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Searching for nitroxyl modulators in Arabidopsis thaliana: a new paradigm of redox signaling in plants
Abstract Through extensive research, nitroxyl (HNO) has emerged as a newly recognized redox signal in plant developmental and stress responses. The interplay between nitric oxide (●NO) and HNO entails a complex network of signaling molecules and regulatory elements sensitive to the environment-specific redox conditions. However, functional implications for HNO in cell signaling require more detailed studies, starting with identifying HNO-level switches. To obtain insight into possible physiologically relevant HNO modulators, we used real-time detection to examine HNO/●NO production triggered by selected plant-related compounds (PRCs), including non-protein amino acids, antioxidants, and phytohormones, both in vitro and in vivo in the model plant Arabidopsis thaliana. Hydrogen sulfide, ascorbic acid, and salicylic acid were identified as superior PRCs in driving HNO/●NO interconversion in the cellular medium, so that these PRCs could provide ubiquitous bioavailability of HNO in plants. Meanwhile, resistance-inducing compounds tended to down-regulate HNO in Arabidopsis leaves. The present study indicates that non-enzymatic HNO/●NO interconversion mediated by functionally important PRCs constitutes a significant route for controlling endogenous HNO concentrations, providing ubiquitous HNO bioavailability in plant cells. Moreover, concurrent HNO/●NO monitoring shows that the redox signals are highly integrated and create a redox code that can be translated into a specific cellular response.
Dynamics of nitration during dark-induced leaf senescence in Arabidopsis reveals proteins modified by tryptophan nitration
Abstract Nitric oxide (NO) is a critical molecule that links plant development with stress responses. Herein, new insights into the role of NO metabolism during leaf senescence in Arabidopsis are presented. A gradual decrease in NO emission accompanied dark-induced leaf senescence (DILS), and a transient wave of peroxynitrite (ONOO–) formation was detected by day 3 of DILS. The boosted ONOO– did not promote tryptophan (Trp) nitration, while the pool of 6-nitroTrp-containing proteins was depleted as senescence progressed. Immunoprecipitation combined with mass spectrometry was used to identify 63 and 4 characteristic 6-nitroTrp-containing proteins in control and individually darkened leaves, respectively. The potential in vivo targets of Trp nitration were mainly related to protein biosynthesis and carbohydrate metabolism. In contrast, nitration of tyrosine-containing proteins was intensified 2-fold on day 3 of DILS. Also, nitrative modification of RNA and DNA increased significantly on days 3 and 7 of DILS, respectively. Taken together, ONOO– can be considered a novel pro-senescence regulator that fine-tunes the redox environment for selective bio-target nitration. Thus, DILS-triggered nitrative changes at RNA and protein levels promote developmental shifts during the plant’s lifespan and temporal adjustment in plant metabolism under suboptimal environmental conditions.
Insights into the expression of DNA (de)methylation genes responsive to nitric oxide signaling in potato resistance to late blight disease
Our previous study concerning the pathogen-induced biphasic pattern of nitric oxide (NO) burst revealed that the decline phase and a low level of NO, due to S-nitrosoglutathione reductase (GSNOR) activity, might be decisive in the upregulation of stress-sensitive genes via histone H3/H4 methylation in potato leaves inoculated with avr P. infestans. The present study refers to the NO-related impact on genes regulating DNA (de)methylation, being in dialog with histone methylation. The excessive amounts of NO after the pathogen or GSNO treatment forced the transient upregulation of histone SUVH4 methylation and DNA hypermethylation. Then the diminished NO bioavailability reduced the SUVH4-mediated suppressive H3K9me2 mark on the R3a gene promoter and enhanced its transcription. However, we found that the R3a gene is likely to be controlled by the RdDM methylation pathway. The data revealed the time-dependent downregulation of the DCL3, AGO4, and miR482e genes, exerting upregulation of the targeted R3a gene correlated with ROS1 overexpression. Based on these results, we postulate that the biphasic waves of NO burst in response to the pathogen appear crucial in establishing potato resistance to late blight through the RdDM pathway controlling R gene expression.
Nitric Oxide Implication in Potato Immunity to Phytophthora infestans via Modifications of Histone H3/H4 Methylation Patterns on Defense Genes
Nitric oxide (NO) is an essential redox-signaling molecule operating in many physiological and pathophysiological processes. However, evidence on putative NO engagement in plant immunity by affecting defense gene expressions, including histone modifications, is poorly recognized. Exploring the effect of biphasic NO generation regulated by S-nitrosoglutathione reductase (GNSOR) activity after avr Phytophthora infestans inoculation, we showed that the phase of NO decline at 6 h post-inoculation (hpi) was correlated with the rise of defense gene expressions enriched in the TrxG-mediated H3K4me3 active mark in their promoter regions. Here, we report that arginine methyltransferase PRMT5 catalyzing histone H4R3 symmetric dimethylation (H4R3sme2) is necessary to ensure potato resistance to avr P. infestans. Both the pathogen and S-nitrosoglutathione (GSNO) altered the methylation status of H4R3sme2 by transient reduction in the repressive mark in the promoter of defense genes, R3a and HSR203J (a resistance marker), thereby elevating their transcription. In turn, the PRMT5-selective inhibitor repressed R3a expression and attenuated the hypersensitive response to the pathogen. In conclusion, we postulate that lowering the NO level (at 6 hpi) might be decisive for facilitating the pathogen-induced upregulation of stress genes via histone lysine methylation and PRMT5 controlling potato immunity to late blight.