OsASR2 regulates the expression of a defence‐related gene,Os2H16, by targeting the GT‐1 cis‐element

The GT‐1 cis‐element widely exists in many plant gene promoters. However, the molecular mechanism that underlies the response of the GT‐1 cis‐element to abiotic and biotic stresses remains elusive in rice. We previously isolated a rice short‐chain peptide‐encoding gene, Os2H16, and demonstrated that it plays important roles in both disease resistance and drought tolerance. Here, we conducted a promoter assay of Os2H16 and identified GT‐1 as an important cis‐element that mediates Os2H16 expression in response to pathogen attack and osmotic stress. Using the repeated GT‐1 as bait, we characterized an abscisic acid, stress and ripening 2 (ASR2) protein from yeast‐one hybridization screening. Sequence alignments showed that the carboxy‐terminal domain of OsASR2 containing residues 80–138 was the DNA‐binding domain. Furthermore, we identified that OsASR2 was specifically bound to GT‐1 and activated the expression of the target gene Os2H16, as well as GFP driven by the chimeric promoter of 2 × GT‐1‐35S mini construct. Additionally, the expression of OsASR2 was elevated by pathogens and osmotic stress challenges. Overexpression of OsASR2 enhanced the resistance against Xanthomonas oryzae pv. oryzae and Rhizoctonia solani, and tolerance to drought in rice. These results suggest that the interaction between OsASR2 and GT‐1 plays an important role in the crosstalk of the response of rice to biotic and abiotic stresses.     

Overexpression of a rice heme activator protein gene (OsHAP2E) confers resistance to pathogens,salinity and drought,and increases photosynthesis and tiller number

Heme activator protein (HAP), also known as nuclear factor Y or CCAAT binding factor (HAP/NF‐Y/CBF), has important functions in regulating plant growth, development and stress responses. The expression of rice HAP gene (OsHAP2E) was induced by probenazole (PBZ), a chemical inducer of disease resistance. To characterize the gene, the chimeric gene (OsHAP2E::GUS) engineered to carry the structural gene encoding β‐glucuronidase (GUS) driven by the promoter from OsHAP2E was introduced into rice. The transgenic lines of OsHAP2Ein::GUS with the intron showed high GUS activity in the wounds and surrounding tissues. When treated by salicylic acid (SA), isonicotinic acid (INA), abscisic acid (ABA) and hydrogen peroxide (H₂O₂), the lines showed GUS activity exclusively in vascular tissues and mesophyll cells. This activity was enhanced after inoculation with Magnaporthe oryzae or Xanthomonas oryzae pv. oryzae. The OsHAP2E expression level was also induced after inoculation of rice with M. oryzae and X. oryzae pv. oryzae and after treatment with SA, INA, ABA and H₂O_2, respectively. We further produced transgenic rice overexpressing OsHAP2E. These lines conferred resistance to M. oryzae or X. oryzae pv. oryzae and to salinity and drought. Furthermore, they showed a higher photosynthetic rate and an increased number of tillers. Microarray analysis showed up‐regulation of defence‐related genes. These results suggest that this gene could contribute to conferring biotic and abiotic resistances and increasing photosynthesis and tiller numbers.     

Overexpression of <Emphasis Type="Italic">OsDT11</Emphasis>, which encodes a novel cysteine-rich peptide,enhances drought tolerance and increases ABA concentration in rice

Short-chain peptides play important roles in plant development and responses to abiotic and biotic stresses. Here, we characterized a gene of unknown function termed OsDT11, which encodes an 88 amino acid short-chain peptide and belongs to the cysteine-rich peptide family. It was found that the expression of OsDT11 can be activated by polyethylene glycol (PEG) treatment. Compared with wild-type lines, the OsDT11-overexpression lines displayed dramatically enhanced tolerance to drought and had reduced water loss, reduced stomatal density, and an increased the concentration of abscisic acid (ABA). The suppression of OsDT11 expression resulted in an increased sensitivity to drought compared to wild-type expression. Several drought-related genes, including genes encoding abscisic acid (ABA) signaling markers, were also strongly induced in the OsDT11-overexpressing lines. Moreover, the expression of OsDT11 was repressed in ABA-insensitive mutant Osbzip23 and Os2H16 RNAi lines. These results suggest that OsDT11-mediated drought tolerance may be dependent on the ABA signaling pathway.     

Design,synthesis, and antibacterial activity against rice bacterial leaf blight and leaf streak of 2,5-substituted-1,3,4-oxadiazole/thiadiazole sulfone derivative

A series of 2,5-substituted-1,3,4-oxadiazole/thiadiazole sulfone derivatives were synthesized and evaluated for their antibacterial activities against rice bacterial leaf blight and leaf streak caused by Xanthomonas oryzae pv. oryzae and Xanthomonas oryzae pv. oryzicolaby via the turbidimeter test in vitro. Antibacterial bioassay results indicated that most compounds demonstrated good inhibitory effect antibacterial bioactivities against rice bacterial leaf blight and leaf streak. Among the title compounds, compound 6c demonstrated the best inhibitory effect against rice bacterial leaf blight and leaf streak with half-maximal effective concentration (EC₅₀) values of 1.07 and 7.14 μg/mL, respectively, which were even better than those of commercial agents such as Bismerthiazol and Thiediazole Copper. In vivo antibacterial activities tests at greenhouse conditions demonstrated that the controlling effect of compounds 6c (43.5%) and 6g (42.4%) against rice bacterial leaf blight were better than those of Bismerthiazol (25.5%) and Thiediazole Copper (37.5%).     

Differential response of cultivated rice to pathogen challenge and abiotic stresses with reference to cationic peroxidase

Seedlings of cultivated rice variety ADT43 was investigated after challenging with two different abiotic (drought and salinity) and biotic (sheath blight and bacterial leaf blight pathogens) stresses. Salinity and drought stress reduced the growth of seedlings, mainly the higher conditions (100 mM NaCl and 10?days of drought, respectively). Increased level of MDA content was observed in biotic and abiotic-stress treated seedlings. The highest H₂O₂ content was observed under salinity-stressed seedlings and lower level observed under biotic stress. Superoxide dismutase activity showed a gradual decrease in all stress conditions compared to control. Salinity stress resulted in highest activity of catalase compared to biotic stress. The peroxidase activity of the seedlings was found to be increased under salt and drought stress conditions and the activity decreased under biotic stress. Drought stress resulted in induced expression of POC1 gene whereas the biotic stress showed lower expression level. Suppression of the rice peroxidase would have been the mechanism of overcoming the intrinsic defence in rice by these pathogens.     

Marker-assisted breeding of Thai fragrance rice for semi-dwarf phenotype, submergence tolerance and disease resistance to rice blast and bacterial blight

Rice is a staple food crop for more than half of the world’s population. However, rice production is affected by many types of abiotic and biotic stress. Genetic breeding by utilizing natural resistance or tolerance genes is the most economic and efficient way to combat or adapt to these stresses. Khao Dawk Mali 105 (KDML 105) is an elite cultivar of aromatic rice mainly grown in Thailand. However, the production of KDML 105 is affected by lodging problems due to its tall plant type, regular flash floods or short-term submergence during the monsoon season, and diseases such as blast and bacterial blight. Here we report the pyramiding of semi-dwarf gene sd1, submergence tolerance gene Sub1A, blast resistance gene Pi9 and bacterial blight resistance genes Xa21 and Xa27 in KDML 105 by marker-assisted selection. The improved line, designated T5105, has a semi-dwarf phenotype with improved lodging resistance and a greater harvest index. T5105 survives after 2 weeks of complete submergence without significant loss of viability. T5105 confers high resistance to all five Magnaporthe oryzae isolates tested and provides resistance or moderate resistance to 25 of the 27 Xanthomonas oryzae pv. oryzae strains tested. In addition, T5105 produced higher yield than KDML 105 in two field trials and retains similar good grain quality to KDML 105. The development of T5105 provides a new line to boost the production of high-quality aromatic rice in tropical regions.     

Jumonji C Domain Protein JMJ705-Mediated Removal of Histone H3 Lysine 27 Trimethylation Is Involved in Defense-Related Gene Activation in Rice

Histone methylation is an important epigenetic modification in chromatin function, genome activity, and gene regulation. Dimethylated or trimethylated histone H3 lysine 27 (H3K27me2/3) marks silent or repressed genes involved in developmental processes and stress responses in plants. However, the role and the mechanism of the dynamic removal of H3K27me2/3 during gene activation remain unclear. Here, we show that the rice (Oryza sativa) Jumonji C (jmjC) protein gene JMJ705 encodes a histone lysine demethylase that specifically reverses H3K27me2/3. The expression of JMJ705 is induced by stress signals and during pathogen infection. Overexpression of the gene reduces the resting level of H3K27me2/3 resulting in preferential activation of H3K27me3-marked biotic stress-responsive genes and enhances rice resistance to the bacterial blight disease pathogen Xanthomonas oryzae pathovar oryzae. Mutation of the gene reduces plant resistance to the pathogen. Further analysis revealed that JMJ705 is involved in methyl jasmonate–induced dynamic removal of H3K27me3 and gene activation. The results suggest that JMJ705 is a biotic stress-responsive H3K27me2/3 demethylase that may remove H3K27me3 from marked defense-related genes and increase their basal and induced expression during pathogen infection.     

Proteomic analysis of bacterial blight defence signalling pathway using transgenic rice overexpressing thaumatin-like protein

Rice overexpressed thaumatin-like protein gene and the proteins from the leaf blades of 2-week-old transgenic rice seedlings were fractionated into cytosolic and membrane fractions, and separated by two-dimensional polyacrylamide gel electrophoresis and stained with Commassie brilliant blue. Among of 440 detected proteins, 5 proteins were up-regulated and 5 proteins were down-regulated by the overexpression of thaumatin-like protein. In the sense thaumatin-like protein transgenic rice and/or in rice inoculated with Xanthomonas oryzae pv. oryzae (Xo7435), 2-cys peroxiredoxin, thaumatin-like protein and glycine cleavage H protein were up-regulated, while oxygen evolving complex protein 2 was down-regulated. These results suggest that thaumatin-like protein-mediated disease resistance of rice against bacterial blight disease is the results of changes in proteins related to oxidative stress and energy metabolism in addition to changes in proteins related to defence.     

Detection of <Emphasis Type="Italic">Xanthomonas oryzae</Emphasis> pv. <Emphasis Type="Italic">oryzae</Emphasis> in seeds using a specific TaqMan probe

Xanthomonas oryzae pv. oryzae is the pathogen that causes bacterial leaf blight in rice. Bacterial leaf blight is the main cause for severe rice underproduction in many countries. However, with conventional methods it is difficult to quickly and reliably distinguish this pathogen from other closely related pathogenic bacteria, especially X. oryzae pv. oryzicola, the causal organism of bacterial leaf streak in rice. We have developed a novel and highly sensitive real-time method for the identification of this specific bacteria based on a TaqMan probe. This probe is designed to recognize the sequence of a putative siderophore receptor gene cds specific to X. oryzae pv. oryzae, and can be identified from either a bacterial culture or naturally infected rice seeds and leaves in only 2 h. The sensitivity of the method is 100 times higher than that of the current polymerase chain reaction (PCR) gel electrophoresis method for diagnosis.     

New PCR-based sequence-tagged site marker for bacterial blight resistance gene Xa38 of rice

Bacterial blight (BB), caused by Xanthomonas oryzae pv. oryzae, is a major disease of rice managed largely through the deployment of resistance genes. Xa38, a BB resistance gene identified from Oryza nivara acc. IRGC 81825, was mapped on chromosome 4L in a 38.4-kb region. The closely linked markers for this gene, identified earlier, were simple sequence repeat marker RM17499 and sequence-tagged site markers developed from loci Os04g53060 and Os04g53120. Marker Os04g53060 is dominant while the other two markers show smaller size differences difficult to resolve accurately on agarose gel. Based on gene annotation, three nucleotide binding site?Cleucine-rich repeat genes present in the target region were cloned from O. nivara and sequenced. One of the loci, LOC_Os04g53050, had a 48-base-pair deletion in O. nivara acc. IRGC 81825 compared to the cultivated rice. Primers were designed around the deletion and the resulting marker is codominant and easy to score in agarose gel. The newly designed marker co-segregated with Xa38, amplifying products of 269?bp in O. nivara and 317?bp in cultivated rice. This marker could be more useful for marker-assisted selection than ones reported earlier.