Hydrogen peroxide generation by the pepper extracellular peroxidase CaPO2 activates local and systemic cell death and defense response to bacterial pathogens

Reactive oxygen species (ROS) are responsible for mediating cellular defense responses in plants. Controversy has existed over the origin of ROS in plant defense. We have isolated a novel extracellular peroxidase gene, CaPO2, from pepper (Capsicum annuum). Local or systemic expression of CaPO2 is induced in pepper by avirulent Xanthomonas campestris pv vesicatoria (Xcv) infection. We examined the function of the CaPO2 gene in plant defense using the virus-induced gene silencing technique and gain-of-function transgenic plants. CaPO2-silenced pepper plants were highly susceptible to Xcv infection. Virus-induced gene silencing of the CaPO2 gene also compromised hydrogen peroxide (H(2)O(2)) accumulation and hypersensitive cell death in leaves, both locally and systemically, during avirulent Xcv infection. In contrast, overexpression of CaPO2 in Arabidopsis (Arabidopsis thaliana) conferred enhanced disease resistance accompanied by cell death, H(2)O(2) accumulation, and PR gene induction. In CaPO2-overexpression Arabidopsis leaves infected by Pseudomonas syringae pv tomato, H(2)O(2) generation was sensitive to potassium cyanide (a peroxidase inhibitor) but insensitive to diphenylene iodonium (an NADPH oxidase inhibitor), suggesting that H(2)O(2) generation depends on peroxidase in Arabidopsis. Together, these results indicate that the CaPO2 peroxidase is involved in ROS generation, both locally and systemically, to activate cell death and PR gene induction during the defense response to pathogen invasion.     

Cotton GhMKK5 affects disease resistance, induces HR-like cell death, and reduces the tolerance to salt and drought stress in transgenic Nicotiana benthamiana

Mitogen-activated protein kinase (MAPK) cascades are involved in various processes from plant growth and development to biotic and abiotic stress responses. MAPK kinases (MAPKKs), which link MAPKs and MAPKK kinases (MAPKKKs), play crucial roles in MAPK cascades to mediate a variety of stress responses in plants. However, few MAPKKs have been functionally characterized in cotton (Gossypium hirsutum). In this study, a novel gene, GhMKK5, from cotton belonging to the group C MAPKKs was isolated and characterized. The expression of GhMKK5 can be induced by pathogen infection, abiotic stresses, and multiple defence-related signal molecules. The overexpression of GhMKK5 in Nicotiana benthamiana enhanced the plants' resistance to the bacterial pathogen Ralstonia solanacearum by elevating the expression of pathogen resistance (PR) genes, including PR1a, PR2, PR4, PR5, and NPR1, but increased the plants' sensitivity to the oomycete pathogen Phytophthora parasitica var. nicotianae Tucker. Importantly, GhMKK5-overexpressing plants displayed markedly elevated expression of reactive oxygen species-related and cell death marker genes, such as NtRbohA and NtCDM, and resulted in hypersensitive response (HR)-like cell death characterized by the accumulation of H(2)O(2). Furthermore, it was demonstrated that GhMKK5 overexpression in plants reduced their tolerance to salt and drought stresses, as determined by statistical analysis of seed germination, root length, leaf water loss, and survival rate. Drought obviously accelerated the cell death phenomenon in GhMKK5-overexpressing plants. These results suggest that GhMKK5 may play an important role in pathogen infection and the regulation of the salt and drought stress responses in plants.     

Characterization of a stress-responsive ankyrin repeat-containing zinc finger protein of Capsicum annuum (CaKR1)

We isolated many genes induced from pepper cDNA microarray data following their infection with the soybean pustule pathogen Xanthomonas axonopodis pv. glycines 8ra. A full-length cDNA clone of the Capsicum annuum ankyrin-repeat domain C(3)H(1) zinc finger protein (CaKR1) was identified in a chili pepper using the expressed sequence tag (EST) database. The deduced amino acid sequence of CaKR1 showed a significant sequence similarity (46%) to the ankyrin-repeat protein in very diverse family of proteins of Arabidopsis. The gene was induced in response to various biotic and abiotic stresses in the pepper leaves, as well as by an incompatible pathogen, such as salicylic acid (SA) and ethephon. CaKR1 expression was highest in the root and flower, and its expression was induced by treatment with agents such as NaCl and methyl viologen, as well as by cold stresses. These results showed that CaKR1 fusion with soluble, modified green fluorescent protein (smGFP) was localized to the cytosol in Arabidopsis protoplasts, suggesting that CaKR1 might be involved in responses to both biotic and abiotic stresses in pepper plants.     

番茄低温响应转录因子SlNAC41克隆及表达分析

NAC转录因子在调控植物生长发育、生物及非生物逆境应答中发挥着重要作用。前期,我们通过对番茄幼苗在低温胁迫下的基因表达谱进行分析,发现Unigene SGN-U212711受低温诱导表达强烈。本研究从番茄中克隆了该基因,命名为Sl NAC41,其开放阅读框（ORF）1 173 bp,编码390个氨基酸,蛋白N端具有典型的NAM结构域,属于NAC转录因子家族成员。预测Sl NAC41蛋白分子量为43.5 k Da,等电点为5.2。实时荧光定量PCR分析表明,Sl NAC41在番茄各组织均有表达,在花中的表达量最高,在红熟果中的表达量最低。低温、干旱、高盐、甲基紫精（MV）、脱落酸（ABA）及乙烯利（ETH）处理均能诱导该基因的表达,其中,以低温和干旱诱导表达最为强烈。利用PLACE和Plant CARE对启动子序列进行预测分析发现,Sl NAC41启动子区含有大量响应光、病原菌侵染、激素、低温、脱水及盐胁迫的顺式作用元件。这些结果表明,Sl NAC41可能在番茄生物及非生物胁迫应答中发挥重要调控作用。     

Identification and functional roles of CaDIN1 in abscisic acid signaling and drought sensitivity

Plants frequently face challenges caused by various abiotic stresses, including drought, and have evolved defense mechanisms to counteract the deleterious effects of these stresses. The phytohormone abscisic acid (ABA) is involved in signal transduction pathways that mediate defense responses of plants to abiotic stress. Here, we report a new function of the CaDIN1 protein in defense responses to abiotic stress. The CaDIN1 gene was strongly induced in pepper leaves exposed to ABA, NaCl, and drought stresses. CaDIN1 proteins share high sequence homology with other known DIN1 proteins and are localized in chloroplasts. We generated CaDIN1-silenced peppers and overexpressing transgenic Arabidopsis plants and evaluated their response to ABA and drought stress. Virus-induced gene silencing of CaDIN1 in pepper plants conferred enhanced tolerance to drought stress, which was accompanied by low levels of lipid peroxidation in dehydrated leaves. CaDIN1-overexpressing transgenic plants exhibited reduced sensitivity to ABA during seed germination and seedling stages. Transgenic plants were more vulnerable to drought than that by the wild-type plants because of decreased expression of ABA responsive stress-related genes and reduced stomatal closure in response to ABA. Together, these results suggest that CaDIN1 modulates drought sensitivity through ABA-mediated cell signaling.     

CASAR82A, a Pathogen-induced Pepper SAR8.2, Exhibits an Antifungal Activity and its Overexpression Enhances Disease Resistance and Stress Tolerance

Pepper SAR8.2 gene (CASAR82A) was previously reported to be locally or systemically induced in pepper plants by biotic and abiotic stresses. In this study, the physiological and molecular functions of the pepper SAR8.2 protein in the plant defense responses were investigated by generating Arabidopsis transgenic lines overexpressing the CASAR82A gene. The transgenic Arabidopsis plants grew faster than the wild-type plants, indicating that the CASAR82A gene was involved in plant development. The ectopic expression of CASAR82A in Arabidopsis was accompanied by the expression of the Arabidopsis pathogenesis-related (PR)-genes including AtPR-1, AtPR-4 and AtPR-5. CASAR82A overexpression enhanced the resistance against infections by Pseudomonas syringae pv. tomato, Fusarium oxysporum f.sp. matthiolae or Botrytis cinerea. The transgenic plants also exhibited increased NaCl and drought tolerance during all growth stages. Moreover, the methyl viologen test showed that the transgenic plants were tolerant to oxidative stress. The purified recombinant CASAR82A protein and crude protein extracts of the transgenic plants exhibited antifungal activity against some phytopathogenic fungi, indicating that the enhanced resistance of the transgenic plants to fungal pathogen infection may be due to the antifungal effect of SAR8.2 protein.     

Plant stress surveillance monitored by ABA and disease signaling interactions

Abiotic and biotic stresses are the major factors that negatively impact plant growth. In response to abiotic environmental stresses such as drought, plants generate resistance responses through abscisic acid (ABA) signal transduction. In addition to the major role of ABA in abiotic stress signaling, ABA signaling was reported to downregulate biotic stress signaling. Conversely recent findings provide evidence that initial activation of plant immune signaling inhibits subsequent ABA signal transduction. Stimulation of effector-triggered disease response can interfere with ABA signal transduction via modulation of internal calcium-dependent signaling pathways. This review overviews the interactions of abiotic and biotic stress signal transduction and the mechanism through which stress surveillance system operates to generate the most efficient resistant traits against various stress condition.