Chinese Journal OF Rice Science >
OsLOX10 Positively Regulates Defense Responses of Rice to Rice Blast and Bacterial Blight
Received date: 2021-06-10
Revised date: 2021-09-30
Online published: 2022-07-12
【Objective】Rice blast caused by Magnaporthe oryzae and bacterial blight caused by Xanthomonas oryzae pv. oryzae (Xoo) seriously affect rice yield and quality. The transgenic materials of OsLOX10 were created to evaluate the resistance to M. oryzae and Xoo. The possible mechanism of OsLOX10 in regulating the defense responses to rice blast and bacterial blight were elucidated. 【Method】The knockout vector of OsLOX10 was constructed using CRISPR/Cas9 system, and the overexpression vector of OsLOX10 was constructed by linearizing pCXUN-HA with restriction enzyme XcmⅠ and linking with TA ligation. Transgenic rice was obtained by genetic transformation, and the homozygous transgenic lines were screened for fungal and bacterial resistance analysis. The expression dynamics of SA (salicylic acid) and JA (jasmonic acid) pathway marker genes were analyzed by qRT-PCR after infection with M. oryzae. The outbreak of reactive oxygen species (ROS) induced by chitin and flg22 in rice was also observed. 【Result】qRT-PCR analysis showed that the OsLOX10 expression was up-regulated 24 h after inoculation with M. oryzae and Xoo. The OsLOX10 knockout lines were more susceptible to the rice blast than that of the wild type (Nipponbare) when inoculated with the spore suspension of M. oryzae Guy11, while the overexpressed lines had no typical disease symptoms. At 6, 12, 24 and 36 hours post inoculation, the transcriptional levels of three pathogenesis-related genes, OsPBZ1, OsPR1a and OsPR1b, and two JA pathway genes, OsAOS2 and OsLOX5, and SA pathway gene OsPAL1, were significantly down-regulated in the knockout transgenic lines. However, they were significantly up-regulated in overexpressed transgenic lines. The OsLOX10 transgenic rice was inoculated with Xoo (PXO99A), and the OsLOX10 knockout lines were more susceptible to bacterial blight. qRT-PCR analysis showed that the relative expression levels of OsPR1b, OsPAL1 and the three genes (OsAOS2, OsAOC and OsJAZ) involved in the JA synthesis pathway were up-regulated in OsLOX10 overexpression lines, significantly down-regulated in OsLOX10 knockout lines seven days after Xoo inoculation. OsLOX10 knockout also significantly reduced the ROS level induced by chitin and flg22 in rice, delayed the peaking time of ROS induced by chitin, and reduced the sensitivity of rice to chitin and flg22.【Conclusion】OsLOX10 could be induced by infection with M. oryzae and Xoo. OsLOX10 may be involved in pathogen-associated molecular patterns triggered immunity, and play a positive regulatory role in rice resistance to rice blast and bacterial blight. Moreover, the OsLOX10 may regulate rice resistance to M. oryzae and Xoo through SA and JA mediated signaling pathways.
Key words： OsLOX10; CRISPR/Cas9; disease resistance; rice blast; bacterial blight; signal pathway
ZHOU Yonglin, SHEN Xiaolei, ZHOU Lishuai, LIN Qiaoxia, WANG Zhaolu, CHEN Jing, FENG Huijie, ZHANG Zhenwen, CHEN Xiaoting, LU Guodong . OsLOX10 Positively Regulates Defense Responses of Rice to Rice Blast and Bacterial Blight[J]. Chinese Journal OF Rice Science, 2022 , 36(4) : 348 -356 . DOI: 10.16819/j.1001-7216.2022.210604
| [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] | Ke Y, Deng H, Wang S. Advances in understanding broad-spectrum resistance to pathogens in rice[J]. The Plant Journal, 2017, 90: 738-748. |
| [3] | Liu W, Wang G L. Plant innate immunity in rice: A defense against pathogen infection[J]. National Science Review, 2016, 3: 295-308 |
| [4] | Ngou B P M, Ahn H K, Ding P, Jones J D G. Mutual potentiation of plant immunity by cell-surface and intracellular receptors[J]. Nature, 2021: 1-6. |
| [5] | Yuan M, Jiang Z, Bi G, Nomura K, Liu M, Wang Y, Cai B, Zhou J M, He S Y, Xin X F. Pattern-recognition receptors are required for NLR-mediated plant immunity[J]. Nature, 2021: 1-5. |
| [6] | Umate P. Genome-wide analysis of lipoxygenase gene family in Arabidopsis and rice[J]. Plant Signaling & Behavior, 2011, 6(3): 335-338. |
| [7] | Marla S S, Singh V K. LOX genes in blast fungus (Magnaporthe grisea) resistance in rice[J]. Functional & Integrative Genomics, 2012, 12(2): 265-275. |
| [8] | Mizuno K, Iida T, Takano A, Yokoyama M, Fujimura T. A new 9-lipoxygenase cDNA from developing rice seeds[J]. Plant Cell Physiology, 2003, 44: 1168-1175. |
| [9] | Shirasawa K, Takeuchi Y, Ebitani T, Suzuki Y. Identification of gene for rice (Oryza sativa) seed lipoxygenase-3 involved in the generation of stale flavor and development of SNP markers for lipoxygenase-3 deficiency[J]. Breeding Science, 2008, 58: 169-176. |
| [10] | Gayen D, Ali N, Ganguly M, Paul S, Datta S K. RNAi mediated silencing of lipoxygenase gene to maintain rice grain quality and viability during storage[J]. Plant Cell Tissue and Organ Culture, 2014, 118(2): 229-243. |
| [11] | Gayen D, Ali N, Sarkar S N, Datta S K, Datta K. Down-regulation of lipoxygenase gene reduces degradation of carotenoids of golden rice during storage[J]. Planta, 2014, 242(1): 353-363. |
| [12] | Ma L, Zhu F, Li Z, Zhang J, Li X, Dong J, Wang T. TALEN-based mutagenesis of lipoxygenase LOX3 enhances the storage tolerance of rice (Oryza sativa) seeds[J]. PLoS ONE, 2015, 10(12): e0143877. |
| [13] | Xu H, Wei Y, Zhu Y, Lian L, Xie H, Cai Q, Chen Q, Lin Z, Xie H, Zhang J. Antisense suppression of LOX3 gene expression in rice endosperm enhances seed longevity[J]. Plant Biotechnology Journal, 2015, 13: 526-539. |
| [14] | Huang J, Cai M, Long Q, Liu L, Lin Q, Jiang L, Chen S, Wan J. OsLOX2, a rice type I lipoxygenase, confers opposite effects on seed germination and longevity[J]. Transgenic Research, 2014, 23(4): 643-655 |
| [15] | Ohta H, Shida K, Peng Y L, Furusawa I, Aibara S, Morita Y. A lipoxygenase pathway is activated in rice after infection with the rice blast fungus Magnaporthe grisea[J]. Plant Physiology, 1991, 97: 94-98. |
| [16] | Ma X, Zhang Q, Zhu Q, Liu W, Chen Y, Qiu R, Wang B, Yang Z, Li H, Lin Y, Xie Y, Shen R, Chen S, Wang Z, Chen Y, Guo J, Chen L, Zhao X, Dong Z, Liu Y G. A robust CRISPR/Cas9 system for convenient, high- efficiency multiplex genome editing in monocot and dicot plants[J]. Molecular Plant, 2015, 8(8): 1274-1284. |
| [17] | Chen S, Songkumarn P, Liu J, Wang G L. A versatile zero background T-vector system for gene cloning and functional genomics[J]. Plant Physiology, 2009, 150(3): 1111-1121. |
| [18] | Hong Y, Liu Q, Cao Y, Zhang Y, Cheng S, Cao L. The OsMPK15 negatively regulates Magnaporthe oryza and Xoo disease resistance via SA and JA signaling pathway in rice[J]. Frontiers in Plant Science, 2019, 10: 752 |
| [19] | Wang G, Ding X, Yuan M, Qiu D, Li X, Xu C, Wang S. Dual function of rice OsDR8 gene in disease resistance and thiamine accumulation[J]. Plant Molecular Biology, 2006, 60: 437-449. |
| [20] | Qiu D, Xiao J, Ding X, Xiong M, Cai M, Li X, Xu C, Wang S. OsWRKY13 mediates rice disease resistance by regulating defense-related genes in salicylate- and jasmonate-dependent signaling[J]. Molecular Plant- Microbe Interactions, 2007, 20(5): 492-499. |
| [21] | Qiu D, Xiao J, Xie W, Liu H, Li X, Wang S. rice gene network inferred from expression profiling of plants overexpressing oswrky13, a positive regulator of disease resistance[J]. Molecular Plant, 2008, 1(3): 538-551. |
| [22] | Tao Z, Liu H, Qiu D, Zhou Y, Xu C, Wang S. A pair of allelic WRKY genes play opposite roles in rice-bacteria interactions[J]. Plant Physiology, 2009, 151: 936-948. |
| [23] | Harkenrider M, Sharma R, de Vleesschauwer D, Tsao L, Zhang X, Chern M, Canlas P, Zuo S, Ronald P C. Overexpression of rice wall-associated kinase25 (OsWAK25) alters resistance to bacterial and fungal pathogens[J]. PLoS ONE, 2016, 11: e0147310. |
| [24] | Tonnessen B W, Manosalva P, Lang J M, Baraoidan M, Bordeos A, Mauleon R, Oard J, Leung H, Leach J E. Rice phenylalanine ammonialyase gene OsPAL4 is associated with broad spectrum disease resistance[J]. Plant Molecular Biology, 2015, 87: 273-286. |
| [25] | Dubouzet J G, Maeda S, Sugano S, Ohtake M, Hayashi N, Ichikawa T, Kondou Y, Horii Y, Matsui M, Hirochika H, Takatsuji H, Mori M. Screening for resistance against Pseudomonas syringae in rice-foxarabidopsis lines identified a putative receptor-like cytoplasmic kinase gene that confers resistance to major bacterial and fungal pathogens in Arabidopsis and rice[J]. Plant Biotechnology Journal, 2010, 9: 466-485. |
| [26] | Zhao X, Zhang T, Feng H, Qiu T, Zhao W. OsNBL1, a multi-organelle localized protein, plays essential roles in rice senescence, disease resistance, and salt tolerance[J]. Rice, 2021, 14(1): 10. |
| [27] | Narayanan S P, Lung S C, Liao P, Lo C, Chye M L. The Overexpression of Osacbp5 protects transgenic rice against necrotrophic, hemibiotrophic and biotrophic pathogens[J]. Scientific Reports, 2020, 10(1): 14918. |
| [28] | Han B, Zhou Y, Zhou Z H, Sun B, Zhou F, Yin C X, Ma W H, Chen H, Lin Y J. Repressed OSMESL expression-triggers reactive oxygen species mediated broad-spectrum disease resistance in rice[J]. Plant Biotechnology Journal, 2021, 19(8): 1511-1522 |
| [29] | Mei C, Qi M, Sheng G, Yang Y Y. inducible overexpression of a rice allele oxide synthase gene increases the endogenous jasmonic acid level, PR gene expression, and host resistance to fungal infection[J]. Molecular Plant Microbe Interactions, 2006, 19(10): 1127-1137. |
| [30] | Nalam V J, Alam S, Keereetaweep J, Shah J. Facilitation of Fusarium graminearum infection by 9-lipoxygenases in Arabidopsis and wheat[J]. Molecular Plant Microbe Interactions, 2015, 28(10): 1142-1152 |
| [31] | Christensen S A, Nemchenko A, Borrego E, Murray I, Sobhy I S, Bosak L, Deblasio S, Vaughn K A, Herrfurth C, Tumlinson J, Nansen C, Meeley R, Kolomiets M V. The maize lipoxygenase, zmlox10, mediates green leaf volatile, jasmonate and-herbivore-induced plant volatile production for defense against insect attack[J]. Plant Journal, 2013, 74(1): 59-73. |
| [32] | Xing Q J, Liao J J, Cao S X, Li M, Lü T H, Qi H Y. Cmlox10 positively regulates drought tolerance through jasmonic acid-mediated stomatal closure in oriental melon (Cucumis melo var. Makuwa Makino)[J]. Scientific Reports, 2020, 10: 17452 |
| [33] | Torres M A, Jones J D, Dangl J L. Reactive oxygen species signaling in response to pathogens[J]. Plant Physiology, 2006, 141: 373-378. |
/
| 〈 |
|
〉 |