水稻抗性蛋白OsRRK1抗褐飞虱机理分析

doi:10.16819/j.1001-7216.2020.0406

中国水稻科学 ›› 2020, Vol. 34 ›› Issue (6): 512-519.DOI: 10.16819/j.1001-7216.2020.0406

水稻抗性蛋白OsRRK1抗褐飞虱机理分析

马银花¹, 李萍芳¹, 董文静¹, 易松望¹, 杨芳², 杜波², 金晨钟^1,^*()

¹ 湖南人文科技学院湖南省农田杂草防控技术与应用协同创新中心/农药无害化应用省高校重点实验室, 湖南娄底 417000
² 武汉大学生命科学学院杂交水稻国家重点实验室, 武汉 430072

收稿日期:2020-04-25 修回日期:2020-05-18 出版日期:2020-11-10 发布日期:2020-11-10
通讯作者: 金晨钟
基金资助:
湖南省自然科学基金青年项目（2019JJ50281）;湖南省教育厅优秀青年项目（18B455）;杂交水稻国家重点实验室（武汉大学）开放课题基金资助项目(KF201901);娄底市应用技术与开发项目（33319013）;湖南省应用特色学科（植物保护）资助项目;湖南人文科技学院卓越人才培养计划资助项目（校教通 2017 第 74 号）;湖南省高等学校科技创新团队(201937924)

Mechanism Analysis of Rice Resistance Protein OsRRK1 Againstthe Brown Planthopper

Yinhua MA¹, Pingfang LI¹, Wenjing DONG¹, Songwang YI¹, Fang YANG², Bo DU², Chenzhong JIN^1,^*()

¹ Hunan Provincial Collaborative Innovation Center for Field Weeds Control/ Key Laboratory of Harmless Application of Pesticide in Hunan Higher Education, Hunan University of Humanities, Science and Technology, Loudi417000,China
² College of Life Sciences, Wuhan University /National Key Laboratory of Hybrid Rice,Wuhan 430072,China

Received:2020-04-25 Revised:2020-05-18 Online:2020-11-10 Published:2020-11-10
Contact: Chenzhong JIN

摘要/Abstract

摘要： 目的 OsRRK1（Rop-interacting receptor-like kinase 1）蛋白是水稻中第一个被研究的类胞质受体激酶RLCKⅥ家族蛋白。在调控水稻叶片的卷曲和对褐飞虱的防御中都起着重要作用,但是其调控机理尚不清楚。本研究试图进一步探究OsRRK1基因抗褐飞虱的机理。方法利用酵母双杂交的方法分析OsRRK1蛋白与OsLecRK（lectin-like receptor kinase）、OsLecRK1、OsLecRK2、OsLecRK3的互作关系,OsLecRK是抗褐飞虱基因BPH15区间的候选基因,是水稻先天免疫系统中一个重要成员,既参与水稻先天免疫反应,包括对褐飞虱、白叶枯病和稻瘟病的抗性,又参与水稻的发育过程。OsLecRK1、OsLecRK2、OsLecRK3的基因簇组成抗褐飞虱基因BPH3,含BPH3基因的水稻具有广谱、持久的抗虫性。同时利用DNAMAN软件分析OsLecRK蛋白与OsLecRK1、OsLecRK2、OsLecRK3的同源性。结果 DNAMAN的分析结果显示OsLecRK与OsLecRK1、OsLecRK2、OsLecRK3的同源性都很高,蛋白一致性在50%以上;酵母双杂交结果显示OsRRK1与OsLecRK、OsLecRK2和OsLecRK3互作。结论 OsRRK1通过与抗褐飞虱基因BPH15和BPH3候选基因的相互作用参与水稻抗褐飞虱过程。

关键词: 水稻, OsRRK1, 褐飞虱, 酵母双杂交系统

Abstract:

【Objective】OsRRK1 (Rop-interacting receptor-like kinase 1) protein in rice is the first RLCK VI family protein in rice that has been studied.It plays an important role inregulating the curling of rice leaves and the defense against brown planthoppers. However, the mechanism of its regulation of resistance to the brown planthopperremains unclear. Weattempt to further explore the mechanism of OsRRK1gene resistance to the brown planthoppers.【Method】The interaction between OsRRK1 protein and OsLecRK (lectin-like receptor kinase), OsLecRK1, OsLecRK2 and OsLecRK3 was analyzed by yeast two-hybrid method.OsLecRK is a candidate gene forBPH15 range of anti-brown planthopper, and is an important member of rice innate immune system. OsLecRK is involved innot only rice innate immune response, including resistance to brown planthoppers, white leaf blight and rice blast, but alsothe development of rice. The gene clusters of OsLecRK1, OsLecRK2 and OsLecRK3 constitute the anti-brown planthopper gene BPH3. Rice containing the BPH3 gene has a broad spectrum and lasting insect resistance. Meanwhile, DNAMAN software was used to analyze the homology of OsLecRK with OsLecRK1, OsLecRK2 and OsLecRK3.【Result】The analysis results of DNAMAN showed that OsLecRK has high homology with OsLecRK1, OsLecRK2 and OsLecRK3, and the proteinsshared more than 50% identity.The results of yeast two-hybrid analysis showed that OsRRK1 interacted with OsLecRK, OsLecRK2 and OsLecRK3.【Conclusion】OsRRK1 participates in the process of resistance to the brownplanthoppersthroughinteracting with BPH15 and BPH3 candidate genes.

Key words: rice, OsRRK1, brown planthopper, yeast two-hybrid system

中图分类号:

Q786

马银花, 李萍芳, 董文静, 易松望, 杨芳, 杜波, 金晨钟. 水稻抗性蛋白OsRRK1抗褐飞虱机理分析[J]. 中国水稻科学, 2020, 34(6): 512-519.

Yinhua MA, Pingfang LI, Wenjing DONG, Songwang YI, Fang YANG, Bo DU, Chenzhong JIN. Mechanism Analysis of Rice Resistance Protein OsRRK1 Againstthe Brown Planthopper[J]. Chinese Journal OF Rice Science, 2020, 34(6): 512-519.

图/表 6

表1 用于构建载体的引物

Table 1 Primers used in vectors constructed.

引物 Primer	引物序列（5'-3'） Primer sequence（5'-3'）	限制性内切酶酶切位点 Restriction endonuclease site
AD-LecRKF	CGGAATTCATGGTTGCTCTGCTACTCTTT	EcoR I
AD-LecRKR	CGGAATTCTTAATGAGAAAACAGAGGATTACATTCC	EcoR I
AD-LecRK1F	TCCCCCGGGATGGTTGCTCTGCTACTCTTTCCC	Sma I
AD-LecRK1R	TCCCCCGGGCTATGGAAGTGAGCTGATGAAGG	Sma I
AD-LecRK2F	TCCCCCGGGTATGGCACCTATCCTGTTCTTGCCC	Sma I
AD-LecRK2R	TCCCCCGGGTCATGCGAGTGAACTGATATAGG	Sma I
AD-LecRK3F	TCCCCCGGGTATGGCTCATCTCCTGTTCTTGCCC	Sma I
AD-LecRK3R	TCCCCCGGGTCATGCAAGTGAACTGATATAGG	SmaI
BK-OsRRK1F	CGGAATTCATGAGGCCTCTGTACCTGCGCAGC	EcoR I
BK-OsRRK1R	CGGAATTCCTAATTGCTCAAAGATGATGAGCA	EcoR I
T7	TAATACGACTCACTATAGGGCGA
3′AD	AGATGGTGCACGATGCACAG
3′BD	TTTTCGTTTTCCTAAGAGTC

图1 OsLecRK与OsLecRK1-OsLecRK3蛋白结构域预测 S_TKc—丝氨酸/苏氨酸蛋白激酶催化结构域。

Fig.1. Prediction of OsLecRK and OsLecRK1-OsLecRK3 protein domains. S_TKc, Serine/Threonine protein kinasescatalytic domain.

图2 OsLecRK与OsLecRK1-OsLecRK3蛋白序列比对

Fig. 2. Alignment of OsLecRK and OsLecRK1-OsLecRK3.

图3 OsLecRK与OsLecRK1-OsLecRK3进化分析

Fig. 3. Phylogenetic tree analysis of OsLecRK and OsLecRK1-OsLecRK3.

图4 利用酵母双杂交对OsRRK1与OsLecRK蛋白互作的验证 pGBKT7-53/ pGADT7-T：阳性对照;pGBKT7-Lam/ pGADT7-T：阴性对照。DDO：SD/-Leu/-Trp（缺乏亮氨酸和色氨酸）双缺;TDO：SD/-Leu/-Trp/-His（缺乏亮氨酸,色氨酸和组氨酸）三缺。

Fig. 4. The interaction between OsRRK1 and OsLecRK protein was verified by yeast two-hybridization. pGBKT7-53/pGADT7-T, Positive control; pGBKT7-Lam/pGADT7-T, Negative control. DDO, SD/-Leu/-Trp; TDO, SD/-Leu/-Trp/-His.

图5 OsRRK1与OsLecRK1-OsLecRK3蛋白互作的验证 DDO：SD/-Leu/-Trp（缺乏亮氨酸和色氨酸）双缺;TDO：SD/-Leu/-Trp/-His（缺乏亮氨酸,色氨酸和组氨酸）三缺。

Fig. 5. Confirmation of positive interactions between OsRRK1 and OsLecRK1-OsLecRK3. DDO: SD/-Leu/-Trp; TDO: SD/-Leu/-Trp/-His.

参考文献 29

[1]	Khush G S. What it will take to feed 5.0 billion rice consumers in 2030[J]. Plant Molecular Biology, 2005, 59(1): 1-6.
[2]	潘磊, 王利华, 朱凤, 韩阳春, 王培, 方继朝. 褐飞虱小分子量热激蛋白基因表达特性和功能[J]. 中国水稻科学, 2020, 34(1): 37-45.
	Pan L,Wang L H,Zhu F, Han Y C, Wang P, Fang J C.Expression profiles and functions of small heat shock proteins in Nilaparvatalugens[J].Chinese Journal of Rice Science, 2020, 34(1): 37-45.(in Chinese with English abstract)
[3]	Heong K, Hardy B.Planthoppers: New threats to the sustainability of intensive rice production systems in Asia[M]. International Rice Research Institute: Los Banos(Philippines),2009: 191-200.
[4]	Qiu Y F, Guo J P, Jing S L, Zhu L, He G C.Development and characterization of japonica rice lines carrying the brown planthopper-resistance genes BPH12 and BPH6[J].Theoretical & Applied Genetics,2012,124(3):485-494.
[5]	刘万才, 刘振东, 黄冲, 陆明红, 刘杰, 杨清坡. 近10年农作物主要病虫害发生危害情况的统计和分析[J]. 植物保护, 2016, 42(5): 1-9, 46.
	Liu W C, Liu Z D, Huang C, Lu M H, Liu J, Yang Q P.Statistics and analysis of crop yield losses caused by main diseases and insect pests in recent 10 years[J].Plant Protection, 2016, 42(5): 1-9, 46.(in Chinese with English abstract)
[6]	杜波, 陈荣智, 何光存. 抗褐飞虱基因的发掘、鉴定与利用[J]. 生命科学, 2018, 30(10): 1072-1082.
	Du B, Chen R Z, He G C.Exploitation, identification and utilization of brown planthopper resistance genes in rice[J].Chinese Bulletin of Life Sciences, 2018, 30(10): 1072-1082.(in Chinese with English abstract)
[7]	Jing S, Zhao Y, Du B, Chen R Z, Zhu L L, He G C.Genomics of interaction between the brown planthopper and rice[J].Current Opinion in Insect Science,2017,19:82-87.
[8]	Du B, Zhang W L, Liu B F, Hu J, Wei Z A, He R F, Zhu L L, Chen R Z, Han B, He G C.Identification and characterization of Bph14, a gene conferring resistance to brown planthopper in rice[J].Proceedings of the National Academy of Sciences of the United States of America,2009,106(52):22163-22168.
[9]	Zhao Y, Huang J, Wang Z Z, Jing S L, Wang Y, Ouyang Y, Cai B, Xin X F, Liu X, Zhang C X, Pan Y F, Ma R, Li Q F, Jiang W H, Zeng Y, Shangguan X, Wang H Y, Du B, Zhu L L, Xu X, Feng Y Q, He S Y, Chen R Z, Zhang Q F, He G C.Allelic diversity in an NLR gene BPH9 enables rice to combat planthopper variation[J].Proceedings of the National Academy of Sciences of the United States of America,2016,113(45):12850-12855.
[10]	Guo J P, Xu C X, Wu D, Zhao Y, Qiu Y F, Wang X X, Ouyang Y, Cai B, Liu X, Jing S L, Shangguan X, Wang H Y, Ma Y H, Hu L, Wu Y, Shi S J, Wang W L, Zhu L L, Xu X, Chen R Z, Feng Y, Du B, He G.Bph6encodes an exocyst-localized protein and confers broad resistance to planthoppers in rice[J].Nature Genetics,2018,50(2):297-306.
[11]	Tamura Y, Hattori M, Yoshioka H, Yoshioka M, Takahashi A, Wu J, Sentoku N, Yasui H.Map-based cloning and characterization of a brown planthopper resistance geneBph26 from Oryza sativa L. ssp. indica cultivar ADR52[J/OL].Scientific Reports,2014,4:5872.
[12]	Wang Y, Cao L, Zhang Y, Cao C X, Liu F, Huang F, Qiu Y F, Li R B, Lou X J.Map-based cloning and characterization ofBph29, a B3 domain-containing recessive gene conferring brown planthopper resistance in rice[J/OL].Journal of Experimental Botany,2015,66(19):6035-6045.
[13]	Ji H, Kim S R, Kim Y H, Suh J P, Park H M, Sreenivasulu N, Misra G, Kim S M, Hechanova S L, Kim H, Lee G S, Yoon U H, Kim T H, Lim H, Suh S C, Yang J, An G, Jena K K.Map-based cloning and characterization of the BPH18gene from wild rice conferring resistance to brown planthopper(BPH) insect pest[J].Scientific Reports,2016,6:34376.
[14]	Ren J, Gao F Y, Wu X T, Lu X J, Zeng L, Lü J Q, Su X W, Luo H, Ren G J.Bph32, a novel gene encoding an unknown SCR domain-containing protein, confers resistance against the brown planthopper in rice[J/OL].Scientific Reports,2016,6:37645.
[15]	Liu Y Q, Wu H, Chen H, Liu Y L, He J A, Sun Z G, Pan G, Wang Q, Hu J L, Zhou F, Zhou K N, Zheng X M, Ren Y A, Wang Y H, Zhao Z G, Lin Q A, Zhang X, Guo X P, Cheng X, Jiang L, Wu C Y, Wang H Y, Wan J M.A gene cluster encoding lectin receptor kinases confers broad-spectrum and durable insect resistance in rice[J].Nature Biotechnology,2015,33(3):301.
[16]	Cheng X, Wu Y, Guo J, Du B, Chen R, Zhu L, He G.A rice lectin receptor-like kinase that is involved in innate immune responses also contributes to seed germination[J].Plant Journal,2013,76(4):687-698.
[17]	曹妮, 陈渊, 季芝娟, 曾宇翔, 杨长登, 梁燕. 水稻抗稻瘟病分子机制研究进展[J]. 中国水稻科学, 2019, 33(6): 489-498.
	Cao N, Chen Y, Ji Z J, Zeng Y X, Yang C D, Liang Y.Recent progress in molecular mechanism of rice blast resistance[J].Chinese Journal of Rice Science, 2019, 33(6): 489-498.(in Chinese with English abstract)
[18]	Vaid N, Pandey P K, Tuteja N.Genome-wide analysis of lectin receptor-like kinase family from Arabidopsis and rice[J].Plant Molecular Biology,2012,80(4-5):365-388.
[19]	Huang Z, He G C, Shu L H, Li XH, Zhang QF.Identification and mapping of two brown planthopper resistance genes in rice[J].Theoretical and Applied Genetics,2001,102(6):929-934.
[20]	Yang H, You A, Yang Z, Zhang F, He R, Zhu L, He G.High-resolution genetic mapping at the Bph15 locus for brown planthopper resistance in rice (Oryza sativa L.)[J].Theoretical and Applied Genetics,2004,110(1):182-191.
[21]	Ma Y, Zhao Y, Shangguan X, Shi S J, Zeng Y, Wu Y, Chen R Z, You A, Zhu L L, Du B, He G C.Overexpression of OsRRK1 changes leaf morphology and defense to insect in rice[J].Frontiers in Plant Science,2017,8:1783.
[22]	Shiu S H, Karlowski W M, Pan R S, Tzeng Y H, Mayer K A.Comparative analysis of the receptor-like kinase family in Arabidopsis and rice[J].Plant Cell,2004,16(5):1220-1234.
[23]	Yamaguchi K, Yamada K, Ishikawa K, Yoshimura S, Hayashi N, Uchihashi K, Ishihama N, Kishi-Kaboshi M, Takahashi A, Tsuge S, Ochiai H, Tada Y, Shimamoto K, Yoshioka H, Kawasaki T.A receptor-like cytoplasmic kinase targeted by a plant pathogen effector is directly phosphorylated by the chitin receptor and mediates rice immunity[J].Cell Host Microbe,2013,13(3):347-357.
[24]	Wang J, Wu G W, Peng C F, Zhou X G, Li W T, He M, Wang J C, Yin J C, Yuan C, Ma W W, Ma B T, Wang Y P, Chen W L, Chen W L, Qin P, Li S G, Chen X W.The Receptor-like cytoplasmic kinase OsRLCK102 regulates XA21-mediated immunity and plant development in rice[J].Plant Molecular Biology Reporter,2016,34(3):628-637.
[25]	Zhou X G, Wang J, Peng C F, Zhu X A, Li W T, He M, Wang J C, Chen M, Yuan C, Wu W, Ma W W, Qin P A, Wu X J, Li S G, Ronald P, Chen X E.Four receptor-like cytoplasmic kinases regulate development and immunity in rice[J].Plant Cell and Environment,2016,39(6):1381-1392.
[26]	Fields S, Song O.A novel genetic system to detect protein-protein interactions[J].Nature,1989,340(6230):245-246.
[27]	Jones J D, Dangl J L.The plant immune system[J].Nature,2006,444(7117):323-329.
[28]	Caplan J,Padmanabhan M,Dinesh-Kumar S P. Plant NB-LRR immune receptors: From recognition to transcriptional reprogramming[J]. Cell Host &Microbe, 2008,3(3): 126-135.
[29]	Spoel S H, Dong X.How do plants achieve immunity? Defense without specialized immune cells[J].Nature Reviews Immunology,2012,12(2):89-100.

水稻抗性蛋白OsRRK1抗褐飞虱机理分析

Mechanism Analysis of Rice Resistance Protein OsRRK1 Againstthe Brown Planthopper

RichHTML

PDF

可视化

摘要/Abstract

引用本文

使用本文

图/表 6

参考文献 29

相关文章 15

编辑推荐

Metrics

[1]	任志奇, 薛可欣, 董铮, 李小湘, 黎用朝, 郭玉静, 刘文强, 郭梁, 盛新年, 刘之熙, 潘孝武. 水稻外卷叶突变体ocl1的鉴定及基因定位[J]. 中国水稻科学, 2023, 37(4): 337-346.
[2]	肖乐铨, 李雷, 戴伟民, 强胜, 宋小玲. 转cry2A/bar*基因水稻与杂草稻杂交后代的苗期生长特性[J]. 中国水稻科学, 2023, 37(4): 347-358.
[3]	李刚, 高清松, 李伟, 张雯霞, 王健, 程保山, 王迪, 高浩, 徐卫军, 陈红旗, 纪剑辉. 定向敲除SD1基因提高水稻的抗倒性和稻瘟病抗性[J]. 中国水稻科学, 2023, 37(4): 359-367.
[4]	汪胜勇, 陈宇航, 陈会丽, 黄钰杰, 张啸天, 丁双成, 王宏伟. 水稻减数分裂期高温对苯丙烷类代谢及下游分支代谢途径的影响[J]. 中国水稻科学, 2023, 37(4): 368-378.
[5]	董立强, 杨铁鑫, 李睿, 商文奇, 马亮, 李跃东, 隋国民. 株行距配置对超高产田水稻产量及根系形态生理特性的影响[J]. 中国水稻科学, 2023, 37(4): 392-404.
[6]	韩聪, 何禹畅, 吴丽娟, 郏丽丽, 王磊, 鄂志国. 水稻碱性亮氨酸拉链(bZIP)蛋白家族功能研究进展[J]. 中国水稻科学, 2023, 37(4): 436-448.
[7]	沈雨民, 陈明亮, 熊焕金, 熊文涛, 吴小燕, 肖叶青. 水稻内外稃异常发育突变体blg1 (beak like grain 1)的表型分析与精细定位[J]. 中国水稻科学, 2023, 37(3): 225-232.
[8]	段敏, 谢留杰, 高秀莹, 唐海娟, 黄善军, 潘晓飚. 利用CRISPR/Cas9技术创制广亲和水稻温敏雄性不育系[J]. 中国水稻科学, 2023, 37(3): 233-243.
[9]	程玲, 黄福钢, 邱一埔, 王心怡, 舒宛, 邱永福, 李发活. 籼稻材料570011抗褐飞虱基因的遗传分析及鉴定[J]. 中国水稻科学, 2023, 37(3): 244-252.
[10]	王文婷, 马佳颖, 李光彦, 符卫蒙, 李沪波, 林洁, 陈婷婷, 奉保华, 陶龙兴, 符冠富, 秦叶波. 高温下不同施肥量对水稻产量品质形成的影响及其与能量代谢的关系分析[J]. 中国水稻科学, 2023, 37(3): 253-264.
[11]	刘嫒桦, 李小坤. 不同肥料施用与稻米品质关系的整合分析[J]. 中国水稻科学, 2023, 37(3): 276-284.
[12]	杨晓龙, 王彪, 汪本福, 张枝盛, 张作林, 杨蓝天, 程建平, 李阳. 不同水分管理方式对旱直播水稻产量和稻米品质的影响[J]. 中国水稻科学, 2023, 37(3): 285-294.
[13]	魏晓东, 宋雪梅, 赵凌, 赵庆勇, 陈涛, 路凯, 朱镇, 黄胜东, 王才林, 张亚东. 硅锌肥及其施用方式对南粳46产量和稻米品质的影响[J]. 中国水稻科学, 2023, 37(3): 295-306.
[14]	林聃, 江敏, 苗波, 郭萌, 石春林. 水稻高温热害模型研究及其在福建省的应用[J]. 中国水稻科学, 2023, 37(3): 307-320.
[15]	郑承梅, 孙金秋, 刘梦杰, 杨永杰, 陆永良, 郭怡卿, 唐伟. 水稻田糠稷种子萌发和出苗特性及化学防除药剂筛选[J]. 中国水稻科学, 2023, 37(3): 321-328.