Generation of fruit postharvest gene datasets and a novel motif analysis tool for functional studies: uncovering links between peach fruit heat treatment and cold storage responses.

MAIN CONCLUSION: A survey of developed fruit gene-specific datasets and the implementation of a novel cis-element analysis tool indicate specific transcription factors as novel regulatory actors under HT response and CI protection. Heat treatment (HT) prior to cold storage (CS) has been successfully applied to ameliorate fruit chilling injury (CI) disorders. Molecular studies have identified several HT-driven benefits and putative CI-protective molecules and mechanisms. However, bioinformatic tools and analyses able to integrate fruit-specific information are necessary to begin functional studies and breeding projects. In this work, a HT-responsive gene dataset (HTds) and four fruit expression datasets (FEds), containing gene-specific information from several species and postharvest conditions, were developed and characterized. FEds provided information about HT-responsive genes, not only validating their sensitivity to HT in different systems but also revealing most of them as CS-responsive. A special focus was given to peach heat treatment-sensitive transcriptional regulation by the development of a novel Perl motif analysis software (cisAnalyzer) and a curated plant cis-elements dataset (PASPds). cisAnalyzer is able to assess sequence motifs presence, localization, enrichment and discovery on biological sequences. Its implementation for the enrichment analysis of PASPds motifs on the promoters of HTds genes rendered particular cis-elements that indicate certain transcription factor (TF) families as responsible of fruit HT-sensitive transcription regulation. Phylogenetic and postharvest expression data of these TFs showed a functional diversity of TF families, with members able to fulfil roles under HT, CS and/or both treatments. All integrated datasets and cisAnalyzer tool were deposited in FruitGeneDB (https://www.cefobi-conicet.gov.ar/FruitGeneDB/search1.php), a new available database with a great potential for fruit gene functional studies, including the markers of HT and CS responses whose study will contribute to unravel HT-driven CI-protection and select tolerant cultivars.

Differential lipidome remodeling during postharvest of peach varieties with different susceptibility to chilling injury.

Peaches ripen and deteriorate rapidly at room temperature. Therefore, refrigeration is used to slow these processes and to extend fruit market life; however, many fruits develop chilling injury (CI) during storage at low temperature. Given that cell membranes are likely sites of the primary effects of chilling, the lipidome of six peach varieties with different susceptibility to CI was analyzed under different postharvest conditions. By using liquid chromatography coupled to mass spectrometry (LC-MS), 59 lipid species were detected, including diacyl- and triacylglycerides. The decreases in fruit firmness during postharvest ripening were accompanied by changes in the relative amount of several plastidic glycerolipid and triacylglyceride species, which may indicate their use as fuels prior to fruit senescence. In addition, levels of galactolipids were also modified in fruits stored at 0°C for short and long periods, reflecting the stabilization of plastidic membranes at low temperature. When comparing susceptible and resistant varieties, the relative abundance of certain species of the lipid classes phosphatidylethanolamine, phosphatidylcholine and digalactosyldiacylglycerol correlated with the tolerance to CI, reflecting the importance of the plasma membrane in the development of CI symptoms and allowing the identification of possible lipid markers for chilling resistance. Finally, transcriptional analysis of genes involved in galactolipid metabolism revealed candidate genes responsible for the observed changes after cold exposure. When taken together, our results highlight the importance of plastids in the postharvest physiology of fruits and provide evidence that lipid composition and metabolism have a profound influence on the cold response.

Synthesis, characterization and biological studies of a cobalt(III) complex of sulfathiazole.

The emergence of old and new antibiotic resistance created in the last decades revealed a substantial medical need for new classes of antimicrobial agents. The antimicrobial activity of sulfa drugs is often enhanced by complexation with metal ions, which is in concordance with the well-known importance of metal ions in biological systems. Besides, sulfonamides and its derivatives constitute an important class of drugs, with several types of pharmacological agents possessing antibacterial, anti-carbonic anhydrase, diuretic, hypoglycemic, antithyroid, antiviral and anticancer activities, among others. The purpose of this work has been the obtainment, characterization and determination of biological properties (antibacterial, antifungal, mutagenicity and phytotoxicity) of a new Co(III)-sulfathiazole complex: Costz, besides of its interaction with bovine serum albumin (BSA). The reaction between sodium sulfathiazole (Nastz) and cobalt(II) chloride in the presence of H2O2 leads to a brown solid, [CoIII(stz)2OH(H2O)3], (Costz). The structure of this compound has been examined by means of elemental analyses, FT-IR, 1H NMR, UV-Visible spectrometric methods and thermal studies. The Co(III) ion, which exhibits a distorted octahedral environment, could coordinate with the N thiazolic atom of sulfathiazolate. The complex quenched partially the native fluorescence of bovine serum albumin (BSA), suggesting a specific interaction with the protein. The Costz complex showed, in vitro, a moderate antifungal activity against Aspergillus fumigatus and A. flavus. As antibacterial, Costz displayed, in vitro, enhanced activity respective to the ligand against Pseudomonas aeruginosa. Costz did not show mutagenic properties with the Ames test. In the Allium cepa test the complex showed cytotoxic properties but not genotoxic ones. These results may be auspicious, however, further biological studies are needed to consider the complex Costz as a possible drug in the future.

Differential Metabolic Rearrangements after Cold Storage Are Correlated with Chilling Injury Resistance of Peach Fruits.

Reconfiguration of the metabolome is a key component involved in the acclimation to cold in plants; however, few studies have been devoted to the analysis of the overall metabolite changes after cold storage of fruits prior to consumption. Here, metabolite profiling of six peach varieties with differential susceptibility to develop mealiness, a chilling-injury (CI) symptom, was performed. According to metabolic content at harvest; after cold treatment; and after ripening, either following cold treatment or not; peach fruits clustered in distinct groups, depending on harvest-time, cold treatment, and ripening state. Both common and distinct metabolic responses among the six varieties were found; common changes including dramatic galactinol and raffinose rise; GABA, Asp, and Phe increase; and 2-oxo-glutarate and succinate decrease. Raffinose content after long cold treatment quantitatively correlated to the degree of mealiness resistance of the different peach varieties; and thus, raffinose emerges as a candidate biomarker of this CI disorder. Xylose increase after cold treatment was found only in the susceptible genotypes, indicating a particular cell wall reconfiguration of these varieties while being cold-stored. Overall, results indicate that peach fruit differential metabolic rearrangements due to cold treatment, rather than differential metabolic priming before cold, are better related with CI resistance. The plasticity of peach fruit metabolism renders it possible to induce a diverse metabolite array after cold, which is successful, in some genotypes, to avoid CI.

Cell wall-related genes studies on peach cultivars with differential susceptibility to woolliness: looking for candidates as indicators of chilling tolerance.

KEY MESSAGE: The results obtained indicate that a ß-xylosidase gene may act as good indicator of chilling tolerance and provide new insights into the complex issue of peach fruit woolliness. The storage of peaches at low temperatures for prolonged periods can induce a form of chilling injury (CI) called woolliness, characterized by a lack of juiciness and a mealy texture. As this disorder has been associated with abnormal cell wall dismantling, the levels of 12 transcripts encoding proteins involved in cell wall metabolism were analysed in cultivars with contrasting susceptibility to this disorder selected from five melting flesh peach cultivars. The resistant ('Springlady') and susceptible ('Flordaking') cultivars displayed differences in the level of expression of some of the selected genes during fruit softening and in woolly versus non-woolly fruits. From these genes, the level of expression of PpXyl, which encodes for a putative ß-xylosidase, was the one that presented the highest correlation (negative) with the susceptibility to woolliness. PpXyl expression was also analysed in a cultivar ('Rojo 2') with intermediate susceptibility to woolliness, reinforcing the conclusion about the correlation of PpXyl expression to the presence of woolliness symptom. Moreover, the level of expression of PpXyl correlated to protein level detected by Western blot. Analyses of the promoter region of the PpXyl gene (1637 bp) isolated from the three cultivars showed no differences suggesting that cis-elements from other regions of the genome and/or trans elements could be responsible of the differential PpXyl expression patterns. Overall, the results obtained indicate that PpXyl may act as a good indicator of woolliness tolerance and that the regulation of expression of this gene in different cultivars does not depend on sequences upstream the coding sequence.

Metabolic profiling of a range of peach fruit varieties reveals high metabolic diversity and commonalities and differences during ripening.

Peach (Prunus persica) fruits from different varieties display differential organoleptic and nutritional properties, characteristics related to their chemical composition. Here, chemical biodiversity of peach fruits from fifteen varieties, at harvest and after post-harvest ripening, was explored by gas chromatography-mass spectrometry. Metabolic profiling revealed that metabolites involved in organoleptic properties (sugars, organic and amino acids), stress tolerance (raffinose, galactinol, maltitol), and with nutritional properties (amino, caffeoylquinic and dehydroascorbic acids) displayed variety-dependent levels. Peach varieties clustered into four groups: two groups of early-harvest varieties with higher amino acid levels; two groups of mid- and late-harvest varieties with higher maltose levels. Further separation was mostly dependent on organic acids/raffinose levels. Variety-dependent and independent metabolic changes associated with ripening were detected; which contribute to chemical diversity or can be used as ripening markers, respectively. The great variety-dependent diversity in the content of metabolites that define fruit quality reinforces metabolomics usage as a tool to assist fruit quality improvement in peach.

Deciphering the metabolic pathways influencing heat and cold responses during post-harvest physiology of peach fruit.

Peaches are highly perishable and deteriorate quickly at ambient temperature. Cold storage is commonly used to prevent fruit decay; however, it affects fruit quality causing physiological disorders collectively termed 'chilling injury' (CI). To prevent or ameliorate CI, heat treatment is often applied prior to cold storage. In the present work, metabolic profiling was performed to determine the metabolic dynamics associated with the induction of acquired CI tolerance in response to heat shock. 'Dixiland' peach fruits exposed to 39 °C, cold stored, or after a combined treatment of heat and cold, were compared with fruits ripening at 20 °C. Dramatic changes in the levels of compatible solutes such as galactinol and raffinose were observed, while amino acid precursors of the phenylpropanoid pathway were also modified due to the stress treatments, as was the polyamine putrescine. The observed responses towards temperature stress in peaches are composed of both common and specific response mechanisms to heat and cold, but also of more general adaptive responses that confer strategic advantages in adverse conditions such as biotic stresses. The identification of such key metabolites, which prime the fruit to cope with different stress situations, will likely greatly accelerate the design and the improvement of plant breeding programs.

Muscle development and differentiation in the urodele Ambystoma mexicanum.

Muscle differentiation has been widely described in zebrafish and Xenopus, but nothing is known about this process in amphibian urodeles. Both anatomical features and locomotor activity in urodeles are known to show intermediate features between fish and anurans. Therefore, a better understanding of myogenesis in urodeles could be useful to clarify the evolutionary changes that led to the formation of skeletal muscle in the trunk of land vertebrates. We report here a detailed morphological and molecular investigation on several embryonic stages of Ambystoma mexicanum and show that the first differentiating muscle fibers are the slow ones, originating from a myoblast population initially localized close to the notochord that forms a superficial layer on the somitic surface afterwards. Subsequently, fast fibers differentiation ensues. We also identified and cloned A. mexicanum Myf5 as a muscle-specific transcriptional factor likely involved in urodele muscle differentiation.

Synthesis, characterization and antimicrobial properties of a Co(II)-phthalylsulfathiazolate complex.

The reaction between phthalylsulfathiazole (H(2)PST), in alkaline aqueous solution, and cobalt(II) nitrate led to a pink solid, [Co(PST)(H(2)O)(4)] (1), which was characterized by elemental and thermogravimetric analysis; FT-IR, Raman and diffuse reflectance spectra. Spectroscopic data reveal that the ligand would be doubly deprotonated and that the Co(II) ion environment is a distorted octahedral one. (1) showed antibacterial activity similar to the ligand.