基于iTRAQ技术的甘蔗受黑穗病菌侵染蛋白组分析

doi:10.3724/SP.J.1006.2019.84001

Abstract

Abstract:

Sugarcane smut has become an important disease affecting sugarcane yield and sugar content. In order to investigate the molecular mechanism of sugarcane responding to smut at protein level, the smut-resistant variety GT29 and the smut-susceptible variety Yacheng 71-374 were used in this study. Both varieties were inoculated with the teliospore suspension of smut pathogen by dipping method whereas the control was treated with sterile water. Leaf samples were collected and used for proteomic analysis by iTRAQ technique at 180 days after treatment. The that 1429 proteins presented in GT29 with quantitative information, including 290 differentially expressed proteins with 153 up-regulated and 137 down-regulated; while 1576 proteins in Yacheng 71-374 with quantitative information, including 125 differentially expressed proteins with 55 up-regulated and 70 down-regulated. The number of differentially expressed proteins in resistant variety was higher than that in susceptible variety, and GT29 enriched more metabolic pathways in KEGG, indicating that the immunomodulatory mechanism of the resistant variety may be more complicated, and the regulatory network involved in response was broader. Through the analysis of photosynthesis, antioxidant system, calcium signal, phenylpropane metabolism, hormone related differential expressed protein and co-owned differentially expressed protein, it was found that the photosynthesis pathway, ROS, ABA, calcium signal pathway related protein were up-regulated in both varieties. The up-regulated expressed proteins were more in GT29 than in Yacheng 71-374, which may be involved in resistance response in sugarcane against smut pathogen at later growth stage. Plant disease resistance is a complex process that requires multiple functions and pathways to participate in regulation. The phenylpropanoid metabolic pathway, enzymes GPX, APX, SOD, GST, and CAT, and hormones IAA, ETH, and GA were not found to be involved in the disease resistance process of sugarcane.

Key words: sugarcane, smut, iTRAQ, proteomics

Qi-Qi SONG,SINGH Pratiksha,Kumar SINGH Rajesh,Xiu-Peng SONG,Hai-Bi LI,You-Ye NONG,Li-Tao YANG,Yang-Rui LI. Proteomic analysis of sugarcane-Sporisorium scitamineum interaction based on iTRAQ technique[J].Acta Agronomica Sinica, 2019, 45(1): 55-69.

Figures/Tables 9

Fig. 1

Table 1

Fig. 2

Supplementary Table 2

Differentially expressed protein GO enrichment analysis for Yacheng 71-374"

分类 Category	GO ID	功能类别 Functional group	P-value
生物过程 Biological process	GO:0006979	Response to oxidative stress	0.0086
生物过程 Biological process	GO:0019222	Regulation of metabolic process	0.0094
	GO:0006560	Proline metabolic process	0.0151
	GO:0006561	Proline biosynthetic process	0.0151
	GO:0060255	Regulation of macromolecule metabolic process	0.0244
	GO:0031323	Regulation of cellular metabolic process	0.0301
	GO:0055114	Oxidation-reduction process	0.0465
分子功能 Molecular function	GO:0004097	Catechol oxidase activity	0.0039
分子功能 Molecular function	GO:0016841	Ammonia-lyase activity	0.0039
	GO:0016682	Oxidoreductase activity, acting on diphenols and related substances as donors, oxygen as acceptor	0.0039
	GO:0004089	Carbonate dehydratase activity	0.0042
	GO:0016684	Oxidoreductase activity, acting on peroxide as acceptor	0.0057
	GO:0004601	Peroxidase activity	0.0057
	GO:0016829	Lyase activity	0.0083
	GO:0016491	Oxidoreductase activity	0.0107
	GO:0016840	Carbon-nitrogen lyase activity	0.0113
	GO:0016209	Antioxidant activity	0.0154
	GO:0020037	Heme binding	0.0174
	GO:0046872	Metal ion binding	0.0187
	GO:0046914	Transition metal ion binding	0.0208
	GO:0043169	Cation binding	0.0242
	GO:0046906	Tetrapyrrole binding	0.0244
	GO:0045735	Nutrient reservoir activity	0.0347
	GO:0016679	Oxidoreductase activity, acting on diphenols and related substances as donors	0.0347
	GO:0004611	Phosphoenolpyruvate carboxykinase activity	0.0347
	GO:0008964	Phosphoenolpyruvate carboxylase activity	0.0347
	GO:0016835	Carbon-oxygen lyase activity	0.0385
细胞组分 Cellular component	GO:0009521	Photosystem	0.0195
细胞组分 Cellular component	GO:0034357	Photosynthetic membrane	0.0266
	GO:0044436	Thylakoid part	0.0266
	GO:0009522	Photosystem I	0.0281
	GO:0009579	Thylakoid	0.0307
	GO:0009538	Photosystem I reaction center	0.0357

Supplementary Table 2

Table 2

Photosynthesis related differentially expressed protein"

蛋白ID Protein ID	蛋白名称 Protein name	差异倍数 Fold change
蛋白ID Protein ID	蛋白名称 Protein name	GT29		Yacheng 71-374
c54318_g1;orf1	光系统I反应中心亚基II Photosystem I reaction center subunit II	1.2995↑		1.2562↑
c69501_g1;orf1	光系统I PsaH, 反应中心亚基III Photosystem I PsaH, reaction centre subunit III	1.9614↑		0.9303
c48673_g1;orf1	光系统I反应中心亚基IV A Photosystem I reaction center subunit IV A	1.1149		1.3041↑
c56023_g1;orf1	光系统I反应中心亚基V Photosystem I reaction center subunit V	1.5387↑		1.0456
c43081_g1;orf1		光系统I PsaH, 反应中心亚基VI Photosystem I PsaH, reaction centre subunit VI	0.8432		1.2040↑
c129327_g1;orf1	光系统I亚基VII Subunit VII of photosystem I	1.1871		1.2700↑
c55121_g1;orf1	光系统I反应中心亚基N Photosystem I reaction center subunit N	1.0602		1.4252↑
c56941_g1;orf1	光系统I反应中心亚基XI Photosystem I reaction center subunit XI	1.3609↑		1.1204
c32636_g1;orf1	铁氧还蛋白[2Fe-2S] Ferredoxin[2Fe-2S]	0.7977↓		1.0785
c70387_g2;orf1	光系统I组装蛋白ycf4 Photosystem I assembly protein ycf4	1.2823↑		0.7661↓
c73345_g1;orf1	叶绿素a-b结合蛋白8 Chlorophyll a-b binding protein 8	0.8225↓		1.1336
c99995_g1;orf1	叶绿素a-b结合蛋白CP26 Chlorophyll a-b binding protein CP26	0.9317		0.7892↓
c52958_g1;orf1	叶绿素a-b结合蛋白 Chlorophyll a-b binding protein	1.3946↑		1.2335↑
c66930_g1;orf1	叶绿素a-b结合蛋白 Chlorophyll a-b binding protein	1.5718↑		1.1641
c69028_g1;orf1	叶绿素a-b结合蛋白 Chlorophyll a-b binding protein	2.4059↑		1.1324
c51419_g1;orf1	叶绿素a-b结合蛋白 Chlorophyll a-b binding protein	1.0915		1.3043↑
c55759_g1;orf1	叶绿素a-b结合蛋白 Chlorophyll a-b binding protein	1.9421↑		1.2397↑
c57415_g1;orf1	叶绿素a-b结合蛋白 Chlorophyll a-b binding protein	1.3181↑		1.0380
c69028_g3;orf1	叶绿素a-b结合蛋白 Chlorophyll a-b binding protein	1.8089↑		1.0351
c55792_g1;orf1	叶绿素a-b结合蛋白 Chlorophyll a-b binding protein	1.8236↑		1.0311
c51241_g4;orf1	叶绿素a-b结合蛋白 Chlorophyll a-b binding protein	1.8423↑		1.0049
c69028_g2;orf1	叶绿素a-b结合蛋白 Chlorophyll a-b binding protein	1.9144↑		0.9817
c121256_g1;orf1	叶绿素a-b结合蛋白 Chlorophyll a-b binding protein	2.4187↑		0.8355
c73247_g2;orf1	光系统II蛋白D1 Photosystem II protein D1	3.4261↑		0.9206
c78076_g1;orf1	光系统II蛋白D2 Photosystem II D2 protein	1.3682↑		0.9089
c54219_g1;orf2	细胞色素b559亚基α Cytochrome b559 subunit α	1.6695↑		0.7424↓
c77580_g1;orf1	光系统II CP43脱辅基蛋白 Photosystem II CP43 chlorophyll apoprotein	1.4055↑		0.9421
蛋白ID Protein ID		蛋白名称 Protein name		差异倍数 Fold change
蛋白ID Protein ID		蛋白名称 Protein name		GT29	Yacheng 71-374
c126236_g1;orf1		放氧增强蛋白1 Oxygen-evolving enhancer protein 1		1.0064	1.2372↑
c59540_g1;orf1		放氧增强蛋白3 Oxygen-evolving enhancer protein 3		0.8754	1.8631↑
c100226_g1;orf1		放氧增强蛋白 Oxygen-evolving enhancer protein		1.5306↑	0.9906
c55903_g1;orf1		叶绿体放氧增强蛋白1 (部分) Chloroplast oxygen-evolving enhancer protein 1 (partial)		1.0456	1.2517↑
c51741_g1;orf1		光系统II PsbR Photosystem II PsbR		1.9037↑	1.0547
c22663_g1;orf1		光系统II 47kDa蛋白 Photosystem II 47 kDa protein		1.7295↑	0.9404
c56158_g2;orf1		细胞色素b6 (部分) Cytochrome b6 (partial)		1.4786↑	1.0816
c72546_g2;orf1		细胞色素f Cytochrome f		1.2540↑	0.9964
c67723_g1;orf1		ATP合成酶CF1α亚基 ATP synthase CF1 α subunit		1.3837↑	1.0172
c57360_g1;orf2		ATP合成酶 ATP synthase		0.6735↓	1.0127
c61429_g2;orf1		叶绿体Ptr-ToxA结合蛋白 Chloroplast Ptr ToxA-binding protein		1.4116↑	0.9843
c54911_g1;orf1		16 kDa膜蛋白 16 kDa membrane protein		1.6364↑	0.7557↓
c47557_g1;orf1		铜蛋白 Blue (type 1) Copper protein Blue (type 1)		1.1170	1.5826↑

Table 2

Table 3

Antioxidant related differentially expressed protein"

蛋白ID Protein ID	蛋白名称 Protein name	差异倍数 Fold change
蛋白ID Protein ID	蛋白名称 Protein name	GT29	Yacheng 71-374
过氧化物酶POD
c50862_g1;orf1	过氧化物酶-5 Peroxiredoxin-5	0.6309↓	0.9930
c67975_g2;orf2	过氧化物酶54 Peroxidase 54	1.1999	2.0388↑
c59322_g3;orf1	过氧化物酶54前体 Peroxidase 54 precursor	1.4571↑	1.5671↑
c67975_g1;orf1	过氧化物酶54前体 Peroxidase 54 precursor	1.4967↑	2.0387↑
c66988_g1;orf1	过氧化物酶12前体 Peroxidase 12 precursor	1.5653↑	1.0408
c53549_g1;orf1	过氧化物酶-2E-1 Peroxiredoxin-2E-1	0.8258↓	0.9233
c59322_g4;orf1	过氧化物酶 Peroxidase	1.4017↑	0.9072
c59146_g1;orf1	过氧化物酶 Peroxidase	0.7301↓	0.6919↓
c72776_g2;orf1	过氧化物酶 Peroxidase	1.1594	0.6714↓
蛋白ID Protein ID	蛋白名称 Protein name	差异倍数 Fold change
蛋白ID Protein ID	蛋白名称 Protein name	GT29		Yacheng 71-374
c58718_g1;orf1	过氧化物酶 Peroxidase	0.4462↓		0.4493↓
c67818_g4;orf1	过氧化物酶 Peroxidase	1.3312↑		1.6457↑
c64648_g9;orf1	过氧化物酶 Peroxidase	2.7030↑		1.4510↑
c70898_g1;orf2	过氧化物酶 Peroxidase	1.2262↑		1.3997↑
c50276_g1;orf1	过氧化物酶 Peroxidase	1.2289↑		1.3558↑
c58269_g2;orf1	过氧化物酶 Peroxidase	1.4748↑		1.2872↑
c46626_g1;orf1	过氧化物酶 Peroxidase	1.4542↑		1.2559↑
c70898_g1;orf1	过氧化物酶 Peroxidase	1.3312↑		1.2195↑
谷胱甘肽过氧化物酶GPX
c73435_g2;orf1	谷胱甘肽过氧化物酶 Glutathione peroxidase	0.8064↓		1.0474
c55250_g2;orf1	磷脂氢过氧化物谷胱甘肽过氧化物酶6 Probable phospholipid hydroperoxide glutathione peroxidase 6	0.7848↓		0.9799
抗坏血酸过氧化物酶APX
c79083_g1;orf1	L-抗坏血酸过氧化物酶 L-ascorbate peroxidase	0.7024↓		0.9451
c113714_g1;orf1	APx1-细胞溶质抗坏血酸过氧化物酶 APx1-cytosolic Ascorbate Peroxidase	0.7028↓		0.9186
c76142_g1;orf1	抗坏血酸过氧化物酶 Ascorbate peroxidase	0.7592↓		0.9356
c64041_g1;orf1	抗坏血酸过氧化物酶 Ascorbate peroxidase	0.7935↓		0.9019
c62865_g1;orf1	抗坏血酸过氧化物酶 Ascorbate peroxidase	0.9393		0.7810↓
超氧化物歧化酶SOD
c73325_g1;orf1	超氧化物歧化酶(Cu/Zn) Superoxide dismutase (Cu/Zn)	0.6040↓		1.4209↑
c73325_g2;orf2	超氧化物歧化酶(Cu/Zn) Superoxide dismutase (Cu/Zn)	0.4810↓		1.1765
c55722_g2;orf1	超氧化物歧化酶 Superoxide dismutase	0.5893↓		1.0278
谷胱甘肽硫转移酶GST
c66937_g2;orf2	谷胱甘肽S-转移酶4 Glutathione S-transferase 4	0.4706↓		1.0162
c71423_g2;orf2	谷胱甘肽S-转移酶F8 Glutathione S-transferase F8	0.7288↓		1.3394↑
c57078_g1;orf1	谷胱甘肽S-转移酶GST 12 Glutathione S-transferase GST 12	1.0751		1.2122↑
c47481_g1;orf1	谷胱甘肽S-转移酶GST 14 Glutathione S-transferase GST 14	0.9491		1.3272↑
c49019_g1;orf1	谷胱甘肽S-转移酶 Glutathione S-transferase	0.9292		1.2484↑
c125211_g1;orf1	谷胱甘肽S-转移酶 Glutathione S-transferase	1.1719		1.2102↑
c67259_g1;orf1	谷胱甘肽S-转移酶 Glutathione S-transferase	0.6751↓		1.0939
c66937_g2;orf1	谷胱甘肽S-转移酶 Glutathione S-transferase	0.5350↓		0.9349
c65037_g3;orf1	谷胱甘肽S-转移酶 Glutathione S-transferase	1.2785↑		0.9165
c66547_g1;orf2	蛋白IN2-1同系物B同型X2 Protein IN2-1 homolog B isoform X2	0.6158↓		1.0739
过氧化氢酶CAT
c63190_g2;orf1	过氧化氢酶 Catalase-1-like	0.7984↓		0.9734
c63190_g2;orf2	过氧化氢酶 Catalase-1-like	0.8116↓		0.8982
c63190_g1;orf1	过氧化氢酶 Catalase	1.2415↑		0.7038↓

Table 3

Fig. 3

Table 4

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品种 Variety	总谱图数 Total profiles	鉴定谱图数 Identified profiles	鉴定肽段数 Identified peptides	鉴定蛋白数 Identified proteins	定量蛋白数 Quantified proteins
GT29	112 817	8009	3512	1452	1429
Yacheng 71-374	114 550	8832	3990	1594	1576

分类 Category	GO ID	功能类别 Functional group	P-value
生物过程 Biological process	GO:0019684	Photosynthesis, light reaction	0
生物过程 Biological process	GO:0009765	Photosynthesis, light harvesting	0.0002
	GO:0015979	Photosynthesis	0.0002
	GO:0009772	Photosynthetic electron transport in photosystem II	0.0156
	GO:0009767	Photosynthetic electron transport chain	0.0167
	GO:0006412	Translation	0.0233
	GO:0006091	Generation of precursor metabolites and energy	0.0283
	GO:0043043	Peptide biosynthetic process	0.0315
	GO:0022900	Electron transport chain	0.0342
	GO:0006518	Peptide metabolic process	0.0347
	GO:0034645	Cellular macromolecule biosynthetic process	0.0375
	GO:0009059	Macromolecule biosynthetic process	0.0375
	GO:0043604	Amide biosynthetic process	0.0381
	GO:0043603	Cellular amide metabolic process	0.0418
	GO:0044267	Cellular protein metabolic process	0.0484
分子功能 Molecular function	GO:0004478	Methionine adenosyltransferase activity	0.0140
分子功能 Molecular function	GO:0004857	Enzyme inhibitor activity	0.0140
	GO:0016859	Cis-trans isomerase activity	0.0145
	GO:0003755	Peptidyl-prolyl cis-trans isomerase activity	0.0145
	GO:0051536	Iron-sulfur cluster binding	0.0216
	GO:0051540	Metal cluster binding	0.0216
	GO:0030234	Enzyme regulator activity	0.0367
	GO:0098772	Molecular function regulator	0.0465
细胞组分 Cellular component	GO:0016020	Membrane	0.0024
细胞组分 Cellular component	GO:0043228	Non-membrane-bounded organelle	0.0386
	GO:0043232	Intracellular non-membrane-bounded organelle	0.0386
	GO:0031224	Intrinsic component of membrane	0.0410

蛋白ID Protein ID	蛋白名称 Annotation	差异倍数 Fold change
蛋白ID Protein ID	蛋白名称 Annotation	GT29	Yacheng 71-374
c62874_g2;orf1	40S核糖体蛋白S5 40S ribosomal protein S5	1.4018↑	0.8042↓
c60900_g1;orf1	40S核糖体蛋白S18 40S ribosomal protein S18	1.4850↑	0.7965↓
c61922_g1;orf1	40S核糖体蛋白S10 40S ribosomal protein S10	2.3718↑	0.7673↓
c53669_g1;orf1	60S核糖体蛋白L35 60S ribosomal protein L35	2.2003↑	0.8276↓
c71804_g5;orf1	NAD(P)H-醌氧化还原酶亚基H NAD(P)H-quinone oxidoreductase subunit H	1.9719↑	0.7819↓
c63190_g1;orf1	过氧化氢酶 Catalase	1.2415↑	0.7038↓
c54219_g1;orf2	细胞色素b559亚基α Cytochrome b559 subunit α	1.6695↑	0.7424↓
c70387_g2;orf1	光系统I组装蛋白ycf4 Photosystem I assembly protein ycf4	1.2823↑	0.7661↓
c58104_g1;orf1	叶绿体光系统II 22kDa蛋白质 Chloroplast photosystem II 22 kDa protein	1.3253↑	0.8263↓
c54911_g1;orf1	16 kDa膜蛋白 16 kDa membrane protein	1.6364↑	0.7557↓
c42694_g1;orf1	C₄磷酸烯醇式丙酮酸羧化酶 Putative C₄ phosphoenolpyruvate carboxylase	1.5936↑	0.7203↓
c42694_g2;orf1	C₄磷酸烯醇式丙酮酸羧化 Putative C₄ phosphoenolpyruvate carboxylase	1.4599↑	0.7465↓
c53868_g3;orf1	C₄磷酸烯醇式丙酮酸羧化酶 Putative C₄ phosphoenolpyruvate carboxylase	1.3213↑	0.7792↓
c107621_g1;orf1	2A型丝氨酸/苏氨酸蛋白磷酸酶65 kDa调节亚基Aβ亚型 Serine/threonine-protein phosphatase 2A 65 kDa regulatory subunit A beta isoform	1.3104↑	0.8029↓
c68372_g4;orf1	γ-生育酚甲基转移酶 γ-tocopherol methyltransferase	1.2630↑	0.7888↓

Proteomic analysis of sugarcane-Sporisorium scitamineum interaction based on iTRAQ technique

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