Visualization of NO3−/NO2− Dynamics in Living Cells by Fluorescence Resonance Energy Transfer (FRET) Imaging Employing a Rhizobial Two-component Regulatory System

Abstract

Ϫ is used as a substrate for respiration and/or assimilation. In animals, including humans, NO 3 Ϫ and NO 2 Ϫ (NO 3 Ϫ /NO 2 Ϫ ) were recognized for a long time as being merely inert oxidants of nitric oxide (NO) (1). NO is a key signaling molecule that regulates a vast range of physiological functions, such as vascular homeostasis, neurotransmission, and host defense (2). Intriguingly, many studies over the last decade revealed that these inert NO 3 Ϫ /NO 2 Ϫ species are physiologically recycled to form NO and other reactive nitrogen species through the "nitrate-nitritenitric oxide (NO 3 Ϫ -NO 2 Ϫ -NO) pathway" (3-5). Currently, NO 3 Ϫ /NO 2 Ϫ are considered as stable reservoirs for NO-like bioactivity, and several beneficial aspects of NO 3 Ϫ /NO 2 Ϫ in the treatment and prevention of cardiovascular diseases by restoring NO homeostasis are reported (5, 6).As the regulation of NO 3 Ϫ /NO 2 Ϫ in physiological processes is an attractive therapeutic target, it is important to understand NO 3 Ϫ /NO 2 Ϫ in biological processes, how intracellular levels are regulated, and how they control cellular processes. The most frequently used method for NO 3 Ϫ /NO 2 Ϫ measurement is based on the Griess reaction (7). The Griess method is also used to assess NO synthesis because of the immediate conversion of NO into NO 3 Ϫ

Regulation of nitrous oxide reductase genes by NasT‐mediated transcription antitermination in Bradyrhizobium diazoefficiens

“…The NasST system is widely distributed in Gram‐negative bacteria (Luque‐Almagro et al ., ), and has been shown to be involved in the nitrate/nitrite ( / )‐responsive regulation of the nas genes, for nitrate assimilation pathway, in Azotobacter vinelandii , Paracoccus denitrificans , Pseudomonas aeruginosa and B. diazoefficiens (Romeo et al ., ; Wang et al ., ; Luque‐Almagro et al ., ; Cabrera et al ., ). NasS and NasT form a complex that dissociates in the presence of certain concentration of nitrate in the micromolar range (Luque‐Almagro et al ., ; Sánchez et al ., ; Hidaka et al ., ). In B. diazoefficiens , nosZ expression in the presence of nitrate is markedly lower in a nasT mutant than in the wild type under both aerobic and anaerobic conditions.…”

Dynamic measurement of cytosolic pH and [NO 3 − ] uncovers the role of the vacuolar transporter AtCLCa in cytosolic pH homeostasis

“…Overall, our results demonstrate that ClopHensor can be used to measure [NO 3 − ] and pH in GCs. Other NO 3 − biosensors have been developed, such as NiTrak, which allows monitoring the activity of the nitrate transporter NRT1.1/NFP5.6 (52), and sNOOOpy, a nitrate/nitrite biosensor that has not been tested in plants yet (53). However, ClopHensor is the first biosensor able to report [NO 3 − ] in the cytosol of plants in parallel with pH.…”

Nitrogen Cycling in Soybean Rhizosphere: Sources and Sinks of Nitrous Oxide (N2O)

“…NasT is an ANTAR (AmiR and NasR transcription antitermination regulator)-family protein (Shu and Zhulin, 2002). NasS and NasT form a complex that dissociates when NasS senses NO 3 – in micromolar concentrations (Luque-Almagro et al, 2013; Sánchez et al, 2014; Hidaka et al, 2016). When NO 3 – is present, nos expression is markedly decreased (∼70%) in the Δ nasT background.…”

“…Then, the application of nos ++ mutants (carrying a mutation in nasS ) may be more effective than that of wild type nos + alone if NO 3 – concentration in the rhizosphere is below the threshold for dissociation of the NasS-NasT complex. Although the concentration threshold in vitro is within the micromolar range (Hidaka et al, 2016), this concentration remains to be fixed under soil conditions.…”