Rice NSF45K microarray experiment to explain Xa21, NH1 and NRR mediated defense responses in rice
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ABSTRACT: Rice Xa21 resistance gene, which encodes a protein with predicted leucine-rich repeat (LRR), transmembrane, juxtamembrane, and intracellular kinase domains, conferred immunity to diverse strains of Xanthomonas oryzae pv. oryzae (Xoo). We generated Xa21 plant on TP309 background (Oryza Sativa Japonica). Systemic Acquired Resistance (SAR) in plants confers durable broad-spectrum resistance to pathogens and requires a phytohormone, salicylic acid (SA). Arabidopsis NPR1/NIM1 is a key regulator of the SAR response. Recently, we found that rice NPR1 homolog 1 (NH1) mediated enhanced resistance responses for Xoo (Chern et al., 2005b). We further investigated relating pathways in rice by identifying proteins that interact with NH1. One of them, constitutive over-expression of NH1 mediated negative regulator of resistance (NRR) gene caused enhanced susceptibility to Xoo , indicating that this gene product negatively affects to basal resistance response (Chern et al., 2005a). To dissect defense responses for rice bacterial blight pathogen, we planed microarray using two resistant mutant named with Xa21-TP309, NH1ox and one super-susceptible mutant (NRRox) before pathogen inoculation and one day post pathogen inoculation. Keywords: Biotic stress response Two or Three-condition experiment, NH1ox vs wild type control (LG) at two durations of Xoo inoculation (0d and 1d); NRRox vs wild type control (LG) at two durations of Xoo inoculation (0d and 1d); and Xa21vs wild type control (TP309) at three durations of Xoo inoculation (0d,1d and 2d);. Biological replicates: 2 or 4, independently grown and harvested.
Project description:Rice Xa21 resistance gene, which encodes a protein with predicted leucine-rich repeat (LRR), transmembrane, juxtamembrane, and intracellular kinase domains, conferred immunity to diverse strains of Xanthomonas oryzae pv. oryzae (Xoo). We generated Xa21 plant on TP309 background (Oryza Sativa Japonica). Systemic Acquired Resistance (SAR) in plants confers durable broad-spectrum resistance to pathogens and requires a phytohormone, salicylic acid (SA). Arabidopsis NPR1/NIM1 is a key regulator of the SAR response. Recently, we found that rice NPR1 homolog 1 (NH1) mediated enhanced resistance responses for Xoo (Chern et al., 2005b). We further investigated relating pathways in rice by identifying proteins that interact with NH1. One of them, constitutive over-expression of NH1 mediated negative regulator of resistance (NRR) gene caused enhanced susceptibility to Xoo , indicating that this gene product negatively affects to basal resistance response (Chern et al., 2005a). To dissect defense responses for rice bacterial blight pathogen, we planed microarray using two resistant mutant named with Xa21-TP309, NH1ox and one super-susceptible mutant (NRRox) before pathogen inoculation and one day post pathogen inoculation. Keywords: Biotic stress response
Project description:Previously, we successfully introduce the bacterial blight resistance trait from Oryza meyeriana into O. sativa using asymmetric somatic hybridization with O. meyeriana as the donor species. After years of breeding, a progeny named Y73 was generated with recurrent parent O. sativa L. ssp. japonica cv. Dalixiang, and it shows high resistance to broad-spectrum of bacterial blight pathogens Xanthomonas oryzae pv. Oryzae (Xoo). However, the resistance mechanism of Y73 is remain undiscovered. To provide insights into the high resistance phenotype of these plants, we examined the transcriptome response in leaves of Y73 to the bacterial blight infection in this study. Xoo inoculated and mock inoculated rice plants were grown in growth room and the global analysis of gene expression events in rice leaves at 24 hours post inoculation (hpi) were analyzed using Affymetrix Rice GeneChip microarrays. We used microarrays to detail the global programme of gene expression underlying Xoo infection in rice Y73. To find out pathways and genes involved in its high and board-spectrum resistance, microanalysis were carried out on Y73 after Xoo infection at 24 hours post inoculation (hpi). Three independant replicates were perfomed for each treatments.
Project description:Xanthomonas oryzae pv. oryzae (Xoo), the causative agent of bacterial blight disease, is one of the major threats to rice productivity. Yet, the molecular mechanism of rice-Xoo interaction is elusive. Here, we report comparative proteome profiles of Xoo susceptible (Dongjin) and resistant (Hwayeong) cultivars of rice in response to two-time points (3 and 6 days) of Xoo infection. Low-abundance proteins were enriched using a protamine sulfate (PS) precipitation method and isolated proteins were quantified by a label-free quantitative analysis, leading to the identification of 3846 protein groups. Of these, 1128 proteins were significantly changed between mock and Xoo infected plants of Dongjin and Hwayeong cultivars. Based on the abundance pattern and functions of the identified proteins, a total of 23 candidate proteins were shortlisted that potentially participate in plant defense against Xoo in the resistant cultivar. Of these candidate proteins, a mitochondrial arginase-1 showed Hwayeong specific abundance and was significantly accumulated following Xoo inoculation. Overexpression of arginase-1 in susceptible rice cultivar (Dongjin) resulted in enhanced tolerance against Xoo as compared to the wild-type (WT). In addition, expression analysis of defense-related genes encoding PR1, glucanase I, and chitinase II by qRT-PCR showed their enhanced expression in the overexpression lines as compared to WT. Mitochondrial localization of the selected arginase was further confirmed by fluorescent microscopy using GFP-tagged arginase. Taken together, our results uncover the proteome changes in the rice cultivars and highlight the functions of arginase in plant defense against Xoo.
Project description:Purpose: The goal of this study is to identify small non-conding RNAs which are involved in rice resistance to Xoo. Methods: Rice leaves were inoculated with the Xoo strain PXO61 at the four-leaf to five-leaf stage by the leaf-clipping method. Control rice plants were inoculated with water (mock inoculation). And then, total RNA was extracted to be sequenced using Illumina GAIIx. Results: Using an optimized data analysis workflow to count the expression level of small ncRNA, we found several differentially expressed small ncRNA which may be participated in the interaction between rice and Xoo. Conclusions: Small ncRNA have be found to function in a variety of biological processes. Our study here has showed that several candidate miRNA or siRNA may play a significant role in rice immunity. Plants were inoculated with the Xoo strain PXO61 at the four-leaf to five-leaf stage by the leaf-clipping method. Control rice plants were inoculated with water (mock inoculation). Samples were collected before inoculation (ck) and at 2, 4, and 24 hours after PXO61 or mock inoculation from Rb49 and MDJ8. Leaf fragments approximately 2 cm in length that were immediately next to the inoculation site were collected.
Project description:affy_xoo_rice - affy_xoo_rice - The Bacterial Leaf Blight disease of rice is due to Xanthomonas oryzae pv. oryzae. As for many pathogenic bacteria, it relies on a type 3 secretion system that is devoted to the injection of type 3 effectors into the eukaryotic host cell. These proteins are meant to suppress host basal defense responses and/or mimic some host regulatory function promoting bacterial survey in the plant. We are interested in the functional analysis of a subgroup of Xoo T3Es, that are specialized in host cell transcriptome remodelling. These effectors, therefore called TAL for Transcription Activator-Like proteins (also named AvrBs3/PthA-like), are often key virulence factors essential to Xoo pathogenicity such as the effector protein Talc of african Xoo strain BAI3. Our goal is to understand its function during disease development, by identifying rice host genes that are being directly up- or down-regulated by Talc. To that end, we aim at performing Affymetrix transcriptomic analysis, comparing leaf samples of a susceptible rice line inoculated with Xoo to leaves challenged with a Talc-deficient mutant and water-treated leaves. Highly induced genes are likely to be Talc primary targets and therefore potentially good susceptibility gene candidates.-The goal of the experiment is to identify the rice genes up- or down-regulated by the type III effector Talc from Xoo African strain BAI3, upon the inoculation of susceptible rice leaves 24 hours post-infection. To that end, the experimental design includes the inoculation of Nipponbare rice leaves with the virulent Xoo strain BAI3, that will be compared to Nipponbare rice leaves inoculated with a talc K.O. mutant strain and water. Keywords: wt vs virulence mutant 9 arrays - rice
Project description:Bacterial blight, caused by Xanthomonas oryzae pv. oryzae (Xoo), is the most destructive bacterial disease of rice. The cloned rice gene Xa21 confers resistance to a broad spectrum of Xoo races. To identify other genes involved in Xa21-mediated immunity, a whole-genome oligonucleotide microarray of rice was used to profile the expression of rice genes between incompatible interactions and mock treatments at 0, 4, 8, 24, 72 and 120 hours post inoculation (hpi) or between incompatible and compatible interactions at 4 hpi, respectively. A total of 441 differentially expressed genes, designated as XDGs (Xa21 mediated Differentially expressed Genes), were identified. Based on their functional annotations, the XDGs were assigned to 14 categories some of which encode resistance/defense related proteins and signaling regulators. Interestingly, most signaling-related genes were down-regulated at 4 and 8 hpi, suggesting that negative regulation of cellular signaling may play a role in the Xa21-mediated defense response. Moreover, a number of pathogenesis-related genes were induced at 72 and 120 hpi. Comparison of expression profiles mediated by other resistance genes revealed an interesting common responses in the plant immune system. Comparison analyses also suggest possible overlaps with hormone signaling pathways and with defense systems including basal defense and hypersensitive responses.
Project description:Bacterial blight, caused by Xanthomonas oryzae pv. oryzae (Xoo), is the most destructive bacterial disease of rice. The cloned rice gene Xa21 confers resistance to a broad spectrum of Xoo races. To identify other genes involved in Xa21-mediated immunity, a whole-genome oligonucleotide microarray of rice was used to profile the expression of rice genes between incompatible interactions and mock treatments at 0, 4, 8, 24, 72 and 120 hours post inoculation (hpi) or between incompatible and compatible interactions at 4 hpi, respectively. A total of 441 differentially expressed genes, designated as XDGs (Xa21 mediated Differentially expressed Genes), were identified. Based on their functional annotations, the XDGs were assigned to 14 categories some of which encode resistance/defense related proteins and signaling regulators. Interestingly, most signaling-related genes were down-regulated at 4 and 8 hpi, suggesting that negative regulation of cellular signaling may play a role in the Xa21-mediated defense response. Moreover, a number of pathogenesis-related genes were induced at 72 and 120 hpi. Comparison of expression profiles mediated by other resistance genes revealed an interesting common responses in the plant immune system. Comparison analyses also suggest possible overlaps with hormone signaling pathways and with defense systems including basal defense and hypersensitive responses. We investigated the transcriptional characteristics of Xa21-mediated defense responses in rice using whole-genome oligonucleotide microarray, by comparing the expression profiles between incompatible interactions and mock treatments at five time-points and between incompatible and compatible interactions at 4 hours post inoculation. Microarray hybridization at each time point was carried out in three replications including two biological replications and one technological replication. To detect the genes that are significantly regulated and to eliminate those that have inconsistent expression data among replicated experiments, a cutoff of 1.5 fold change and 1.96 for intensity-dependent Z-scores were required in at least two experimental samples.
Project description:Rice-Xanthomonas oryzae is an economically important pathosystem owing to the loss caused by bacterial blight disease. Understanding the moleulcar dynbamics that occur during rice-Xoo interaction is crucial for understanding disaease susceptible and resistance mechanisms. SM210, harbors the resistance alleles xa5, xa13, and Xa21 and is resistant against Xoo. Transcriptome analysis of Xoo-treated. SM210 and SM, its parent variety, indicated different pathways that are altered in these varieties after Xoo treatment.
Project description:With the availability of a high quality draft rice genome sequence, large mutant collections, and gene expression oligo arrays for rice, we are now well positioned to dissect rice defense pathways. To do this, we performed global expression analyses to identify genes that are differentially expressed in 10 mutant lines (i.e., Xa21, NH1ox, NRR1ox, GR978, spl11, spl17, NBS2-PI9ox, MPK5ox, OsCoi1ox, and OsNahG1ox), exhibiting altered defense responses. As controls, we used wild typle varieties same with these mutants. Two-condition experiment, 10 mutants vs wild type control with treatment or without treatment. Biological replicates: 2-4 control, independently grown and harvested. Technical replicates: 1-2 control. One replicate per array.
Project description:affy_riz_2011_7 - affy_riz_2011_7 - The Bacterial Leaf Blight disease of rice is due to Xanthomonas oryzae pv. oryzae. As for many pathogenic bacteria, it relies on a type 3 secretion system (TTSS) that is devoted to the injection of type 3 effectors (T3Es) into the eukaryotic host cell. These proteins are meant to suppress host basal defense responses and/or mimic some host regulatory function promoting bacterial survey in the plant. During an incompatible interaction, T3Es may act as Avr proteins and stimulate Effector-Triggered-Immunity. We aim at evaluating the transcriptomic response of rice leaves challenged with avirulent strains of Xoo BAI3 and MAI1 on resistant lines IR64 and IRBB4 versus the reference susceptible rice line Nipponbare. In addition, we investigated the transcriptomic response of rice leaves upon inoculation of an XoohrcC mutant strain affected in the production of a functional TTSS.-The goal of the experiment is to characterize the rice leaf transcriptome response, upon the inoculation of susceptible and resistant rice leaves 24 hours post-infection. To that end, the experimental design includes the inoculation of susceptible Nipponbare rice leaves with Xoo strains BAI3 (race A1) and MAI1 (race A3), that will be compared to the response of resistant lines IRBB4 and IR64 rice lines. In addition, Nipponbare rice leaves will also be challenged with the BAI3hrcC mutant that is affected in the production of a functional TTSS. 18 arrays - rice; avirulent vs virulent