研究报告

复苏植物旋蒴苣苔J结构域蛋白编码基因BhDNAJC2的 克隆、表达与功能

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  • 1西北农林科技大学生命科学学院, 杨凌 712100;
    2.宜宾学院生命科学与食品工程学院, 宜宾 644000;
    3中国科学院植物研究所北方资源植物重点实验室, 北京 100093;
    4.黑龙江八一农垦大学生命科学技术学院, 大庆 163319

收稿日期: 2014-03-12

  修回日期: 2014-05-08

  网络出版日期: 2015-04-10

基金资助

国家高技术研究发展计划(No.2007AA021403)、国家转基因生物新品种培育科技重大专项(No.2009ZX08009-060B)、国家自然科学基金委员会青年科学基金(No.31100195)和宜宾学院重点科研项目(No.2013QD07)

Cloning, Expression and Functional Analysis of a J-domain Protein- coding Gene, BhDNAJC2, from the Resurrection Plant Boea hygrometrica

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  • 1College of Life Sciences, Northwest Agriculture & Forestry University, Yangling 712100, China;
    2.College of Life Sciences and Food Engineering, Yibin University, Yibin 644000, China;
    3.Key Laboratory of Plant Resources, Institute of Botany, Chinese Academy of Sciences, Beijing 100093, China;
    4.College of Life Sciences and Technology, Heilongjiang Bayi Agricultural University, Daqing 163319, China

Received date: 2014-03-12

  Revised date: 2014-05-08

  Online published: 2015-04-10

摘要

热激蛋白(HSP)是一类在受到逆境刺激后大量表达的蛋白质, 能够帮助蛋白质正确折叠, 促使变性蛋白质降解, 缓解逆境胁迫对生物体的损伤。为揭示热激蛋白在耐旱的复苏植物中的保护作用, 该研究对复苏植物旋蒴苣苔(Boea hygrometrica)HSP40家族中J结构域蛋白BhDNAJC2的编码基因进行了克隆、表达与功能分析。Real-time PCR检测表明, 该基因受脱水、低温、热激等多种逆境条件和脱落酸(ABA)诱导表达。BhDNAJC2-YFP定位于细胞质、内质网和细胞核。过表达BhDNAJC2的拟南芥(Arabidopsis thaliana)株系在干旱、热激、盐胁迫和碱胁迫下均表现出明显的抗逆性。综上所述, BhDNAJC2可能在旋蒴苣苔抗旱、耐热及耐盐碱等胁迫反应中起关键作用。

本文引用格式

陈世璇, 张振南, 王波, 朱燕, 龚月桦, 孙冬梅, 邓馨 . 复苏植物旋蒴苣苔J结构域蛋白编码基因BhDNAJC2的 克隆、表达与功能[J]. 植物学报, 2015 , 50(2) : 180 -190 . DOI: 10.3724/SP.J.1259.2015.00180

Abstract

Heat shock proteins (Hsps) can relieve environmental stress damage by aiding proper folding or degradation of denatured proteins. The resurrection plant Boea hygrometrica can tolerate desiccation and revive after rehydration. In desiccated leaves of B. hygrometrica, an Hsp-coding gene was cloned and designated BhDNAJC2. Sequence analysis revealed that BhDNAJC2 contained a J-domain and thus was a novel member of the Hsp40 family. Real-time PCR revealed that BhDNAJC2 was induced by dehydration, cold, heat, oxidative, alkaline stresses and abscisic acid treatment. Confocal microscopy revealed the BhDNAJC2-YFP fusion protein in the endoplasmic reticulum, nucleus and cytoplasm. Compared to the wild type, Arabidopsis plants overexpressing BhDNAJC2 showed improved stress tolerance under drought, heat, salt and alkaline stresses, as indicated by better growth, higher Fv/Fm, and lower electrolytic leakage. BhDNAJC2 likely plays a key role in resistance to osmotic, saline, alkaline and heat stresses in B. hygrometrica.

参考文献

1 张兰军, 姬飞腾, 王丽丽, 亓岽东, 朱燕, 邓馨 (2012). 复苏植物旋蒴苣苔C2结构域小蛋白BhC2DP1参与植物对ABA的反应. 植物学报 47, 11-27.
2 Al-Whaibi MH (2011). Plant heat-shock proteins: a mini review. J King Saud Univ-Sci 23, 139-150.
3 Bekh-Ochir D, Shimada S, Yamagami A, Kanda S, Ogawa K, Nakazawa M, Matsui M, Sakuta M, Osada H, Asami T, Nakano T (2013). A novel mitochondrial DnaJ/Hsp40 family protein BIL2 promotes plant growth and resistance against environmental stress in brassinosteroid signaling. Planta 237, 1509-1525.
4 Chaves MM, Flexas J, Pinheiro C (2009). Photosynthesis under drought and salt stress: regulation mechanisms from whole plant to cell. Ann Bot 103, 551-560.
5 De Rybel B, van den Berg W, Lokerse AS, Liao CY, van Mourik H, Möller B, Llavata-Peris CI, Weijers D (2011). A versatile set of ligation-independent cloning vectors for functional studies in plants. Plant Physiol 156, 1292- 1299.
6 Deng X, Hu ZA, Wang HX, Wen XG, Kuang TY (2003). A comparison of photosynthetic apparatus of the detached leaves of the resurrection plant Boea hygrometrica with its non-tolerant relative Chirita heterotrichia in response to dehydration and rehydration. Plant Sci 165, 851-861.
7 Glynn JM, Froehlich JE, Osteryoungc KW (2008). Arabidopsis ARC6 coordinates the division machineries of the inner and outer chloroplast membranes through interaction with PDV2 in the intermembrane space. Plant Cell 20, 2460-2470.
8 Horne BE, Li TF, Genevaux P, Georgopoulos C, Landry SJ (2010). The Hsp40 J-domain stimulates Hsp70 when tethered by the client to the ATPase domain. J Biol Chem 285, 21679-21688.
9 Horton P, Park KJ, Obayashi T, Fujita N, Harada H, Adams-Collier CJ, Nakai K (2007). WoLF PSORT: protein localization predictor. Nucleic Acids Res 35, 585-587.
10 Ingle RA, Schmidt UG, Farrant JM, Thomson JA, Mundree SG (2007). Proteomic analysis of leaf proteins during dehydration of the resurrection plant Xerophyta viscosa . Plant Cell Environ 30, 435-446.
11 Katavic V, Haughn GW, Reed D, Martin M, Kunst L (1994). In planta transformation of Arabidopsis thaliana . Mol Gene Genet 245, 363-370.
12 Kelley WL (1998). The J-domain family and the recruitment of chaperone power. Trends Biochem Sci 23, 222-227.
13 Kong FY, Deng YS, Zhou B, Wang GD, Wang Y, Meng QW (2014). A chloroplast-targeted DnaJ protein contri- butes to maintenance of photosystem II under chilling stress. J Exp Bot 65, 143-158.
14 Kregel KC (2002). Heat shock proteins: modifying factors in physiological stress responses and acquired thermotolerance. J Appl Physiol 92, 2177-2186.
15 Kroczynska B, Blond SY (2001). Cloning and characterization of a new soluble murine J-domain protein that stimulates BiP, Hsc70 and DnaK ATPase activity with different efficiencies. Gene 273, 267-274.
16 Li JW, Zhang H, Hu JB, Liu JQ, Liu KK (2012). A heat shock protein gene, CsHsp45.9 , involved in the response to diverse stresses in cucumber. Biochem Genet 50, 565-578.
17 Liu X, Wang Z, Wang LL, Wu RH, Phillipsb J, Deng X (2009). LEA 4 group genes from the resurrection plant Boea hygrometrica confer dehydration tolerance in trans- genic tobacco. Plant Sci 176, 90-98.
18 Livak KJ, Schmittgen TD (2001). Analysis of relative gene expression data using real-time quantitative PCR and the 2 -ΔΔCT method. Methods 25, 402-408.
19 Miernyk JA (2001). The J-domain proteins of Arabidopsis thaliana : an unexpectedly large and diverse family of chaperones. Cell Stress Chaperon 6, 209-218.
20 Mitra J, Xu GH, Wang B, Li MJ, Deng X (2013). Understanding desiccation tolerance using the resurrection plant Boea hygrometrica as a model system. Front Plant Sci 4, 446.
21 Rajan VBV, D’Silva P (2009). Arabidopsis thaliana J-class heat shock proteins: cellular stress sensors. Funct Integr Genomics 9, 433-446.
22 Rowley N, Carina PB, Westermann B, Brown C, Schwarz E, Barrell B, Neupert W (1994). Mdj1p, a novel chaperone of the DnaJ family, is involved in mitochondrial biogenesis and protein folding. Cell 77, 249-259.
23 Shen LS, Kang YG, Lu L, Yu H (2011). The J-domain protein J3 mediates the integration of flowering signals in Arabidopsis. Plant Cell 23, 499-514.
24 Shinozaki K, Yamaguchi-Shinozaki K (2007). Gene networks involved in drought stress response and tolerance. J Exp Bot 58, 221-227.
25 So HA, Chung E, Lee JH (2013). Molecular characterization of soybean GmDjp1 encoding a type III J-protein induced by abiotic stress. Gene Genom 35, 247-256.
26 Sparkes IA, Runions J, Kearns A, Hawes C (2006). Rapid, transient expression of fluorescent fusion proteins in tobacco plants and generation of stably transformed plants. Nat Protoc 1, 2019-2025.
27 Thompson JD, Gibson TJ, Plewniak F, Jeanmougin F, Higgins DG (1997). The CLUSTAL_X windows interface: flexible strategies for multiple sequence alignment aided by quality analysis tools. Nucleic Acids Res 25, 4876-4882.
28 Verslues PE, Bray EA (2004). LWR1 and LWR2 are required for osmoregulation and osmotic adjustment in Arabidopsis. Plant Physiol 136, 2831-2842.
29 Wang XX, Jia N, Zhao CL, Fang YL, Lv TT, Zhou W, Sun YZ, Li B (2014). Knockout of AtDjB1, a J-domain protein from Arabidopsis thaliana , alters plant responses to osmotic stress and abscisic acid. Physiol Plant 152, 286- 300.
30 Wang Z, Zhu Y, Wang LL, Liu X, Liu YX, Phillipsb J, Deng X (2009). A WRKY transcription factor participates in dehydration tolerance in Boea hygrometrica by binding to the W-box elements of the galactinol synthase ( BhGolS1 ) promoter. Planta 230, 1155-1166.
31 Yang CW, Xu HH, Wang LL, Liu J, Shi DC, Wang DL (2009). Comparative effects of salt-stress and alkali- stress on the growth, photosynthesis, solute accumulation, and ion balance of barley plants. Photosynthetica 47, 79-86.
32 Zhang ZN, Wang B, Sun DM, Deng X (2013). Molecular cloning and differential expression of sHSP gene family members from the resurrection plant Boea hygrometrica in response to abiotic stresses. Biologia 68, 651-661.
33 Zhao ZC, Zhang WR, Yan JP, Zhang JJ, Liu Z, Li XF, Yi Y (2010). Over-expression of Arabidopsis DnaJ (Hsp40) contributes to NaCl-stress tolerance. Afr J Biotechnol 9, 972-978.
34 Zhou W, Zhou T, Li MX, Zhao CL, Jia N, Wang XX, Sun YZ, Li GL, Xu M, Zhou RG, Li B (2012). The Arabidopsis J-protein AtDjB1 facilitates thermotolerance by protecting cells against heat-induced oxidative damage. New Phytologist 194, 364-378.
35 Zhu Y, Wang Z, Jing YJ, Wang LL, Liu X, Liu YX, Deng X (2009). Ectopic over-expression of BhHsf1 , a heat shock factor from the resurrection plant Boea hygrometrica , leads to increased thermotolerance and retarded growth in transgenic Arabidopsis and tobacco. Plant Mol Biol 71, 451-467.
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