Hydrogen peroxide is involved in nitric oxide‐induced cell death in maize leaves

Recent evidence indicates that nitric oxide (NO) plays an important role in plant hypersensitive cell death. Here, we report that NO treatment led to rapid cell death and induced hydrogen peroxide (H₂O₂) accumulation in maize leaves. We also show that NO induced the expression of Zmrboh genes. Pharmacological study suggests that NO‐induced cell death is in part mediated via H₂O₂. In addition, semi‐quantitative RT‐PCR revealed that NO induced expression of the systemic acquired resistance (SAR) genes, ZmPR1 and ZmPR5.     

<Emphasis Type="Italic">N</Emphasis>-acetyl cysteine suppresses the foam cell formation that is induced by oxidized low density lipoprotein via regulation of gene expression

Foam cells derived from macrophages have been implicated as markers of early stage atherosclerosis development. In this study, we found that N-acetyl cysteine (NAC), a well-known inhibitor of reactive oxygen species (ROS), decreased the generation of ROS and suppressed foam cell formation in the presence of oxidized low density lipoprotein through down-regulation of cluster of differentiation 36 expression. We investigated gene expression profiles in order to determine the effects of NAC on foam cell formation using a microarray analysis. The level of apolipoprotein E, which is involved in lipid efflux, was increased and the levels of the antioxidant genes glutathione peroxidase 1 and 3 were also increased. The expression levels of the oxidative stress response and the DNA repair genes were decreased. These results were confirmed using quantitative real-time PCR. Our results indicate that oxidative stress plays an important role in foam cell formation, and that regulation of oxidation using antioxidants is a potential therapeutic method for blocking atherosclerosis development.     

Bax-induced cell death of Arabidopsisis meditated through reactive oxygen-dependent and -independent processes

An Arabidopsisprotoplast system was developed for dissecting plant cell death in individual cells. Bax, a mammalian pro-apoptotic member of the Bcl-2 family, induces apoptotic-like cell death in Arabidopsis. Bax accumulation in Arabidopsismesophyll protoplasts expressing murine BaxcDNA from a glucocorticoid-inducible promoter results in cytological characteristics of apoptosis, namely DNA fragmentation, increased vacuolation, and loss of plasma membrane integrity. In vivotargeting analysis monitored using jellyfish green fluorescent protein (GFP) reporter indicated full-length Bax was localized to the mitochondria, as it does in animal cells. Deletion of the carboxyl-terminal transmembrane domain of Bax completely abolished targeting to mitochondria. Bax expression was followed by reactive oxygen species (ROS) accumulation. Treatment of protoplasts with the antioxidant N-acetyl- -cysteine (NAC) during induction of Bax expression strongly suppressed Bax-mediated ROS production and the cell death phenotype. However, some population of the ROS depleted cells still induced cell death, indicating that there is a process that Bax-mediated plant cell death is independent of ROS accumulation. Accordingly, suppression of Bax-mediated plant cell death also takes place in two different processes. Over-expression of a key redox-regulator, Arabidopsisnucleoside diphosphate kinase 2 (AtNDPK2) down-regulated ROS accumulation and suppressed Bax-mediated cell death and transient expression of ArabidopsisBax inhibitor-1 (AtBI-1) substantially suppressed Bax-induced cell death without altering cellular ROS level. Taken together, our results collectively suggest that the Bax-mediated cell death and its suppression in plants is mediated by ROS-dependent and -independent processes.     

Reactive oxygen species generated in chloroplasts contribute to tobacco leaf infection by the necrotrophic fungus Botrytis cinerea

Reactive oxygen species (ROS) play fundamental roles in plant responses to pathogen infection, including modulation of cell death processes and defense‐related gene expression. Cell death triggered as part of the hypersensitive response enhances resistance to biotrophic pathogens, but favors the virulence of necrotrophs. Even though the involvement of ROS in the orchestration of defense responses is well established, the relative contribution of specific subcellular ROS sources to plant resistance against microorganisms with different pathogenesis strategies is not completely known. The aim of this work was to investigate the role of chloroplastic ROS in plant defense against a typical necrotrophic fungus, Botrytis cinerea. For this purpose, we used transgenic Nicotiana tabacum (tobacco) lines expressing a plastid‐targeted cyanobacterial flavodoxin (pfld lines), which accumulate lower chloroplastic ROS in response to different stresses. Tissue damage and fungal growth were significantly reduced in infected leaves of pfld plants, as compared with infected wild‐type (WT) counterparts. ROS build‐up triggered by Botrytis infection and associated with chloroplasts was significantly decreased (70–80%) in pfld leaves relative to the wild type. Phytoalexin accumulation and expression of pathogenesis‐related genes were induced to a lower degree in pfld plants than in WT siblings. The impact of fungal infection on photosynthetic activity was also lower in pfld leaves. The results indicate that chloroplast‐generated ROS play a major role in lesion development during Botrytis infection. This work demonstrates that the modulation of chloroplastic ROS levels by the expression of a heterologous antioxidant protein can provide a significant degree of protection against a canonical necrotrophic fungus.     

Cardamonin induces G2/M arrest and apoptosis via activation of the JNK–FOXO3a pathway in breast cancer cells

Cardamonin (CD), a naturally occurring chalcone isolated from large black cardamom, was previously reported to suppress the proliferation of breast cancer cells. However, its precise molecular anti‐tumor mechanisms have not been well elucidated. In this study, we found that CD markedly inhibited the proliferation of MDA‐MB 231 and MCF‐7 breast cancer cells through the induction of G2/M arrest and apoptosis. Reactive oxygen species (ROS) plays a pivotal role in the inhibition of CD‐induced cell proliferation. Treatment with N‐acetyl‐cysteine (NAC), an ROS scavenger, blocked CD‐induced G2/M arrest and apoptosis in this study. Quenching of ROS by overexpression of catalase also blocked CD‐induced cell cycle arrest and apoptosis. We showed that CD enhanced the expression and nuclear translocation of Forkhead box O3 (FOXO3a) via upstream c‐Jun N‐terminal kinase, inducing the expression of FOXO3a and its target genes, including p21, p27, and Bim. This process led to the reduction of cyclin D1 and enhancement of activated caspase‐3 expression. The addition of NAC markedly reversed these effects, knockdown of FOXO3a using small interfering RNA also decreased CD‐induced G2/M arrest and apoptosis. In vivo, CD efficiently suppressed the growth of MDA‐MB 231 breast cancer xenograft tumors. Taken together, our data provide a molecular mechanistic rationale for CD‐induced cell cycle arrest and apoptosis in breast cancer cells.     

Dickeya dadantii pectic enzymes necessary for virulence are also responsible for activation of the Arabidopsis thaliana innate immune system

Soft‐rot diseases of plants attributed to Dickeya dadantii result from lysis of the plant cell wall caused by pectic enzymes released by the bacterial cell by a type II secretion system (T2SS). Arabidopsis thaliana can express several lines of defence against this bacterium. We employed bacterial mutants with defective envelope structures or secreted proteins to examine early plant defence reactions. We focused on the production of AtrbohD‐dependent reactive oxygen species (ROS), callose deposition and cell death as indicators of these reactions. We observed a significant reduction in ROS and callose formation with a bacterial mutant in which genes encoding five pectate lyases (Pels) were disrupted. Treatment of plant leaves with bacterial culture filtrates containing Pels resulted in ROS and callose production, and both reactions were dependent on a functional AtrbohD gene. ROS and callose were produced in response to treatment with a cellular fraction of a T2SS‐negative mutant grown in a Pels‐inducing medium. Finally, ROS and callose were produced in leaves treated with purified Pels that had also been shown to induce the expression of jasmonic acid‐dependent defence genes. Pel catalytic activity is required for the induction of ROS accumulation. In contrast, cell death observed in leaves infected with the wild‐type strain appeared to be independent of a functional AtrbohD gene. It was also independent of the bacterial production of pectic enzymes and the type III secretion system (T3SS). In conclusion, the work presented here shows that D. dadantii is recognized by the A. thaliana innate immune system through the action of pectic enzymes secreted by bacteria at the site of infection. This recognition leads to AtrbohD‐dependent ROS and callose accumulation, but not cell death.