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作物学报 ›› 2020, Vol. 46 ›› Issue (8): 1217-1224.doi: 10.3724/SP.J.1006.2020.92060

• 作物遗传育种·种质资源·分子遗传学 • 上一篇    下一篇

OsRPK1基因过表达和RNA干涉对水稻苗期耐盐性的影响

李晶岚,陈鑫欣,石翠翠,刘方惠,孙静,葛荣朝()   

  1. 河北师范大学生命科学学院, 河北石家庄 050024
  • 收稿日期:2019-11-25 接受日期:2020-03-24 出版日期:2020-08-12 网络出版日期:2020-04-21
  • 通讯作者: 葛荣朝
  • 作者简介:李晶岚, E-mail: 83494429@qq.com|陈鑫欣, E-mail: 1046433763@qq.com
  • 基金资助:
    国家自然科学基金项目(30900104);北省自然科学基金项目(C2016205158)

Effects of OsRPK1 gene overexpression and RNAi on the salt-tolerance at seedling stage in rice

LI Jing-Lan,CHEN Xin-Xin,SHI Cui-Cui,LIU Fang-Hui,SUN Jing,GE Rong-Chao()   

  1. College of Life Science, Hebei Normal University, Shijiazhuang 050024, Hebei, China
  • Received:2019-11-25 Accepted:2020-03-24 Published:2020-08-12 Published online:2020-04-21
  • Contact: Rong-Chao GE
  • Supported by:
    National Natural Science Foundation of China(30900104);Hebei Provincial Natural Science Foundation(C2016205158)

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摘要：

水稻OsRPK1基因属于富含亮氨酸重复序列的类受体蛋白激酶, 在盐胁迫下其表达量暂时升高后出现明显下降。前期研究表明, OsRPK1基因过表达会造成拟南芥非生物胁迫耐受性明显降低。本研究对OsRPK1基因在水稻中实现过表达和RNA干涉, 进而对OsRPK1的抗逆性功能加以探究。结果表明, OsRPK1基因表达量增加时, 水稻幼苗表现为耐盐性下降; RNA干涉则使水稻幼苗表现为耐盐性增加。盐胁迫后OsRPK1过表达植株脯氨酸含量低于野生型、MDA含量和相对电导率显著高于野生型, 而OsRPK1-RNAi水稻植株的脯氨酸含量显著高于野生型、MDA含量和相对电导率显著低于野生型。因此, OsRPK1基因的表达量对水稻耐盐性具有明显影响, 脯氨酸含量及细胞质膜受破损程度的改变可能是造成转基因水稻耐盐性变化的内在原因。本研究为进一步阐明OsRPK1基因的抗逆作用机制奠定了基础, 对于通过调整该基因表达改良水稻的耐盐性具有重要意义。

关键词: 水稻, OsRPK1基因, 过表达, RNA干涉, 生理指标

Abstract:

OsRPK1 is a leucine-rich repeat receptor-like protein kinase (LRR-RLK) gene in rice, and its expression level decreases evidently after temporarily increasing under salt stress. Previous studies have shown that the overexpression of OsRPK1 can significantly reduce the abiotic stress tolerance of Arabidopsis. In this study, OsRPK1 gene was overexpressed and RNA interfered in rice. The salt tolerance of the 35S:OsRPK1 rice seedlings was significantly lower than that of the control plants. The OsRPK1-RNAi rice seedlings exhibited higher salt tolerance than the wild-type plants. The proline content of OsRPK1 overexpressed plants was lower than that of the wild type, MDA content and relative conductivity were significantly higher than those of the wild type, while the proline content of OsRPK1-RNAi rice plants was significantly higher than those of wild type, MDA content and relative conductivity were significantly lower than those of wild type. The results of this study indicate that OsRPK1 plays an important role in salt tolerance of rice. The changes of proline content and the damage degree of cytoplasmic membrane may be the internal reasons for the salt-tolerance variations of the transgenic rice lines. This study lays a foundation for further elucidating the resistance mechanism of OsRPK1 gene, which is very important for cultivating salt-tolerance rice by adjusting the expression of OsRPK1 gene.

Key words: rice, OsRPK1 genes, overexpressed, RNA interference, physiological indexes

图1

OsRPK1基因在140 mmol L-1 NaCl胁迫的水稻中表达量的变化 差异显著性分析采用t-test方法, *P < 0.05、**P < 0.01、***P < 0.001。"

图2

35S:OsRPK1植株与OsRPK1-RNAi植株的qRT-PCR检测 差异显著性分析采用t-test方法, *P < 0.05、**P < 0.01、 ***P < 0.001。"

图3

OsRPK1过表达水稻在盐胁迫下的生长情况 A: NaCl胁迫前; B: 140 mmol L-1 NaCl胁迫6 d; C: 140 mmol L-1 NaCl胁迫后恢复9 d; D: 盐胁迫恢复培养后存活率统计。差异显著性分析采用t-test方法, *P < 0.05、**P < 0.01、***P < 0.001。"

图4

OsRPK1-RNAi水稻在胁迫下生长情况 A: NaCl胁迫前; B: 140 mmol L-1 NaCl胁迫10 d; C: 140 mmol L-1 NaCl胁迫后恢复7 d; D, E: 盐胁迫恢复培养后存活率统计。差异显著性分析采用t-test方法, *P < 0.05、**P < 0.01、***P < 0.001。"

图5

140 mmol L-1 NaCl盐胁迫后转基因水稻脯氨酸含量 差异显著性分析采用t-test方法, *P < 0.05、**P < 0.01、***P < 0.001。"

图6

140 mmol L-1 NaCl盐胁迫后转基因水稻株系中MDA含量 差异显著性分析采用t-test方法, *P < 0.05、**P < 0.01、***P<0.001。"

图7

转基因水稻的相对电导率 差异显著性分析采用t-test方法, *P < 0.05、**P < 0.01、***P < 0.001。"

[1] Liang W J, Ma X L, Wan P, Liu L Y. Plant salt-tolerance mechanism: a review. Biochem Biophy Res Commun, 2017,495:286-291.
[2] 李彬, 王志春, 孙志高, 陈渊, 杨福. 中国盐碱地资源与可持续利用研究. 干旱地区农业研究, 2005,23(3):154-158.
Li B, Wang Z C, Sun Z G, Chen Y, Yang F. Resources and sustainable resource exploitation of salinized land in China. Agric Res Arid Areas, 2005,23(3):154-158 (in Chinese with English abstract).
[3] 张建峰, 张旭东, 周金星, 刘国华, 李冬雪. 世界盐碱地资源及其改良利用的基本措施. 水土保持研究, 2005,12(6):28-30.
Zhang J F, Zhang X D, Zhou J X, Liu G H, Li D X. World resources of saline soil and main amelioratior measures. Res Soil Water Conserv, 2005,12(6):28-30 (in Chinese with English abstract).
[4] Lee S K, Kim B G, Kwon T R, Jeong M J, Park S R, Lee J W, Byun M O, Kwon H B, Matthews B F, Hong C B, Park S C. Overexpression of the mitogen-activated protein kinase gene OsMAPK33 enhances sensitivity to salt stress in rice(Oryza sativa L.). J Biosci, 2011,36:139-151.
pmid: 21451255
[5] Xing H T, Guo P, Xia X L, Yin W L. PdERECTA, a leucine-rich repeat receptor-like kinase of poplar, confers enhanced water use efficiency in Arabidopsis. Planta, 2011,234:229-241.
pmid: 21399949
[6] Mondal T K, Rawal H C, Chowrasia S, Varshney D, Panda A K, Mazumdar A, Kaur H, Gaikwad K, Sharma T R, Singh N K. Draft genome sequence of first monocot-halophytic species Oryza coarctata reveals stress-specific genes. Sci Rep, 2018,8:13698-13708.
pmid: 30209320
[7] 牛吉山. 植物和小麦蛋白激酶的研究现状. 西北植物学报, 2003,23:143-150.
Niu J S. Studies on plant and wheat protein kinases. Acta Bot Boreali-Occident Sin, 2003,23:143-150 (in Chinese with English abstract).
[8] Julie M S, John C W. Plant protein kinase families and signal transduction. Plant Physiol, 1995,108:451-457.
[9] Depuydt S, Rodriguez-Villalon A, Santuari L, Wyser-Rmili C, Ragni L, Hardtke C S. Suppression of Arabidopsis protophloem differentiation and root meristem growth by CLE45 requires the receptor-like kinase BAM3. Proc Natl Acad Sci USA, 2013,110:7074-7079.
doi: 10.1073/pnas.1222314110 pmid: 23569225
[10] Bryan A C, Obaidi A, Wierzba M, Tax F E. XYLEM INTERMIXED WITH PHLOEM1, a leucine-rich repeat receptor-like kinase required for stem growth and vascular development in Arabidopsis thaliana. Planta, 2012,235:111-122.
pmid: 21853254
[11] Burr C A, Leslie M E, Orlowski S K, Chen I, Wright C E, Daniels M J, Liljegren S J. CAST AWAY, a membrane-associated receptor-like kinase, inhibits organ abscission in Arabidopsis. Plant Physiol, 2011,156:1837-1850.
doi: 10.1104/pp.111.175224 pmid: 21628627
[12] Mizuno S, Osakabe Y, Maruyama K, Ito T, Osakabe K, Sato T, Shinozaki K, Yamaguchi-Shinozaki K. Receptor-like protein kinase 2 (RPK2) is a novel factor controlling anther development in Arabidopsis thaliana. Plant J, 2007,50:751-766.
pmid: 17419837
[13] Engelsdorf T, Hamann T. An update on receptor-like kinase involvement in the maintenance of plant cell wall integrity. Ann Bot, 2014,114:1339-1347.
doi: 10.1093/aob/mcu043 pmid: 24723447
[14] Wu Y, Zhou J M. Receptor-like kinases in plant innate immunity. J Integr Plant Biol, 2013,55:1271-1286.
doi: 10.1111/jipb.12123 pmid: 24308571
[15] Phillips S M, Dubery I A, Heerden H. Identification and molecular characterisation of a lectin receptor-like kinase (GHLECRK-2) from cotton. Plant Mol Biol Rep, 2013,31:9-20.
[16] Singh P, Kuo Y C, Mishra S, Tsai C H, Chien C C, Chen C W, Desclos-Theveniau M, Chu P W, Schulze B, Chinchilla D, Boller T, Zimmerli L. The lectin receptor kinase-VI.2 is required for priming and positively regulates Arabidopsis pattern-triggered immunity. Plant Cell, 2012,24:1256-1270.
pmid: 22427336
[17] Tanaka H, Osakabe Y, Katsura S, Mizuno S, Maruyama K, Kusakabe K, Mizoi J, Shinozaki K, Yamaguchi-Shinozaki K. Abiotic stress-inducible receptor-like kinases negatively control ABA signaling in Arabidopsis. Plant J: Cell Mol Biol, 2012,70:599-613.
[18] Hua D, Wang C, He J, Liao H, Duan Y, Zhu Z, Guo Y, Chen Z, Gong Z. A plasma membrane receptor kinase, GHR1, mediates abscisic acid- and hydrogen peroxide-regulated stomatal movement in Arabidopsis. Plant Cell, 2012,24:2546-2561.
doi: 10.1105/tpc.112.100107 pmid: 22730405
[19] Yuriko O, Shinji M, Hidenori T, Maruyama K, Osakabe K, Todaka D, Fujita Y, Kobayashi M, Shinozaki K, Yamaguchi- Shinozaki K. Overproduction of the membrane-bound receptor- like protein kinase1, RPK1, enhances abiotic stress tolerance in Arabidopsis. J Biol Chem, 2010,285:9190-9201.
doi: 10.1074/jbc.M109.051938 pmid: 20089852
[20] Yang T, Chaudhuri S, Yang L, Du L, Poovaiah B W. A calcium/calmodulin-regulated member of the receptor-like kinase family confers cold tolerance in plants. J Biol Chem, 2010,285:7119-7126.
doi: 10.1074/jbc.M109.035659 pmid: 20026608
[21] Sicilia A, Testa G, Santoro D F, Cosentino S L, Lo Piero A R. RNASeq analysis of giant cane reveals the leaf transcriptome dynamics under long-term salt stress. BMC Plant Biol, 2019,19:355-379.
doi: 10.1186/s12870-019-1964-y pmid: 31416418
[22] Song W Y, Wang G L, Chen L L, Kim H S, Pi L Y, Holsten T, Gardner J, Wang B, Zhai W X, Zhu L H, Fauquet C, Ronald P. A receptor kinase-like protein encoded by the rice disease resistance gene Xa21. Science, 1995,270:1804-1806.
doi: 10.1126/science.270.5243.1804 pmid: 8525370
[23] Kobe B, Deisenhofer J. The leucine-rich repeat: a versatile binding motif. Trends Biochem Sci, 1994,19:415-421.
doi: 10.1016/0968-0004(94)90090-6 pmid: 7817399
[24] Jones D A, Jones J D G. The role of leucine-rich repeat proteins in plant defences. Adv Bot Res, 1997,24:89-167.
[25] Hong S W, Jon J H, Kwak J M, Nam H G. Identification of a receptor-like protein kinase gene rapidly induced by abscisic acid, dehydration, high salt, and cold treatments in Arabidopsis thaliana. Plant Physiol, 1997,113:1203-1212.
doi: 10.1104/pp.113.4.1203 pmid: 9112773
[26] Osakabe Y, Maruyama K, Seki M, Satou M, Shinozaki K, Yamaguchi-Shinozaki K. Leucine-rich repeat receptor-like kinase1 is a key membrane-bound regulator of abscisic acid early signaling in Arabidopsis. Plant Cell, 2005,17:1105-1119.
doi: 10.1105/tpc.104.027474 pmid: 15772289
[27] Shi C C, Feng C C, Yang M M, Li J L, Li X X, Zhao B C, Huang Z J, Ge R C. Overexpression of the receptor-like protein kinase genes AtRPK1 and OsRPK1 reduces the salt tolerance of Arabidopsis thaliana. Plant Sci, 2014, 217-218:63-70.
doi: 10.1016/j.plantsci.2013.12.002 pmid: 24467897
[28] Wang J, Li C, Yao X, Liu S H, Zhang P Y, Chen K S. The Antarctic moss leucine-rich repeat receptor-like kinase (PnLRR-RLK2) functions in salinity and drought stress adaptation. Polar Biol, 2018,41:353-364.
[29] Sun X, Sun M, Luo X, Ding X, Cai H, Bai X, Liu X, Zhu Y. A Glycine soja, ABA-responsive receptor-like cytoplasmic kinase, GsRLCK, positively controls plant tolerance to salt and drought stresses. Planta, 2013,237:1527-1545.
doi: 10.1007/s00425-013-1864-6 pmid: 23494614
[30] An S H, Choi H W, Hwang I S, Hong J K, Hwang B K. A novel pepper membrane-located receptor-like protein gene CaMRP1 is required for disease susceptibility, methyl jasmonate insensitivity and salt tolerance. Plant Mol Biol, 2008,67:519-533.
doi: 10.1007/s11103-008-9337-1 pmid: 18427932
[31] Li C H, Wang G, Zhao J L, Zhang L Q, Ai L F, Han Y F, Sun D Y, Zhang S W, Sun Y. The receptor-like kinase SIT1 mediates salt sensitivity by activating MAPK3/6 and regulating ethylene homeostasis in rice. Plant Cell, 2014,26:2538-2553.
doi: 10.1105/tpc.114.125187
[32] Liu S H, Wang J, Chen K S, Zhang Z H, Zhang P Y. The L-type lectin receptor-like kinase(PnLecRLK1) from the Antarctic moss Pohlia nutans enhances chilling-stress tolerance and abscisic acid sensitivity in Arabidopsis. Plant Growth Regul, 81:409-418.
doi: 10.1007/s10725-016-0217-4
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