Abstract
WRKY proteins are a large family of transcriptional regulators involved in a variety of biological processes in plants. Here we report functional characterization of a rice WRKY gene, OsWRKY89. RNA gel blot analysis indicated that OsWRKY89 was strongly induced by treatments of methyl jasmonate and UV-B radiation. The transient expression analysis of the OsWRKY89–eGFP reporter in onion epidermal cells revealed that OsWRKY89 was targeted to nuclei. Transcriptional activity assays of OsWRKY89 and its mutants fused with a GAL4 DNA binding domain indicated that the 67 C-terminal amino acids were required for the transcriptional activation and that the leucine zipper region at the N-terminus enhanced its transcriptional activity. Overexpression of OsWRKY89 led to growth retardation at the early stage and reduction of internode length. Scanning electron microscopy revealed an increase in wax deposition on leaf surfaces of the OsWRKY89 overexpression lines and a decrease in wax loading in the RNAi-mediated OsWRKY89 suppression lines. Moreover, extractable and cell-wall-bound phenolic compounds were decreased in the overexpressor lines, but its SA levels were increased. Lignin staining showed an increase in lignification in culms of the overexpressor lines. Interestingly, overexpression of the OsWRKY89 gene enhanced resistance to the rice blast fungus and white-backed planthopper as well as tolerance to UV-B irradiation. These results suggest that OsWRKY89 plays an important role in response to biotic and abiotic stresses.
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References
Bieza K, Lois R (2001) An Arabidopsis mutant tolerant to lethal ultraviolet B level shows constitutively elevated accumulation of flavonoids and phenolics. Plant Physiol 126:1105–1115
Bohnert HJ, Nelson DE, Jensen RG (1995) Adaptations to environmental stress. Plant Cell 7:1099–1111
Brown BA, Cloix C, Jiang GH, Kaiserli E, Herzyk P, Kliebenstein DJ, Jenkins GI (2005) A UV-B-specific signaling component orchestrates plant UV protection. Proc Natl Acad Sci USA 102:18225–18230
Casati P, Walbot V (2003) Gene expression profiling in response to ultraviolet radiation in maize genotypes with varying flavonoid content. Plant Physiol 132:1739–1754
Chen C, Chen Z (2000) Isolation and characterization of two pathogen- and salicylic acid-induced genes encoding WRKY DNA-binding proteins from tobacco. Plant Mol Biol 42:387–396
Chen X, Goodwin SM, Boroff VL, Liu X, Jenks MA (2003) Cloning and characterization of the WAX2 gene of Arabidopsis involved in cuticle membrane and wax production. Plant Cell 15:1170–1185
Chomczyski P, Sacchi N (1987) Single-step method of RNA isolation by acid quanidinium thiocyanate-phenol-chloroform extraction. Annu Biochem 162:156–159
Cormack RS, Eulgem T, Rushton PJ, Kochner P, Hahlbrock K, Somssich IE (2002) Leucine zipper-containing WRKY proteins widen the spectrum of immediate early elicitor-induced WRKY transcription factors in parsley. Biochim Biophys Acta 1576:92–100
Day TA, Howells BW, Rice WJ (1994) Ultraviolet absorption and epidermal- transmittance spectra in foliage. Physiol Plant 92:207–218
Deslandes L, Oliver J, Peeters N, Feng DX, Khounlotham M, Boucher C, Somssich I, Genin S, Marco Y (2003) Physical interaction between RRS1-R, a protein conferring resistance to bacterial wilt, and PopP2, a type III effector targeted to the plant nucleus. Proc Natl Acad Sci USA 100:8024–8029
Dixon RA, Paiva NL (1995) Stress-induced phenylpropanoid metabolism. Plant Cell 7:1085–1097
Dong J, Chen C, Chen Z (2003) Expression profiles of the Arabidopsis WRKY gene superfamily during plant defense response. Plant Mol Biol 51:21–37
Eulgem T, Rushton PJ, Robatzek S, Somssich IE (2000) The WRKY superfamily of plant transcription factors. Trends Plant Sci 5:199–206
Fujimoto SY, Ohta M, Usui A, Shinshi H, Ohme-Takagi M (2000) Arabidopsis ethylene-responsive element binding factors act as transcriptional activators or repressors of GCC box-mediated gene expression. Plant Cell 12:393–404
Genty B, Briantais MJ, Baker NR (1989) The relationship between the quantum yield of photosynthetic electron transport and quenching of chlorophyll fluorescence. Biochim Biophys Acta 990:87–92
Green R, Fluhr R (1995) UV-B-induced PR-1 accumulation is mediated by active oxygen species. Plant Cell 7:203–212
Guo ZJ, Kan YC, Chen XJ, Li DB, Wang DW (2004a) Characterization a rice WRKY gene whose expression is induced upon pathogen attack and mechanical wounding. Acta Bot Sin 46:955–964
Guo ZJ, Chen XJ, Wu XL, Ling JQ, Xu P (2004b) Overexpression of the AP2/EREBP transcription factor OPBP1 enhances disease resistance and salt tolerance in tobacco. Plant Mol Biol 55:607–618
Hammond-Kosack KE, Jones JDG (1996) Resistance gene-dependent plant defense responses. Plant Cell 8:1773–1791
Hiei Y, Ohta S, Komari T, Kumashiro T (1994) Efficient transformation of rice (Oryza sativa L.) mediated by Agrobacterium and sequence analysis of the boundaries of the T-DNA. Plant J 6:271–282
Jansen MAK, Gaba V, Greenberg BM (1998) Higher plants and UV-B radiation: balancing damage, repair and acclimation. Trends Plant Sci 3:131–135
Jefferson RA (1987) Assaying chimeric genes implants: the Gus gene fusion system. Plant Mol Biol Rep 5:387–405
Jin H, Cominelli E, Bailey P, Parr A, Mehrtens F, Jones J, Tonelli C, Weisshaar B, Martin C (2000) Transcriptional repression by AtMYB4 controls production of UV-protecting sunscreens in Arabidopsis. EMBO J 19:6150–6161
Johnson CS, Kolevski B, Smyth DR (2002) TRANSPARENT TESTA GLABRA2, a trichome and seed coat development gene of Arabidopsis, encodes a WRKY transcription factor. Plant Cell 14:1359–1375
Landry LG, Chapple CCS, Last RL (1995) Arabidopsis mutants lacking phenolic sunscreen exhibit enhanced ultraviolet-B injury and oxidative damage. Plant Physiol 109:1159–1166
Li J, Ou-Lee T, Raba R, Amoundson R, Last R (1993) Arabidopsis flavonoid mutants are hypersensitive to UV-B irradiation. Plant Cell 5:171–179
Liu XQ, Bai XQ, Qian Q, Wang XJ, Chen MS, Chu CC (2005) OsWRKY03, a rice transcriptional activator that functions in defense signaling pathway upstream of OsNPR1. Cell Res 15:593–603
Logemann E, Tavernaro A, Schulz W, Somssich IE, Hahlbrock K (2000) UV light selectively coinduces supply pathways from primary metabolism and flavonoid secondary product formation in parsley. Proc Natl Acad Sci USA 97:1903–1907
Lolle SJ, Berlyn GP, Engstrom EM, Krolikowski KA, Reiter WD, Pruitt RE (1997) Developmental regulation of cell interactions in the Arabidopsis fiddlehead-1 mutant: a role for the epidermal cell wall and cuticle. Dev Biol 189:311–321
Luo M, Dennis ES, Berger F, Peacock WJ, Chaudhury A (2005) MINISEED3 (MINI3), a WRKY family gene, and HAIKU2 (IKU2), a leucine-rich repeat (LRR) KINASE gene, are regulators of seed size in Arabidopsis. Proc Natl Acad Sci USA 102:17531–17536
Mackerness S, John CF, Jordan BR, Thomas B (2001) Early signaling components in ultraviolet-B responses: distinct roles for different reactive oxygen species and nitric oxide. FEBS Lett 489:237–242
Mackerness S, Surplus SL, Blake P, John CF, Buchana-Wollaston V, Jordan BR, Thomas B (1999) Ultraviolet-B-induced stress and changes in gene expression in Arabidopsis thaliana: role of signaling pathways controlled by jasmonic acid, ethylene and reactive oxygen species. Plant Cell Environ 22:1413–1423
Maleck K, Levine A, Eulgem T, Morgan A, Schmid J, Lawton KA, Dangl JL, Dietrich RA (2000) The transcriptome of Arabidopsis thaliana during systemic acquired resistance. Nature Genet 26:403–410
Mulroy TW (1979) Spectral properties of heavily glaucous and non-glaucous leaves of a succulent rosette-plant. Oecologia 38:349–357
Müsel G, Schindler T, Bergfeld R, Ruel K, Jacquet G, Lapierre C, Speth V, Schopfer P (1997) Structure and distribution of lignin in primary and secondary cell walls of maize coleoptiles analyzed by chemical and immunological probes. Planta 201:146–159
Nakai K, Kanehisa M (1992) A knowledge base for predicting protein localization sites in eukaryotic cells. Genomics 14:897–911
Paul ND, Gwynn-Jones D (2003) Ecological roles of solar UV radiation: towards an integrated approach. Trends Ecol Evol 18:48–55
Peng YL, Shishiyama J (1988) Temporal sequence of cytological events in rice leaves affected with Pyriculria oryzae. Can J Bot 66:730–735
Rakwala R, Agrawala GK, Jwa NS (2001) Characterization of a rice (Oryza sativa L.) Bowman-Birk proteinase inhibitor: tightly light regulated induction in response to cut, jasmonic acid, ethylene and protein phosphatase 2A inhibitors. Gene 263:189–198
Reddy VS, Goud KV, Sharma R, Reddy AR (1994) Ultraviolet-B-responsive anthocyanin production in a rice cultivar is associated with a specific phase of phenylalanine ammonia-lyase biosynthesis. Plant Physiol 105:1059–1066
Robatzek S, Somssich IE (2002) Targets of AtWRKY6 regulation during plant senescence and pathogen defense. Genes Dev 16:1139–1149
Rozema J, van de Staaij J, Björn LO, Caldwell M (1997) UV-B as an environmental factor in plant life: stress and regulation. Trends Ecol Evol 12:22–28
Ryu H-S, Han M, Lee S-K, Cho J-I, Ryoo N, Heu S, Lee Y-H, Bhoo SH, Wang G-L, Hahn T-R, Jeon J-S (2006) A comprehensive expression analysis of the WRKY gene superfamily in rice plants during defense response. Plant Cell Rep 25:836–847
Sakamoto A, Lan VTT, Hase Y, Shikazono N, Matsunaga T, Tanaka A (2003) Disruption of the AtREV3 gene causes hypersensitivity to ultraviolet B light and γ-rays in Arabidopsis: implication of the presence of a translesion synthesis mechanism in plants. Plant Cell 15:2042–2057
Scheideler M, Schlaich ML, Fellenberg K, Beissbarth T, Hauser NC, Vingron M, Slusarenko AJ, Hoheisel JD (2002) Monitoring the switch from housekeeping to pathogen defense metabolism in Arabidopsis thaliana using cDNA arrays. J Biol Chem 277:10555–10561
Stratmann J (2003) Ultraviolet-B radiation co-opts defense signaling pathways. Trends Plant Sci 8:526–533
Suesslin C, Frohnmeyer H (2003) An Arabidopsis mutant defective in UV-B light-mediated responses. Plant J 33:591–601
Surplus SL, Jordan BR, Murphy AM, Carr JP, Thomas B, AH Mackerness S (1998) Ultraviolet-B-induced responses in Arabidopsis thaliana: role of salicylic acid and reactive oxygen species in the regulation of transcripts encoding photosynthetic and acidic pathogenesis-related proteins. Plant Cell Environ 21:685–694
Teramura AH, Sullivan JH (1994) Effects of UV-B radiation on photosynthesis and growth of terrestral plants. Photosynth Res 39:463–473
Turck F, Zhou A, Somssich IE (2004) Stimulus-dependent, promoter-specific binding of transcription factor WRKY1 to its native promoter and the defense-related gene PcPR1–1 in Parsley. Plant Cell 16:2573–2585
Wang D Amornsiripanitch N, Dong X (2006) A genomic approach to identify regulatory nodes in the transcriptional network of systemic acquired resistance in plants. PloS Pathog 2:1042–1050
Wang HH, Hao ZN, Xie K, Wu KL, Guo ZJ (2005) Leucine zipper like structure in rice WRKY89 enhances its affinity for binding with W box elements. Chinese Sci Bull 50:980–989
Wesley SV, Helliwell CA, Smith NA, Wang M, Rouse DT, Liu O, Gooding PS, Singh SP, Abbott D, Stoutjesdijk PA, Robinson SP, Gleave AP, Green AG, Waterhouse PM (2001) Construct design for efficient, effective and high through put gene silencing in plants. Plant J 27:581–590
Winkrel-Shirley B (2002) Biosynthesis of flavonoids and effects of stress. Curr Opin Plant Biol 5:218–223
Wu KL, Guo ZJ, Wang HH, Li J (2005) The WRKY family of transcription factors in rice and Arabidopsis and their origins. DNA Res 12:9–26
Xie Z, Zhang Z-L, Zou X, Huang J, Ruas P, Thompson D, Shen QJ (2005) Annotations and functional analyses of the rice WRKY gene superfamily reveal positive and negative regulators of abscisic acid signaling in aleurone cells. Plant Physiol 137:176–189
Xie Z, Zhang Z-L, Zou X, Yang G, Komatsu S, Shen Q (2006) Interactions of two abscisic-acid induced WRKY genes in repressing gibberellin signaling in aleurone cells. Plant J 46:231–242
Xu X, Chen C, Fan F, Chen Z (2006) Physical and functional interactions between pathogen-induced Arabidopsis WRKY18, WRKY40, and WRKY60 transcription factors. Plant Cell 18:1310–1326
Yang Y, Shah J, Klessig DF (1997) Signal perception and transduction in plant defense response. Genes Dev 11:1621–1639
Yu D, Chen C, Chen Z (2001) Evidence for an important role of WRKY DNA binding proteins in the regulation of NPR1 gene expression. Plant Cell 13:1527–1540
Zhang Y, Wang L (2005) The WRKY transcription factor superfamily: its origin in eukaryotes and expansion in plants. BMC Evol Biol 5:1–12
Zheng Z, Qamar SA, Chen Z, Mengiste T (2006) Arabidopsis WRKY33 transcription factor is required for resistance to necrotrophic fungal pathogens. Plant J 48:592–605
Zhu Q, Dabi T, Beeche A, Yamamoto R, Lawton MA, Lamb C (1995) Cloning and properties of a rice gene encoding phenylalanine ammonia-lyase. Plant Mol Biol 29:535–550
Acknowledgements
We thank Drs. Xia Y. and Dawson R. W. for critical reading of the manuscript. This research was supported by the State Basic Research and Development Plan (2006CB101905) and National Nature Science of Foundation (30370139 and 30471122).
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Haihua Wang, Junjie Hao, and Xujun Chen contribute equally to this manuscript.
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Wang, H., Hao, J., Chen, X. et al. Overexpression of rice WRKY89 enhances ultraviolet B tolerance and disease resistance in rice plants. Plant Mol Biol 65, 799–815 (2007). https://doi.org/10.1007/s11103-007-9244-x
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DOI: https://doi.org/10.1007/s11103-007-9244-x