Malus hupehensis NPR1 induces pathogenesis-related protein gene expression in transgenic tobacco

Most commercially grown apple cultivars are susceptible to fungal diseases. Malus hupehensis has high resistance to many diseases affecting apple cultivars. Understanding innate defence mechanisms would help to develop disease-resistant apple crops. Non-expressor of pathogenesis-related genes 1 (NPR1) plays a key role in regulating salicylic acid (SA)-mediated systemic acquired resistance (SAR). MhNPR1 cDNA, corresponding to genomic DNA and its 5' flanking sequences, was isolated from M. hupehensis. Sequence analysis showed that the regulatory mechanism for oligomer-monomer transition of the MhNPR1 protein in apple might be similar to that of GmNPR1 in soybean, but different from that of AtNPR1 in Arabidopsis. No significant differences in MhNPR1 expression were found in M. hupehensis after infection with Botryosphaeria berengeriana, showing that MhNPR1 might be regulated by pathogens at the protein level, as described for Arabidopsis and grapevine. SA treatment significantly induced MhNPR1 expression in leaves, stems and roots, while methyl jasmonate (MeJA) treatment induced MhNPR1 expression in roots, but not in leaves or stems. The expression of MhNPR1 was highly increased in roots, moderately in leaves, and did not change in stems after treatment with 1-aminocyclopropane-1-carboxylic acid (ACC). SAR marker genes (MhPR1 and MhPR5) were induced by SA, MeJA and ACC in leaves, stems and roots. Overexpression of MhNPR1 significantly induced the expression of pathogenesis-related genes (NtPR1, NtPR3 and NtPR5) in transgenic tobacco plants and resistance to the fungus Botrytis cinerea, suggesting that MhNPR1 orthologues are a component of the SA defence signalling pathway and SAR is induced in M. hupehensis.     

Induction of resistance to Verticillium dahliae in Arabidopsis thaliana by the biocontrol agent K-165 and pathogenesis-related proteins gene expression

The biocontrol bacterium Paenibacillus alvei K165 has the ability to protect Arabidopsis thaliana against Verticillium dahliae. A direct antagonistic action of strain K165 against V. dahliae was ruled out, making it likely that K165-mediated protection results from induced systemic resistance (ISR) in the host. K165-mediated protection was tested in various Arabidopsis mutants and transgenic plants impaired in defense signaling pathways, including NahG (transgenic line degrading salicylic acid [SA]), etr1-1 (insensitive to ethylene), jar1-1 (insensitive to jasmonate), npr1-1 (nonexpressing NPR1 protein), pad3-1 (phytoalexin deficient), pad4-1 (phytoalexin deficient), eds5/sid1 (enhanced disease susceptibility), and sid2 (SA-induction deficient). ISR was blocked in Arabidopsis mutants npr1-1, eds5/sid1, and sid2, indicating that components of the pathway from isochorismate and a functional NPR1 play a crucial role in the K165-mediated ISR. Furthermore, the concomitant activation and increased transient accumulation of the PR-1, PR-2, and PR-5 genes were observed in the treatment in which both the inducing bacterial strain and the challenging pathogen were present in the rhizosphere of the A. thaliana plants.     

拟南芥NPR1基因的克隆与表达载体的构建

NPR1基因为植物抗病基因表达和系统获得性抗性中的一个关键基因。该文以DNA PCR扩增的方法,从拟南芥基因组DNA中克隆出NPR1基因,通过序列分析,所克隆的 NPR1 基因与报道的基因序列完全一致。将其构建成植物表达载体,为今后植物抗病基因工程的开展奠定了基础。     

The CCCH zinc finger protein gene AtZFP1 improves salt resistance in Arabidopsis thaliana

The CCCH type zinc finger proteins are a super family involved in many aspects of plant growth and development. In this study, we investigated the response of one CCCH type zinc finger protein AtZFP1 (At2g25900) to salt stress in Arabidopsis. The expression of AtZFP1 was upregulated by salt stress. Compared to transgenic strains, the germination rate, emerging rate of cotyledons and root length of wild plants were significantly lower under NaCl treatments, while the inhibitory effect was significantly severe in T-DNA insertion mutant strains. At germination stage, it was mainly osmotic stress when treated with NaCl. Relative to wild plants, overexpression strains maintained a higher K⁺, K⁺/Na⁺, chlorophyll and proline content, and lower Na⁺ and MDA content. Quantitative real-time PCR analysis revealed that the expression of stress related marker genes KIN1, RD29B and RD22 increased more significantly in transgenic strains by salt stress. Overexpression of AtZFP1 also enhanced oxidative and osmotic stress tolerance which was determined by measuring the expression of a set of antioxidant genes, osmotic stress genes and ion transport protein genes such as SOS1, AtP5CS1 and AtGSTU5. Overall, our results suggest that overexpression of AtZFP1 enhanced salt tolerance by maintaining ionic balance and limiting oxidative and osmotic stress.     

Uncoupling PR gene expression from NPR1 and bacterial resistance: characterization of the dominant Arabidopsis cpr6-1 mutant.

In Arabidopsis, NPR1 mediates the salicylic acid (SA)-induced expression of pathogenesis-related (PR) genes and systemic acquired resistance (SAR). Here, we report the identification of another component, CPR 6, that may function with NPR1 in regulating PR gene expression. The dominant CPR 6-1 mutant expresses the SA/NPR1-regulated PR genes (PR-1, BGL 2, and PR-5) and displays enhanced resistance to Pseudomonas syringae pv maculicola ES4326 and Peronospora parasitica Noco2 in the absence of SAR induction. cpr 6-1-induced PR gene expression is not suppressed in the cpr 6-1 npr1-1 double mutant but is suppressed when SA is removed by salicylate hydroxylase. Thus, constitutive PR gene expression in cpr 6-1 requires SA but not NPR1. In addition, resistance to P. s. maculicola ES4326 is suppressed in the cpr 6-1 npr1-1 double mutant, despite expression of PR-1, BGL 2, and PR-5. Resistance to P. s. maculicola ES4326 must therefore be accomplished through unidentified antibacterial gene products that are regulated through NPR1. These results show that CPR 6 is an important regulator of multiple signal transduction pathways involved in plant defense.     

Biologically induced systemic acquired resistance in Arabidopsis thaliana

Local infection with a necrotizing pathogen can render plants resistant to subsequent infection by normally virulent pathogens. A system for biological induction of such systemic acquired resistance (SAR) in Arabidopsis thaliana is reported. When plants were immunized by local inoculation of a single leaf with avirulent Pseudomonas syringae pv. tomato (Pst) carrying the avrRpt2 avirulence gene, after 2 days other leaves became resistant, as measured symptomatically and by in planta bacterial growth, to challenge with a virulent Pst strain lacking this avirulence gene. Resistance was systemic and protected the plants against infection by other virulent pathogens including P. syringae pv. maculicola. Low-dose inoculation induced a strong SAR and double immunizations did not increase the level of protection indicating that the response of only a few cells to the immunizing bacteria is required. SAR was not induced by the virulent strain of Pst lacking avrRpt2. However, experiments with the Arabidopsis RPS2 disease resistance gene mutant rps2-201, which does not exhibit a local hypersensitive response to Pst carrying the corresponding avirulence gene avrRpt2, indicate that a hypersensitive response contributes to, but is not essential for, the induction of SAR. Thus, avrRpt2 activates either a branching signal pathway or separate parallel pathways for induction of localized hypersensitive resistance and SAR, with downstream potentiation of the systemic response by the local response. Using this system for the biological induction of SAR in Arabidopsis, it should be possible to dissect the molecular genetics of SAR by the isolation of mutants affected in the production, transmission, perception and transduction of the systemic signal(s).     

Hydrogen peroxide-induced gene expression in Arabidopsis thaliana

Hydrogen peroxide (H(2)O(2)) is generated in plants after exposure to a variety of biotic and abiotic stresses, and has been shown to induce a number of cellular responses. Previously, we showed that H(2)O(2) generated during plant-elicitor interactions acts as a signaling molecule to induce the expression of defense genes and initiate programmed cell death in Arabidopsis thaliana suspension cultures. Here, we report for the first time the identification by RNA differential display of four genes whose expression is induced by H(2)O(2). These include genes that have sequence homology to previously identified Arabidopsis genes encoding a late embryogenesis-abundant protein, a DNA-damage repair protein, and a serine/threonine kinase. Their putative roles in H(2)O(2)-induced defense responses are discussed.     

Genetically engineered resistance to Fusarium head blight in wheat by expression of Arabidopsis NPR1

Fusarium head blight (FHB) is a devastating disease of wheat and barley which causes extensive losses worldwide. Monogenic, gene-for-gene resistance to FHB has not been reported. The best source of resistance to FHB is a complex, quantitative trait derived from the wheat cv. Sumai 3. Here, we show that the Arabidopsis thaliana NPR1 gene (AtNPR1), which regulates the activation of systemic acquired resistance, when expressed in the FHB-susceptible wheat cv. Bobwhite, confers a heritable, type II resistance to FHB caused by Fusarium graminearum. The heightened FHB resistance in the transgenic AtNPRI -expressing wheat is associated with the faster activation of defense response when challenged by the fungus. PR1 expression is induced rapidly to a high level in the fungus-challenged spikes of the AtNPR1-expressing wheat. Furthermore, benzothiadiazole, a functional analog of salicylic acid, induced PR1 expression faster and to a higher level in the AtNPR1-expressing wheat than in the nontransgenic plants. We suggest that FHB resistance in the AtNPR1-expressing wheat is a result of these plants being more responsive to an endogenous activator of plant defense. Our results demonstrate that NPR1 is an effective candidate for controlling FHB.