Iron is essential for all organisms but can be toxic in excess. Iron homeostasis is typically regulated by cytoplasmic iron binding proteins, but here we describe a signal transduction system (PmrA/PmrB) that responds to extracytoplasmic ferric iron. Iron promoted transcription of PmrA-activated genes and resistance to the antibiotic polymyxin in Salmonella. The PmrB protein bound iron via its periplasmic domain which harbors two copies of the sequence ExxE, a motif present in the Saccharomyces FTR1 iron transporter and in mammalian ferritin light chain. A pmrA mutant was hypersensitive to killing by iron but displayed wild-type resistance to a variety of oxidants, suggesting PmrA/PmrB controls a novel pathway mediating the avoidance of iron toxicity.

The rates of RNA decay and transcription determine the steady state levels of all mRNAs and both can be subject to regulation. While the details of transcriptional regulation are becoming increasingly understood, the mechanism(s) controlling mRNA decay remain unclear. In yeast, a major pathway of mRNA decay begins with deadenylation followed by decapping and 5’-3’ exonuclease digestion. Importantly, it is hypothesized that ribosomes must be removed from mRNA before transcripts are destroyed. Contrary to this prediction, here we show that decay takes place while mRNAs are associated with actively translating ribosomes. The data indicate that dissociation of ribosomes from mRNA is not a prerequisite for decay and we suggest that the 5’-3’ polarity of mRNA degradation has evolved to ensure that the last translocating ribosome can complete translation.

Transgenic tobacco deficient in the H 2 O 2 -removing enzyme catalase (Cat1AS) was used as an inducible and noninvasive system to study the role of H 2 O 2 as an activator of pathogenesis-related (PR) proteins in plants. Excess H 2 O 2 in Cat1AS plants was generated by simply increasing light intensities. Sustained exposure of Cat1AS plants to excess H 2 O 2 provoked tissue damage, stimulated salicylic acid and ethylene production, and induced the expression of acidic and basic PR proteins with a timing and magnitude similar to the hypersensitive response against pathogens. Salicylic acid production was biphasic, and the first peak of salicylic acid as well as the peak of ethylene occurred within the first hours of high light, which is long before the development of tissue necrosis. Under these conditions, accumulation of acidic PR proteins was also seen in upper leaves that were not exposed to high light, indicating systemic induction of expression. Short exposure of Cat1AS plants to excess H 2 O 2 did not cause damage, induced local expression of acidic and basic PR proteins, and enhanced pathogen tolerance. However, the timing and magnitude of PR protein induction was in this case more similar to that in upper uninfected leaves than to that in hypersensitiveresponse leaves of pathogen-infected plants. Together, these data demonstrate that sublethal levels of H 2 O 2 activate expression of acidic and basic PR proteins and lead to enhanced pathogen tolerance. However, rapid and strong activation of PR protein expression, as seen during the hypersensitive response, occurs only when excess H 2 O 2 is accompanied by leaf necrosis.

We initially compared lipid peroxidation profiles in tobacco (Nicotiana tabacum) leaves during different cell death events. An upstream oxylipin assay was used to discriminate reactive oxygen species (ROS)-mediated lipid peroxidation from 9-and 13-lipoxygenase (LOX)-dependent lipid peroxidation. Free radical-mediated membrane peroxidation was measured during H 2 O 2 -dependent cell death in leaves of catalase-deficient plants. Taking advantage of these transgenic plants, we demonstrate that, under light conditions, H 2 O 2 plays an essential role in the execution of cell death triggered by an elicitor, cryptogein, which provokes a similar ROS-mediated lipid peroxidation. Under dark conditions, however, cell death induction by cryptogein was independent of H 2 O 2 and accompanied by products of the 9-LOX pathway. In the hypersensitive response induced by the avirulent pathogen Pseudomonas syringae pv syringae, both 9-LOX and oxidative processes operated concurrently, with ROS-mediated lipid peroxidation prevailing in the light. Our results demonstrate, therefore, the tight interplay between H 2 O 2 and lipid hydroperoxides and underscore the importance of light during the hypersensitive response.Different defense mechanisms are used by plants to cope with pathogen assaults. A major source of resistance is conditioned by the interaction between plant resistance and pathogen avirulence gene products (Martin et al., 2003;Rathjen and Moffett, 2003). This defense strategy is characterized by (1) the activation of the hypersensitive response (HR), typified by a localized programmed cell death activation; (2) the initiation of defense responses, including cell wall reinforcement, accumulation of phytoalexins, and expression of antimicrobial proteins; and (3) the onset of a local and systemic acquired resistance (Lamb and Dixon, 1997;Beers and McDowell, 2001;Greenberg and Yao, 2004).The signaling cascades leading to the HR are starting to be elucidated. Several resistance genes have been cloned, and protein kinases, phosphatases, and GTP-binding proteins have all been implicated downstream of these recognition proteins (Martin et al., 2003;Rathjen and Moffett, 2003). Changes in ion fluxes across the plasma membrane and the production of reactive oxygen species (ROS) and nitric oxide (NO) are among the earliest events following pathogen infection or elicitor treatment in cultured plant cells (Doke, 1997;Grant and Loake, 2000;Wendehenne et al., 2004). The production of ROS is biphasic, with a sustained second oxidative burst in response to an avirulent pathogen, and monophasic and transient with a virulent pathogen. This suggests that ROS production may be responsible for some of the defense-associated processes (Lamb and Dixon, 1997). Furthermore, ROS, and specifically H 2 O 2 , are key modulators of NO in triggering plant cell death (Delledonne et al., 2001;Neill et al., 2002;Wendehenne et al., 2004).In order to obtain further insight into the role of H 2 O 2 in plant signaling and cell death, we used transgenic tobacco ...

Transgenic tobacco deficient in either Cat1 (Cat1AS), Cat2 (Cat2AS), or both (CatGH) was generated through sense and antisense technology. Cat1AS, Cat2AS, and CatGH plants showed no visible phenotype when grown at low light (100 mu mol m(-2) sec(-1)). Under these conditions, deficiency in Cat1 and/or Cat2 did not lead to constitutive pathogenesis-related (PR-1) expression and did not potentiate PR-1 induction by exogenous salicylic acid. This demonstrates that catalase suppression per se is not a sufficient signal for PR-1 induction. In Cat1-deficient plants exposed to higher light intensities (250-1000 mu mol m(-2) sec(-1)), PR-1 expression was induced without pathogenic challenge and multiplication of Pseudomonas syringae pv, syringae was repressed. Yet, it is unlikely that Cat1 deficiency is mimicking the mode of action of salicylic acid in tobacco, because, concurrent with PR-1 induction, Cat1 deficiency at high light provoked severe leaf damage, characterized by white necrotic lesions. Taken together, these results do not support the model that catalase inactivation is the key route by which salicylic acid induces PR defense responses in healthy tissue. However, because catalase deficiency is potentially lethal to leaves, catalase inactivation by salicylic acid could be of importance in the establishment of hypersensitive responses

We have isolated a new organic hydroperoxide resistance (ohr) gene from Xanthomonas campestris pv. phaseoli. This was done by complementation of anEscherichia coli alkyl hydroperoxide reductase mutant with an organic hydroperoxide-hypersensitive phenotype. ohrencodes a 14.5-kDa protein. Its amino acid sequence shows high homology with several proteins of unknown function. An ohr mutant was subsequently constructed, and it showed increased sensitivity to both growth-inhibitory and killing concentrations of organic hydroperoxides but not to either H2O2 or superoxide generators. No alterations in sensitivity to other oxidants or stresses were observed in the mutant. ohr had interesting expression patterns in response to low concentrations of oxidants. It was highly induced by organic hydroperoxides, weakly induced by H2O2, and not induced at all by a superoxide generator. The novel regulation pattern of ohrsuggests the existence of a second organic hydroperoxide-inducible system that differs from the global peroxide regulator system, OxyR. Expression of ohr in various bacteria tested conferred increased resistance totert-butyl hydroperoxide killing, but this was not so for wild-type Xanthomonas strains. The organic hydroperoxide hypersensitivity of ohr mutants could be fully complemented by expression of ohr or a combination of ahpC and ahpF and could be partially complemented by expression ahpC alone. The data suggested that Ohr was a new type of organic hydroperoxide detoxification protein.

No abstract

Catalase is an important protective enzyme against H 2 O 2 toxicity. Here, we report the characterization of a Xanthomonas oryzae pv. oryzae catalase gene (katX). The gene was localized and its nucleotide sequence was determined. The gene codes for a 77-kDa polypeptide. The deduced katX amino acid sequence shares regions of high identity with other monofunctional catalases in a range of organisms from bacteria to eukaryotes. The transcriptional regulation of katX was atypical of bacterial monofunctional kat genes. Northern (RNA) analysis showed that katX transcription was highly induced by treatments with low concentrations of menadione, a superoxide generator, and methyl methanesulfonate, a mutagen. It was only weakly induced by H 2 O 2 . Unlike in other bacteria, a high level of catalase in Xanthomonas spp. provided protection from the growth-inhibitory and killing effects of H 2 O 2 but not from those of organic peroxides and superoxide generators. Unexpectedly, heterologous expression of katX in Escherichia coli was both growth phase and temperature dependent. Catalase activity in E. coli kat mutants harboring katX on an expression vector was detectable only when the cells entered the stationary phase of growth and at 28؇C. The patterns of transcription regulation, heterologous expression, and physiological function of katX are different from previously studied bacterial kat genes.Catalase is a heme-containing enzyme involved in dismutation of H 2 O 2 to oxygen and water. The enzyme plays an important role in detoxifying H 2 O 2 and minimizing oxidative stress caused by highly reactive oxygen species (ROS, i.e., OH⅐) which arise from H 2 O 2 degradation in the Fenton reaction (13). Mutations in kat have always resulted in increased sensitivity to H 2 O 2 stress, and this is an indication of the important physiological role of the enzyme (10, 13). In many bacteria, there are two types of catalase enzyme, namely a monofunctional catalase and a bifunctional catalase/peroxidase. Each enzyme is encoded by a different gene (e.g., in Escherichia coli, katE and katG code for monofunctional and bifunctional catalases, respectively) (35, 45). Monofunctional catalases share regions of an amino acid sequence that is highly conserved among microbial, plant, and mammalian enzymes (5,26,47). In many bacteria, the two kat genes are regulated differently in terms of growth phase and response to oxidative stress, suggesting that they may have different physiological functions (10,23,29).Xanthomonas oryzae pv. oryzae (Xoo) is the causative agent for the most destructive bacterial disease (the bacterial leaf blight) in rice (37). Oxidative stress is an important component of normal aerobic life and in bacterial-plant interactions. Increasing production of ROS, including superoxide, H 2 O 2 , and OH⅐, is associated with an active plant defense response and with aerobic respiration (44). ROS are highly toxic to all cellular components, and their rapid detoxification is essential for microbial survival.Xoo monofunctional catalase ...