The alkaliphilic bacterium Pseudomonas pseudoalcaligenes CECT5344 uses free cyanide and several metal−cyanide complexes as the sole nitrogen source and tolerates high concentrations of metals like copper, zinc and iron, which are present in the jewelry wastewaters. To understand deeply the regulatory mechanisms involved in the transcriptional regulation of cyanide-containing wastewaters detoxification by P. pseudoalcaligenes CECT5344, RNA-Seq has been performed from cells cultured with a cyanide-containing jewelry wastewater, sodium cyanide or ammonium chloride as the sole nitrogen source. Small RNAs (sRNAs) that may have potential regulatory functions under cyanotrophic conditions were identified. In total 20 sRNAs were identified to be differentially expressed when compared the jewelry residue versus ammonium as nitrogen source, 16 of which could be amplified successfully by RT-PCR. As predicted targets of these 16 sRNAs were several components of the nit1C gene cluster encoding the nitrilase NitC essential for cyanide assimilation, the cioAB gene cluster that codes for the cyanide-insensitive cytochrome bd-type terminal oxidase, the medium length-polyhydroxyalkanoates (ml-PHAs) gene cluster, and gene clusters related with a global nitrogen limitation response like those coding for glutamine synthase and urease. Other targets were non-clustered genes (or their products) involved in metal resistance and iron acquisition, such as metal extrusion systems and the ferric uptake regulatory (Fur) protein, and a GntR-like regulatory family member probably involved in the regulation of the cyanide assimilation process in the strain CECT5344. Induction of genes targeted by sRNAs in the jewelry residue was demonstrated by qRT-PCR.

Common bean (Phaseolus vulgaris L.), one of the most important legume crops, uses atmospheric nitrogen through symbiosis with soil rhizobia, reducing the need for nitrogen fertilization. However, this legume is particularly sensitive to drought conditions, prevalent in arid regions where this crop is cultured. Therefore, studying the response to drought is important to sustain crop productivity. We have used integrated transcriptomic and metabolomic analysis to understand the molecular responses to water deficit in a marker-class common bean accession cultivated under N₂ fixation or fertilized with nitrate (NO₃^–). RNA-seq revealed more transcriptional changes in the plants fertilized with NO₃^– than in the N₂-fixing plants. However, changes in N₂-fixing plants were more associated with drought tolerance than in those fertilized with NO₃^–. N₂-fixing plants accumulated more ureides in response to drought, and GC/MS and LC/MS analysis of primary and secondary metabolite profiles revealed that N₂-fixing plants also had higher levels of abscisic acid, proline, raffinose, amino acids, sphingolipids, and triacylglycerols than those fertilized with NO₃^–. Moreover, plants grown under nitrogen fixation recovered from drought better than plants fertilized with NO₃^–. Altogether we show that common bean plants grown under symbiotic nitrogen fixation were more protected against drought than the plants fertilized with nitrate.

Drought is probably the most harmful stress affecting common bean crops. Domestication, worldwide spread and local farming practices has entailed the development of a wide variety of common bean genotypes with different degrees of resistance to water stress. In this work, physiological and molecular responses to water stress have been compared in two common bean accessions, PHA-0683 and PMB-0220, previously identified as highly and moderately resistant to water stress, respectively. Our hypothesis was that only quantitative differences in the expression patterns of key genes should be found if molecular mechanisms regulating drought resistance are similar in the two accessions. However, results presented here indicate that the resistance to drought in PMB-0220 and PHA-0683 common bean accessions is regulated by different molecular mechanisms. Differential regulation of ABA synthesis and ABA signaling related genes among the two genotypes, and the control of the drought-induced senescence have a relevant contribution to the higher resistance level of PHA-0683 accession. Our results also suggest that expression patterns of key senescence-related transcription factors could be considered in the screening for drought resistance in common bean germplasm collections.

Drought is one of the most critical factors limiting legume crop productivity. Understanding the molecular mechanisms of drought tolerance in the common bean is required to improve the yields of this important crop under adverse conditions. In this work, RNA-seq analysis was performed to compare the transcriptome profiles of drought-stressed and well-irrigated plants of a previously characterized drought-tolerant common bean landrace. The analysis revealed responses related with the abscisic acid signaling, including downregulation of a phosphatase 2C (PP2C) and an abscisic acid-8′ hydroxylase, and upregulation of several key transcription factors and genes involved in cell wall remodeling, synthesis of osmoprotectants, protection of photosynthetic apparatus, and downregulation of genes involved in cell expansion. The results also highlighted a significant proportion of differentially expressed genes related to phosphate starvation response. In addition, the moderate detrimental effects of drought in the biomass of these tolerant plants were abolished by the addition of phosphate, thus indicating that, besides the ABA-mediated response, acquisition of phosphate could be crucial for the drought tolerance of this common bean genotype. These results provided information about the mechanisms involved in drought response of common bean response that could be useful for enhancing the drought tolerance of this important crop legume.

Adenine metabolism is important for common bean (Phaseolus vulgaris L) productivity since this legume uses ureides derived from the oxidation of purine nucleotides, as their primary nitrogen storage molecules. Purine nucleotides are produced from de novo synthesis or through salvage pathways. Adenine phosphoribosyl transferase (APRT) is the enzyme dedicated to adenine nucleobase salvage for nucleotide synthesis, but it also acts on the inactivation of cytokinin bases. In common bean, the APRT enzyme is encoded by four genes. Gene expression analysis, biochemical properties and subcellular location suggest functional differences among the common bean APRT isoforms. CRISPR/Cas9 targeted downregulation of two of the four PvAPRTs followed by metabolomics and physiological analyses of targeted hairy roots reveals that, although the two proteins have redundant functions, PvAPRT1 mostly participates in the salvage of adenine, whereas PvAPRT5 is the predominant form in the regulation of cytokinin homeostasis and stress responses with a high impact in root and nodule growth.

Drought stress reduces plant growth and yield of crops. Common bean (Phaseolus vulgaris L.) establishes symbiosis with rhizobia, ensuring an adequate nitrogen supply without fertilizers. However, the relationship with rhizobia is constrained by limited water availability which inhibits both nitrogen fixation and plant growth. In addition, physiological and molecular responses of common bean to drought are conditioned by the form of nitrogen assimilated. Therefore, understanding the molecular mechanism(s) triggered in common bean under water-deficit conditions is relevant to identify the best strategies to resist drought stress. With the objective of understanding the molecular responses of roots and nodules from common bean to water-deficit stress, plants cultivated under N₂-fixation or nitrate fertilization were exposed to ten days of water deprivation. Afterwards, transcriptomic analysis was performed in roots, while metabolome profiling was carried out in roots and nodules. Physiological results showed that under water-deficit, N₂-fixing plants increased their root biomass more than nitrate-fertilized plants. Furthermore, water-deficit stress induced more transcriptional changes in nitrate-fertilized plants than in N₂-fixing plants, including a larger number of transcription factors in these plants compared with the N₂-fixing plants. On the other hand, roots from N₂-fixing plants accumulated more metabolites with potential protective functions such as allantoin, proline, raffinose, abscisic acid, and flavonoids in response to water-deficit stress than plants fertilized with nitrate, indicating that symbiosis might facilitate a faster and more efficient response to water-deficit stress. Moreover, common bean nodules exposed to water-deficit stress accumulated proline and erythritol, but reduced their content of maltose, pyruvic acid and allantoin compared to their respective controls. Taken collectively, these findings suggest that, despite the inhibition of nodule activity, N₂-fixing plants respond better to water-deficit stress than nitrate-fertilized plants.