 2008, 455: 189-194.
[11]	Snowden K C, Simkin A J, Janssen B J, et al.The decreased apical dominance1/petunia hybrid CAROTENOID CLEAVAGE DIOXYGENASE 8 gene affects branch production and plays a role in leaf senescence, root growth, and flower development.Plant Cell, 2005, 17: 746-759.
[12]	Cook C G, Whichard L P, Turner B, et al.Germination of witchweed (Striga lutea Lour.): Isolation and properties of a potent stimulant.Science, 1966, 154: 1189-1190.
[13]	Akiyama K, Matsuzaki K, Hayashi H, et al.Plant sesquiterpenes induce hyphal branching in arbuscular mycorrhizal fungi.Nature, 2005, 435: 824-827.
[14]	Umehara M, Hanada A, Yoshida S, et al.Inhibition of shoot branching by new terpenoid plant hormones.Nature, 2008, 455: 195-200.
[15]	Kohlen W, Charnikhova T, Lammers M, et al.Strigolactones are transported through the xylem and play a key role in shoot architectural response to phosphate deficiency in nonarbuscular mycorrhizal host Arabidopsis.Plant Physiol, 2011, 155(2): 974-987.
[16]	Zhou F, Lin Q B, Zhu L H, et al.D14-SCFD3-dependent degradation of D53 regulates strigolactone signalling.Nature, 2013, 504: 7480.
[17]	Zou J, Zhang S Y, Zhang W P, et al.The rice HIGH-TILLERING DWARF1 encoding an ortholog of Arabidopsis MAX3 is required for negative regulation of the outgrowth of axillary buds.Plant J, 2006, 48(5): 687-698.
[18]	Arite T, Umehara M, Ishikawa S, et al.d14, a strigolactone-insensitive mutant of rice, shows an accelerated outgrowth of tillers.Plant Cell Physiol, 2009, 50: 1416-1424.
[19]	Lin H, Wang R, Qian Q, et al.DWARF27, an iron-containing protein required for the biosynthesis of strigolactones, regulates rice tiller bud outgrowth.Plant Cell, 2009, 21: 1512-1525.
[20]	Arite T, Iwata H, Ohshima K, et al.DWARF10, an RMS1/MAX4/DAD1 ortholog, controls lateral budoutgrowth in rice.Plant J, 2007, 51: 1019-1029.
[21]	Jiang L, Liu X, Xiong G S, et al.DWARF 53 acts as a repressor of strigolactone signalling in rice.Nature, 2013, 504(7480): 401-405.
[22]	Booker J, Auldridge M, Wills S, et al.MAX3/CCD7 is a carotenoid cleavage dioxygenase required for the synthesis of a novel plant signaling molecule.Curr Biol, 2004, 14: 1232-1238.
[23]	Sorefan K, Booker J, Haurogné K, et al.MAX4 and RMS1 are orthologous dioxygenase-like genes that regulate shoot branching in Arabidopsis and pea.Genes Dev, 2003, 17: 1469-1474.
[24]	Beveridge C A, Symons G M, Turnbull C G.Long-distance signaling and a mutation analysis of branching in pea.Plant Growth Regul, 2000, 32: 193-203.
[25]	Hamiaux C, Drummond R S, Janssen B J, et al.DAD2 is an a/b hydrolase likely to be involved in the perception of the plant branching hormone, strigolactone.Curr Biol, 2012, 22: 2032-2036.
[26]	Huang J Q, Wei Z M, An H L, et al.Agrobacterium tumefaciens-mediated transformation of rice with the spider insecticidalgene conferring resistance to leaffolder and striped stem borer.Cell Res, 2001, 11: 149-155.
[27]	Hiei Y, Ohta S, Komari T, et al.Efficient transformation of rice (Oryza sativa L.) mediated by Agrobacterium and sequence analysis of the boundaries of the T-DNA.Plant J, 1994, 6: 271-282.
[28]	许红梅, 张立军, 刘淳. 农杆菌蘸花法侵染拟南芥的研究. 北方园艺, 2010, 14: 143-146.
[29]	Yoshid A S, Forno D A, Cock J H, et al.Laboratory manual for physiological studies of rice. 3rd ed. Manila, Philippines: IRRI, 1976: 61-64.
[30]	Turnbull C G N, Booker J P, Leyser H M O. Micrografting techniques for testing long-distance signalling in Arabidopsis.Plant J, 2002, 32: 255-262.
[31]	Booker J, Sieberer T, Wright W, et al.MAX1 encodes a cytochrome P450 family member that acts downstream of MAX3/4 to produce a carotenoid-derived branch-inhibiting hormone.Dev Cell, 2005, 8: 443-449.
[32]	Alder A, Holdermann I, Beyer P, et al.Carotenoid oxygenases involved in plant branching catalyse a highly specific conserved apocarotenoid cleavage reaction.Biochem J, 2008, 416: 289-296.
[33]	Alder A, Jamil M, Marzorati M, et al.The path from b-carotene to carlactone, astrigolactone-like plant hormone.Science, 2012, 335: 1348-1351.
[34]	Drummond R S, Sheehan H, Simons J L, et al.The expression of petunia strigolactone pathway genesis altered as part of the endogenous developmental program.Front Plant Sci, 2012, 2: 115.
[35]	Yoneyama K, Takeuchi T, Sekimoto H, et al.Phosphorus deficiency in red clover promotes exudation of orobanchol, the signal for mycorrhizal symbionts and germination stimulant for root parasites.Planta, 2007, 225(4): 1031-1038.