Structure diversity of nitric oxide synthases (NOS): the emergence of new forms in photosynthetic organisms

Humans have enormously increased the level of nitrogen (N) circulating in the troposphere and the earth surface during the last century, correlating with the population increase. As an undesirable consequence, high levels of reactive N are polluting the environment where humans inhabit. Nitric oxide (NO) is one of the reactive N species with both positive and negative impact on life. NO synthases (NOS) are enzymes that oxidize arginine to citrulline and generate the denitrifying intermediate NO which can be subsequently reduced to N2O and N2. NOS are large modular enzymes present in all kingdoms which through evolution were the result of multiple gene and genome duplication events together with changes in protein architecture (Andreakis et al. 2011). A recently described NOS from the marine unicellular microalgae Ostreococcus tauri, belonging to the picoplankton in oceans, adds new insights to study the evolution of the complex organization of these enzymes. In this opinion we discuss the structure diversity of the emerging new NOS forms described in prokaryotes and eukaryotes. Regarding the controversy about the existence of canonical NOS in higher plants, we propose that the latest findings support the existence of a high diversity of NOS forms in different lineages. Thereby, since higher plant species whose genomes have been fully sequenced, which are scarce, it cannot be discarded that a new form of NOS may have evolved in higher plants.Fil: Correa Aragunde, Maria Natalia. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Mar del Plata. Instituto de Investigaciones Biológicas; Argentina. Universidad Nacional de Mar del Plata; ArgentinaFil: Foresi, Noelia Pamela. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Mar del Plata. Instituto de Investigaciones Biológicas; Argentina. Universidad Nacional de Mar del Plata; ArgentinaFil: Lamattina, Lorenzo. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Mar del Plata. Instituto de Investigaciones Biológicas; Argentina. Universidad Nacional de Mar del Plata; Argentin

A singular nitric oxide synthase with a globin domain found in Synechococcus PCC 7335 mobilizes N from arginine to nitrate

The enzyme nitric oxide synthase (NOS) oxidizes L-arginine to NO and citrulline. In this work, we characterise the NOS from the cyanobacteria Synechococcus PCC 7335 (SyNOS). SyNOS possesses a canonical mammalian NOS architecture consisting of oxygenase and reductase domains. In addition, SyNOS possesses an unusual globin domain at the N-terminus. Recombinant SyNOS expressed in bacteria is active, and its activity is suppressed by the NOS inhibitor L-NAME. SyNOS allows E. coli to grow in minimum media containing L-arginine as the sole N source, and has a higher growth rate during N deficiency. SyNOS is expressed in Synechococcus PCC 7335 where NO generation is dependent on L-arginine concentration. The growth of Synechococcus is dramatically inhibited by L-NAME, suggesting that SyNOS is essential for this cyanobacterium. Addition of arginine in Synechococcus increases the phycoerythrin content, an N reservoir. The role of the novel globin domain in SyNOS is discussed as an evolutionary advantage, conferring new functional capabilities for N metabolism.Fil: Correa Aragunde, Maria Natalia. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Biológicas. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Instituto de Investigaciones Biológicas; Argentina. Universidad Nacional de Mar del Plata; ArgentinaFil: Foresi, Noelia Pamela. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Biológicas. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Instituto de Investigaciones Biológicas; Argentina. Universidad Nacional de Mar del Plata; ArgentinaFil: del Castello, Fiorella Paola. Universidad Nacional de Mar del Plata; ArgentinaFil: Lamattina, Lorenzo. Universidad Nacional de Mar del Plata; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Biológicas. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Instituto de Investigaciones Biológicas; Argentin

Chimera of Globin/Nitric Oxide Synthase: Toward Improving Nitric Oxide Homeostasis and Nitrogen Recycling and Availability

As a result of anthropogenic activities, there are regions suffering extreme climate changes (hot temperatures, droughts, floods), that generate serious and harmful environmental and socio-economic consequences (Hoegh-Guldberg et al., 2018). Climate change has a strong impact on agriculture, mainly by increasing soil degradation and reducing land productivity (Olsson et al., 2019). Desertification decreases soil macronutrients as organic carbon (OC), phosphorus (P) and nitrogen (N) (Shang et al., 2013; Tang et al., 2015). Considerable reductions of OC, P and N are caused by a decline in soil water, vegetation and wind erosion. It is estimated that when soil water is less than 30%, OC and N decrease approximately 50% (Shang et al., 2013). The intensity and frequency of extreme climate events predicted will increase the competition for nutrients, notably N among plants and soil microorganisms (Bennett and Klironomos, 2019; Pugnaire et al., 2019).N availability is essential for net primary production and determines changes in total vegetation biomass and soil OC (Tharammal et al., 2019). Approximately 150 Tg/yr of N is spilled to the land surface as a result of industrial activities and fossil fuel combustion (Schlesinger, 2009). Huge amounts of N fertilizers are used to increase crop productivity, but only 25%?30% are retained in plant biomass (Nadelhoffer et al., 1999; Schlesinger, 2009). The excess of N is then transported to aquatic environments resulting in eutrophication and reduction of dissolved O2, percolated to the groundwater or loss to the atmosphere, increasing greenhouse gas (GHG) emissions such as nitrogen oxides (NOx) (Schlesinger, 2009; Breitburg et al., 2018). In this context, bio/technological solutions like genetic modification of crops are required to avoid, reduce and reverse GHG emissions and water eutrophication, contributing to climate change mitigation. New biotechnological strategies developed for the agriculture to increase N use efficiency (NUE) in crops would help to fulfill this purpose. In this opinion, we will discuss some studies of nitric oxide (NO) synthases (NOS) and nitrate reductase (NR) enzymes playing a collaborative role with globin (Hb) proteins, leading to NO and/or nitrate (NO3-) homeostasis in different organisms. We propose that chimeric globin-NOS such as the NOS from Synechococcus PCC 7335 (SyNOS) may have evolved in photosynthetic microorganisms contributing to a more efficient N recycling and sustaining growth in N fluctuating conditions. Engineered crops that maximize NUE would result in a better adaptation to changing climatic conditions with less N fertilization, preserving aquatic ecosystems and atmosphere.Fil: del Castello, Fiorella Paola. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Biológicas. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Instituto de Investigaciones Biológicas; ArgentinaFil: Nejamkin, Andres. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Biológicas. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Instituto de Investigaciones Biológicas; ArgentinaFil: Foresi, Noelia Pamela. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Biológicas. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Instituto de Investigaciones Biológicas; ArgentinaFil: Lamattina, Lorenzo. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Biológicas. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Instituto de Investigaciones Biológicas; ArgentinaFil: Correa Aragunde, Maria Natalia. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Biológicas. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Instituto de Investigaciones Biológicas; Argentin

Do photosynthetic cells communicate with each other during cell death? From cyanobacteria to vascular plants

As in metazoans, life in oxygenic photosynthetic organisms relies on the accurate regulation of cell death. During development and in response to the environment, photosynthetic cells activate and execute cell death pathways that culminate in the death of a specific group of cells, a process known as regulated cell death (RCD). RCD control is instrumental, as its misregulation can lead to growth penalties and even the death of the entire organism. Intracellular molecules released during cell demise may act as ‘survival’ or ‘death’ signals and control the propagation of cell death to surrounding cells, even in unicellular organisms. This review explores different signals involved in cell-cell communication and systemic signalling in photosynthetic organisms, in particular Ca2+, reactive oxygen species, lipid derivates, nitric oxide, and eATP. We discuss their possible mode-of-action as either ‘survival’ or ‘death’ molecules and their potential role in determining cell fate in neighbouring cells. By comparing the knowledge available across the taxonomic spectrum of this coherent phylogenetic group, from cyanobacteria to vascular plants, we aim at contributing to the identification of conserved mechanisms that control cell death propagation in oxygenic photosynthetic organisms.Fil: Aguilera, Anabella. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Distefano, Ayelen Mariana. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Biológicas. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Instituto de Investigaciones Biológicas; ArgentinaFil: Jauzein, Cécile. No especifíca;Fil: Correa Aragunde, Maria Natalia. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Biológicas. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Instituto de Investigaciones Biológicas; ArgentinaFil: Martinez, Dana Ethel. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Fisiología Vegetal. Universidad Nacional de La Plata. Facultad de Ciencias Naturales y Museo. Instituto de Fisiología Vegetal; ArgentinaFil: Martin, María Victoria. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones en Biodiversidad y Biotecnología; ArgentinaFil: Sueldo, Daniela Jorgelina. Norwegian University of Science and Technology; Norueg

Nitrogen Depletion Blocks Growth Stimulation Driven by the Expression of Nitric Oxide Synthase in Tobacco

Nitric oxide (NO) is a messenger molecule widespread studied in plant physiology. Latter evidence supports the lack of a NO-producing system involving a NO synthase (NOS) activity in higher plants. However, a NOS gene from the unicellular marine alga Ostreococcus tauri (OtNOS) was characterized in recent years. OtNOS is a genuine NOS, with similar spectroscopic fingerprints to mammalian NOSs and high NO producing capacity. We are interested in investigating whether OtNOS activity alters nitrogen metabolism and nitrogen availability, thus improving growth promotion conditions in tobacco. Tobacco plants were transformed with OtNOS under the constitutive CaMV 35S promoter. Transgenic tobacco plants expressing OtNOS accumulated higher NO levels compared to siblings transformed with the empty vector, and displayed accelerated growth in different media containing sufficient nitrogen availability. Under conditions of nitrogen scarcity, the growth promoting effect of the OtNOS expression is diluted in terms of total leaf area, protein content and seed production. It is proposed that OtNOS might possess a plant growth promoting effect through facilitating N remobilization and nitrate assimilation with potential to improve crop plants performance.Fil: Nejamkin, Andres. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Biológicas. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Instituto de Investigaciones Biológicas; ArgentinaFil: Foresi, Noelia Pamela. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Biológicas. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Instituto de Investigaciones Biológicas; ArgentinaFil: Mayta, Martín Leonardo. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario. Instituto de Biología Molecular y Celular de Rosario. Universidad Nacional de Rosario. Facultad de Ciencias Bioquímicas y Farmacéuticas. Instituto de Biología Molecular y Celular de Rosario; ArgentinaFil: Lodeyro, Anabella Fernanda. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario. Instituto de Biología Molecular y Celular de Rosario. Universidad Nacional de Rosario. Facultad de Ciencias Bioquímicas y Farmacéuticas. Instituto de Biología Molecular y Celular de Rosario; ArgentinaFil: del Castello, Fiorella Paola. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Biológicas. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Instituto de Investigaciones Biológicas; ArgentinaFil: Correa Aragunde, Maria Natalia. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Biológicas. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Instituto de Investigaciones Biológicas; ArgentinaFil: Carrillo, Nestor Jose. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario. Instituto de Biología Molecular y Celular de Rosario. Universidad Nacional de Rosario. Facultad de Ciencias Bioquímicas y Farmacéuticas. Instituto de Biología Molecular y Celular de Rosario; ArgentinaFil: Lamattina, Lorenzo. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Biológicas. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Instituto de Investigaciones Biológicas; Argentin

Regulation of SCF TIR1/AFB E3 ligase assembly by S-nitrosylation of Arabidopsis SKP1-like1 impacts on auxin signaling

The F-box proteins (FBPs) TIR1/AFBs are the substrate recognition subunits of SKP1–cullin–F-box (SCF) ubiquitin ligase complexes and together with Aux/IAAs form the auxin co-receptor. Although tremendous knowledge on auxin perception and signaling has been gained in the last years, SCFTIR1/AFBs complex assembly and stabilization are emerging as new layers of regulation. Here, we investigated how nitric oxide (NO), through S-nitrosylation of ASK1 is involved in SCFTIR1/AFBs assembly. We demonstrate that ASK1 is S-nitrosylated and S-glutathionylated in cysteine (Cys) 37 and Cys118 residues in vitro. Both, in vitro and in vivo protein-protein interaction assays show that NO enhances ASK1 binding to CUL1 and TIR1/AFB2, required for SCFTIR1/AFB2 assembly. In addition, we demonstrate that Cys37 and Cys118 are essential residues for proper activation of auxin signaling pathway in planta. Phylogenetic analysis revealed that Cys37 residue is only conserved in SKP proteins in Angiosperms, suggesting that S-nitrosylation on Cys37 could represent an evolutionary adaption for SKP1 function in flowering plants. Collectively, these findings indicate that multiple events of redox modifications might be part of a fine-tuning regulation of SCFTIR1/AFBs for proper auxin signal transduction.This work has been supported by grants from CONICET (PIP 0202 to M.C.T), Universidad Nacional de Mar del Plata and Agencia Nacional de Promoción Científica y Tecnológica, -Argentina (PICT 1167 to M.C.T and PICT 2421 to C.A.C); from the Spanish Government (PS09/00101 and PI12/00875 to A.M.R., cofinanced by the European Union ERDF), and by the Argentinian-Spanish Integrated Action ES/11/02 / PRIAIBAR- 2011-0782. A.M.R. is supported by the I3SNS programme (ISCIII, Spanish Government). The Proteomics Service of the CBMSO is a member of Proteored (PRB2-ISCIII), and is supported by grants PT13/ 0001/0024 and PT17/0019/0018 of Spanish Government (cofinanced by the European Union ERDF). Work in the laboratory of M.E is supported by grants from the National Institutes of Health (NIH) GM43644, The Howard Hughes Medical Institute and the Gordon and Betty Moore Foundation, US

Nitric oxide modulates the expression of cell cycle regulatory genes during lateral root formation in tomato

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Nitric oxide is required for the auxin-induced activation of NADPH-dependent thioredoxin reductase and protein denitrosylation during root growth responses in arabidopsis

Background and Aims: Auxin is the main phytohormone controlling root development in plants. This study uses pharmacological and genetic approaches to examine the role of auxin and nitric oxide (NO) in the activation of NADPH-dependent thioredoxin reductase (NTR), and the effect that this activity has on root growth responses in Arabidopsis thaliana. Methods: Arabidopsis seedlings were treated with auxin with or without the NTR inhibitors auranofin (ANF) and 1-chloro-2, 4-dinitrobenzene (DNCB). NTR activity, lateral root (LR) formation and S-nitrosothiol content were measured in roots. Protein S-nitrosylation was analysed by the biotin switch method in wild-type arabidopsis and in the double mutant ntra ntrb. Key Results: The auxin-mediated induction of NTR activity is inhibited by the NO scavenger 2-(4-carboxyphenyl)-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide (CPTIO), suggesting that NO is downstream of auxin in this regulatory pathway. The NTR inhibitors ANF and DNCB prevent auxin-mediated activation of NTR and LR formation. Moreover, ANF and DNCB also inhibit auxin-induced DR5 : : GUS and BA3 : : GUS gene expression, suggesting that the auxin signalling pathway is compromised without full NTR activity. Treatment of roots with ANF and DNCB increases total nitrosothiols (SNO) content and protein S-nitrosylation, suggesting a role of the NTR-thioredoxin (Trx)-redox system in protein denitrosylation. In agreement with these results, the level of S-nitrosylated proteins is increased in the arabidopsis double mutant ntra ntrb as compared with the wild-type. Conclusions: The results support for the idea that NTR is involved in protein denitrosylation during auxin-mediated root development. The fact that a high NO concentration induces NTR activity suggests that a feedback mechanism to control massive and unregulated protein S-nitrosylation could be operating in plant cells.Fil: Correa Aragunde, Maria Natalia. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Mar del Plata. Instituto de Investigaciones Biológicas; Argentina. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales; ArgentinaFil: Cejudo, Francisco J.. Universidad de Sevilla; EspañaFil: Lamattina, Lorenzo. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Mar del Plata. Instituto de Investigaciones Biológicas; Argentina. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales; Argentin

Auxin and Nitric Oxide: A Counterbalanced Partnership Ensures the Redox Cue Control Required for Determining Root Growth Pattern

Auxin is the main hormone that controls growth and developmental processes in plants. In the last decade, many studies confirmed the interplay between reactive oxygen species (ROS) and reactive nitrogen species (RNS) in the auxin signaling transduction pathway. Nitric oxide (NO) is an RNS induced by, and required for the auxin-mediated lateral root formation and inhibition of the primary root elongation. Auxin induces both ROS and NO that, in turn, modulates reductases and peroxidases activities resulting in the attenuation of auxin signaling. As a consequence, a balance between ROS and NO concentrations appears to be essential for the control of the auxin action during root growth and developmental processes. In this review, we summarize and discuss the recent progress in the understanding of signaling and interacting components participating of the auxin-, NO- and ROS-modulated redox balance determining root growth pattern.Fil: Correa Aragunde, Maria Natalia. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Biológicas. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Instituto de Investigaciones Biológicas; ArgentinaFil: Foresi, Noelia Pamela. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Biológicas. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Instituto de Investigaciones Biológicas; ArgentinaFil: Lamattina, Lorenzo. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Biológicas. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Instituto de Investigaciones Biológicas; Argentin