Role of nitric oxide (NO) in plant response to cadmium and Fusarium oxysporumpossible crosstalk

Resumen

Plants are sessile organisms closely linked to their habitat and have thus developed a series of strategies that facilitate the adaptation of their metabolism to changing environmental conditions. A change in the environment can cause stress in plants and trigger metabolic changes, beginning with stress perception in order to optimize the plant response. All this alters gene expression, as well as the regulation of secondary metabolites and protein modifications, which are fundamental to a plant’s efficient response to stress. The production of reactive oxygen and nitrogen species (ROS/RNS) plays a key role in the plant's response to stress. Nitric oxide (NO) and ROS are capable of interacting with each other, as well as with other signal molecules, such as hormones. In addition, the concentration and subcellular location of RNS/ROS in plant tissues, which are essential for the functioning of these molecules, can have a signalling effect on many processes or be cytotoxic at high concentrations, leading to cellular damage. Plants have developed a series of highly regulated mechanisms that enable them to produce and eliminate NO and ROS in due measure. Furthermore, peroxisomes, which are highly dynamic and metabolically active organelles present in almost all eukaryotic cells, play an essential role in ROS/RNS homeostasis under control conditions and in stress responses. These organelles can interact with mitochondria and chloroplasts, share metabolic pathways and signalling and are involved in ROS detoxification, signalling and ROS/RNS sensing, as well in the import and transport of proteins to other organelles. Over the last twenty years, NO has been shown to be a key signal molecule in plant responses to stress. However, the role of NO in plant responses to cadmium is not fully understood, while little is known about its role in plant-pathogenic fungus interactions. On the other hand, transcriptomic analyses carried out in our laboratory showed a possible relationship between the plant's response to Cd and biotic stress, that can also be regulated by NO, which may play a key role in plant responses to both these stress conditions. Given the diversity of treatments in terms of criteria such as the metal concentrations used, plant species, plant growth and time of treatment, it is difficult to draw conclusions as to the role of NO in plant responses to heavy metals. In this thesis, we made a bioinformatics analysis of articles published over the last ten years on the production and/or function of NO in plant responses to heavy metals, including Cd. This analysis showed that exogenous applications of NO to the plant protect against heavy metals, particularly Cd, and that, in response to exposure to heavy metals, particularly Cd, plants initially produce NO, which can act as a signal molecule. At a later stage, the plant appears to be equipped with mechanisms to control NO levels, thus preventing further symptoms of toxicity. Using both biochemical techniques, involving modifications in NO levels through chemical donors and scavengers, and molecular approaches, involving the use of mutants with altered levels of NO, we subsequently analysed the role of NO in Arabidopsis seedling responses to Cd stress and its relationship to ROS. We also show that prolonged production of NO in plant responses to Cd can indeed affect antioxidant systems and induce oxidative stress, suggesting yet again that NO levels need to be strictly regulated in plant responses to Cd stress in order to prevent further damage to the plant. Peroxisomes have recently been shown to play a key role in a plant's early response to Cd stress. While both NO and RNS have been detected in these organelles, little is known about their effect on peroxisomal metabolism and dynamics. In this thesis, we analysed the role of NO in peroxisomal metabolism, distribution and dynamics under control conditions and in response to Cd stress. We showed that NO is involved in changes observed in peroxisomal dynamics which are necessary for the plant to respond to Cd stress. We also demonstrated the effect of NO on the oxidative metabolism of peroxisomes and their cellular distribution, as well as on organelle-dependent signalling. In addition, we analysed the little-known role of NO in Arabidopsis-Fusarium oxysporum interactions. We found that NO metabolism mutants have differential fungal responses with respect to WT in terms of ROS production, phenols, secondary and iron metabolisms; as well as defence gene induction Also, nitrate reductase appears to be essential for adequate cell wall assembly through the regulation of CESA4 and MYB46, given that the cell wall is a key barrier in the plant’s defence against Fusarium oxysporum. Finally, pre-treatment with Cd was found to protect plants against Fusarium oxysporum and to increase their survival. This could be explained by a priming effect, as certain genes are common to different plant responses to Cd and fungi, particularly Fusarium, suggesting that crosstalk takes place between both these stress conditions. Laura C. Terrón-Camero, María Rodríguez-Serrano, Luisa M. Sandalio and María C. Romero-Puertas. Nitric oxide is essential for peroxule production and peroxisome proliferation and signalling in response to Cd stress. Plant, Cell and Environment. In press Laura C. Terrón-Camero, Coral del Val, Luisa M. Sandalio and María C. Romero-Puertas (2020). Low endogenous NO levels in roots and antioxidant systems are determinants for the resistance of Arabidopsis seedlings grown in Cd. Environmental Pollution. 256, 113411. https://doi.org/10.1016/j.envpol.2019.113411 Laura C. Terrón-Camero, M. Ángeles Peláez-Vico, Coral del Val, Luisa M. Sandalio and María C. Romero-Puertas (2019). Role of nitric oxide in plant responses to heavy metal stress: exogenous application vs. endogenous production. Journal of Experimental Botany 70 (17), 4477-4488. María C. Romero-Puertas, Laura C. Terrón-Camero, M. Ángeles Peláez-Vico, Adela Olmedilla and Luisa M. Sandalio (2019). Reactive oxygen and nitrogen species as key indicators of plant responses to Cd stress. Environmental and Experimental Botany. 161, 107-119 Laura C. Terrón-Camero, Eliana Molina-Moya, María Sanz-Fernández, Luisa M. Sandalio, María C. Romero-Puertas (2018) Detection of Reactive Oxygen and Nitrogen Species (ROS/RNS) during Hypersensitive Cell Death. Methods in Molecular Biology 1743:97-105. doi: 10.1007/978-1-4939-7668-3_9.