小麦籽粒次生物质对麦红吸浆虫幼虫解毒酶活性及基因表达的影响

doi:10.3864/j.issn.0578-1752.2020.20.009

Abstract

Abstract:

【Objective】The objective of this study is to clarify the relationship between secondary metabolite contents at grain filling stage of wheat and detoxification enzyme activities of glutathione S-transferase (GST), carboxylesterase (CarE) and cytochrome P450 (CYP450) and related gene expression in Sitodiplosis mosellena, and to explore the role of secondary metabolite in wheat resistance to S. mosellena and the metabolic mechanism.【Method】The numbers of S. mosellana larvae in wheat kernels of four resistant and four susceptible wheat varieties (lines) planted in experimental fields in Zhouzhi County, Shaanxi Province were investigated by dissecting wheat ears in May 2016. Wheat kernels at filling stage from eight varieties (lines) and S. mosellena larvae feeding wheat kernels of eight varieties (lines) were simultaneously collected. The contents of ferulic acid, tannin, total phenol and total flavonoid were determined by colorimetry, vanillin assay, Folin-Ciocalteu assay and sodium nitrite-aluminium nitrite assay, respectively. The activities of GST, CarE and CYP450 and expression levels of GST1, CarE2 and CYP6A1 in collected S. mosellena larvae were determined using enzyme linked immunosorbent assay (ELISA) and qPCR methods, respectively. And then, the relationship between secondary metabolite contents in various wheat varieties (lines) and resistance indictors of wheat to S. mosellena, as well as the interaction of secondary metabolites with detoxification enzyme activity and gene expression in S. mosellena larvae were studied by correlation analysis.【Result】The contents of ferulic acid, tannin, total phenol and total flavonoid of wheat kernel at filling stage of eight wheat varieties (lines) were significantly different. The highest contents were in the resistant variety Kenong 1006 for ferulic acid and tannin, in the resistant variety Jinmai 47 for total phenol, and in the susceptible variety Xinong 88 for total flavonoid. The activities of GST, CarE and CYP450 and expression levels of GST1, CarE2 and CYP6A1 in S. mosellena larvae from eight wheat varieties (lines) were also significantly different. The highest enzyme activities were in larvae from resistant varieties Kenong 1006 and Shaanmai 139 for GST and CarE, respectively, and from the susceptible variety Xiaoyan 6 for CYP450. The expression levels of GST1 and CarE2 were higher in larvae feeding four resistant varieties than those feeding four susceptible varieties, but CYP6A1 expression level had not obvious regularity between larvae feeding resistant and susceptible varieties. Correlation analysis results showed that the ferulic acid content of wheat kernel had significant negative correlation (P<0.05) with the four resistance indicators of wheat to S. mosellena including percentage of infested ears, percentage of infested grains, insect number per ear and estimated loss rate. There was no significant correlation between the contents of tannin, total phenol and total flavonoid and the four resistance indicators. The GST and CarE activities of S. mosellena feeding wheat kernels were significantly positively correlated to the ferulid acid content of wheat kernels, and the CYP450 activity of S. mosellena was significantly negatively correlated to the tannin content (P<0.05). No significant correlation was observed between the total phenol and total flavonoid contents of wheat kernels and the activity of any of the three detoxification enzymes. The expression levels of S. mosellena GST1 and CarE2 were significantly positively correlated with the activities of GST and CarE (P<0.05), respectively, while S. mosellena CYP6A1 expression level was not significantly related with the activity of CYP450.【Conclusion】Ferulic acid is the main secondary metabolite of wheat anti-S. mosellena, and can induce GST and CarE activities of S. mosellena larvae and related gene expression. GST and CarE jointly participate in the detoxification metabolism and adaptation of S. mosellena to wheat secondary metabolites.

Key words: wheat kernel, secondary metabolite, Sitodiplosis mosellena, detoxification enzyme activity, gene expression

CHEN Rui,GAO He,ZHANG GuoJun,ZHU KeYan,CHENG WeiNing. Effects of Secondary Metabolites in Wheat Kernels on Activities of Three Detoxifying Enzymes and Related Gene Expression in Sitodiplosis mosellena[J].Scientia Agricultura Sinica, 2020, 53(20): 4204-4214.

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References 42

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引物名称 Primer name	上游引物 Forward primer (5′-3′)	下游引物 Reverse primer (5′-3′)
GST1	ATAACGGCGTGCTAATAG	AACGAGAATCCACTTGAG
CarE2	AAATGGCTCGGACTCTTTACG	ATGGCTCGGATGCTGACT
CYP6A1	TTACGAAGCGATGATGGA	GCAACTGGTGGATACTTTC
GAPDH	CCATCAAAGCAAGCAAGA	CAGCACGGAGCACAAGAC

小麦品种（系） Wheat variety (line)	穗被害率 Percentage of infested ears (%)	粒被害率 Percentage of infested grains (%)	单穗虫口数 Insect number per ear	估计损失率 Estimated loss rate (%)
陕麦139 Shaanmai 139	30.67±0.88f	3.20±0.35e	1.10±0.14f	0.42±0.05f
科农1006 Kenong 1006	50.33±1.76e	3.75±0.41e	2.30±0.27ef	1.11±0.13e
晋麦47 Jinmai 47	59.67±1.76d	5.44±0.41d	3.61±0.32de	1.80±0.15d
陕农33 Shaannong 33	69.67±2.60c	7.11±0.49c	5.07±0.39d	2.43±0.19d
小偃6号Xiaoyan 6	89.33±0.88b	11.51± 0.75b	7.57±0.53c	4.25±0.30c
西农88 Xinong 88	95.33±1.20a	12.13±0.62b	9.94±0.63b	4.97±0.29b
西农822 Xinong 822	92.00±1.53ab	12.90±0.61b	10.91±0.69b	5.02±0.29b
小偃22 Xiaoyan 22	91.33±1.86ab	15.15±0.73a	14.24±0.88a	6.12±0.36a

小麦品种（系） Wheat variety (line)	阿魏酸含量 Ferulic acid content (mg·g^-1)	单宁含量 Tannin content (mg·g^-1)	总酚含量 Total phenol content (mg·g^-1)	总黄酮含量 Total flavonoid content (mg·g^-1)
陕麦139 Shaanmai 139	6.23±0.09b	1.69±0.05b	24.64±0.85a	2.19±0.08ef
科农1006 Kenong 1006	6.57±0.10a	2.17±0.04a	23.74± 0.15a	2.94±0.06c
晋麦47 Jinmai 47	6.44±0.02ab	1.66±0.03bc	24.86±1.15a	2.67±0.05cd
陕农33 Shaannong 33	6.33±0.11ab	1.28±0.01d	12.56±0.30c	3.29±0.12b
小偃6号 Xiaoyan 6	5.51±0.16c	1.02±0.06e	23.99±1.34a	1.91±0.05f
西农88 Xinong 88	4.77±0.10d	1.55±0.02c	24.09±1.04a	3.93±0.02a
西农822 Xinong 822	4.61±0.04d	1.12±0.01e	14.55±0.61bc	2.39±0.15de
小偃22 Xiaoyan 22	5.72±0.15c	1.55±0.05c	15.83±1.25b	3.44±0.06b

次生物质 Secondary metabolite	穗被害率 Percentage of infested ears		粒被害率 Percentage of infested grains		单穗虫口数 Insect number per ear		估计损失率 Estimated loss rate
次生物质 Secondary metabolite	r	P	r	P	r	P	r	P
阿魏酸 Ferulic acid	-0.774	0.024*	-0.778	0. 023*	-0.726	0.041*	-0.774	0.024*
单宁 Tannin	-0.620	0.101	-0.597	0.118	-0.477	0.232	-0.550	0.158
总黄酮Total flavonoid	0.332	0.422	0.275	0.509	0.378	0.356	0.337	0.415
总酚 Total phenol	-0.392	0.337	-0.438	0.278	-0.495	0.212	-0.426	0.292

小麦品种（系） Wheat variety (line)	GST活性 GST activity (U/mg protein)	CarE 活性 CarE activity (U/mg protein)	CYP450活性 CYP450 activity (U/mg protein)
陕麦139 Shaanmai 139	0.1633±0.0185bcd	0.2284±0.0054a	0.1446±0.0173bc
科农1006 Kenong 1006	0.2144±0.012a	0.1823±0.0103b	0.1124±0.0046c
晋麦47 Jinmai 47	0.1707±0.0242abc	0.1414±0.0087c	0.1248±0.0107c
陕农33 Shaannong 33	0.2038±0.0177ab	0.2116±0.0135a	0.1163±0.0065c
小偃6号 Xiaoyan 6	0.1913±0.0022ab	0.1784±0.0079b	0.1885±0.0038a
西农88 Xinong 88	0.0963±0.0055e	0.1016±0.0071d	0.1304±0.0143c
西农822 Xinong 822	0.1206 ±0.0101de	0.1074±0.0037d	0.1791±0.0217ab
小偃22 Xiaoyan 22	0.1333±0.0064cde	0.1242±0.0038cd	0.1306±0.0001c

Effects of Secondary Metabolites in Wheat Kernels on Activities of Three Detoxifying Enzymes and Related Gene Expression in Sitodiplosis mosellena

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解毒酶 Detoxification enzyme	阿魏酸Ferulic acid		单宁Tannin		总酚 Total phenol		总黄酮 Total flavonoid
解毒酶 Detoxification enzyme	r	P	r	P	r	P	r	P
GST	0.805	0.016*	0.229	0.586	0.083	0.845	-0.375	0.361
CarE	0.717	0.045*	0.167	0.693	0.114	0.788	-0.412	0.311
CYP450	-0.608	0.110	-0.742	0.035*	-0.003	0.995	-0.682	0.063

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