Chrysanthi Kalloniati

Sheep and goats differences in CLA and fatty acids milk fat content in relation with mRNA stearoyl-CoA desaturase and lipogenic genes expression in their mammary gland

An experiment was conducted with 12 lactating dairy ewes and 12 goats with the objective to determine whether, under the same dietary treatments, the differences in their fatty acid (FA) profile with emphasis on cis-9 trans-11 CLA milk fat content, are reflected in the transcript levels of genes involved in FA and cis-9, trans-11 CLA biosynthesis. The animals were fed with two diets (A, B) in different days of milk (DIM) due to the different milk yield, body weight etc, in order to have the same food intake and to avoid dietary effects. Diet A was fed to the animals on a group basis as it is traditionally used in practice, while diet B was chosen to avoid individual feed intake variation which is usually observed in group feeding. The results showed that there are significantly lower mRNA levels of acetyl-CoA carboxylase (ACC) in sheep mammary gland compared with those of goats, independently from the diet fed. The same trend was observed with the mRNA level of FA synthase (FAS), but the results were significant only for diet A. The mRNA level of lipoprotein lipase (LPL) in the mammary gland did not differ between sheep and goats fed with diet A. In addition, the concentration of cis-9 trans-11 CLA content was significantly higher in sheep milk fat compared with those of goats. This is in accordance with the significant higher levels on mRNA of stearoyl-CoA desaturase (SCD) which were observed in their mammary adipocytes of sheep compared with those of goats, independently of the fed diet (A or B). In conclusion, these findings demonstrate that the differences between sheep and goats, concerning cis-9, trans-11 CLA and FA milk fat content, under the same dietary treatments could be explained in part by the differences in mRNA of SCD and lipogenic genes in their mammary gland.

The study of two barley Type I-like MADS-box genes as potential targets of epigenetic regulation during seed development

BackgroundMADS-box genes constitute a large family of transcription factors functioning as key regulators of many processes during plant vegetative and reproductive development. Type II MADS-box genes have been intensively investigated and are mostly involved in vegetative and flowering development. A growing number of studies of Type I MADS-box genes in Arabidopsis, have assigned crucial roles for these genes in gamete and seed development and have demonstrated that a number of Type I MADS-box genes are epigenetically regulated by DNA methylation and histone modifications. However, reports on agronomically important cereals such as barley and wheat are scarce.ResultsHere we report the identification and characterization of two Type I-like MADS-box genes, from barley (Hordeum vulgare), a monocot cereal crop of high agronomic importance. Protein sequence and phylogenetic analysis showed that the putative proteins are related to Type I MADS-box proteins, and classified them in a distinct cereal clade. Significant differences in gene expression among seed developmental stages and between barley cultivars with varying seed size were revealed for both genes. One of these genes was shown to be induced by the seed development- and stress-related hormones ABA and JA whereas in situ hybridizations localized the other gene to specific endosperm sub-compartments. The genomic organization of the latter has high conservation with the cereal Type I-like MADS-box homologues and the chromosomal position of both genes is close to markers associated with seed quality traits. DNA methylation differences are present in the upstream and downstream regulatory regions of the barley Type I-like MADS-box genes in two different developmental stages and in response to ABA treatment which may be associated with gene expression differences.ConclusionsTwo barley MADS-box genes were studied that are related to Type I MADS-box genes. Differential expression in different seed developmental stages as well as in barley cultivars with different seed size was evidenced for both genes. The two barley Type I MADS-box genes were found to be induced by ABA and JA. DNA methylation differences in different seed developmental stages and after exogenous application of ABA is suggestive of epigenetic regulation of gene expression. The study of barley Type I-like MADS-box genes extends our investigations of gene regulation during endosperm and seed development in a monocot crop like barley.

Characterization of a Mesorhizobium loti α-Type Carbonic Anhydrase and Its Role in Symbiotic Nitrogen Fixation

Carbonic anhydrase (CA) (EC 4.2.1.1) is a widespread enzyme catalyzing the reversible hydration of CO(2) to bicarbonate, a reaction that participates in many biochemical and physiological processes. Mesorhizobium loti, the microsymbiont of the model legume Lotus japonicus, possesses on the symbiosis island a gene (msi040) encoding an alpha-type CA homologue, annotated as CAA1. In the present work, the CAA1 open reading frame from M. loti strain R7A was cloned, expressed, and biochemically characterized, and it was proven to be an active alpha-CA. The biochemical and physiological roles of the CAA1 gene in free-living and symbiotic rhizobia were examined by using an M. loti R7A disruption mutant strain. Our analysis revealed that CAA1 is expressed in both nitrogen-fixing bacteroids and free-living bacteria during growth in batch cultures, where gene expression was induced by increased medium pH. L. japonicus plants inoculated with the CAA1 mutant strain showed no differences in top-plant traits and nutritional status but consistently formed a higher number of nodules exhibiting higher fresh weight, N content, nitrogenase activity, and delta(13)C abundance. Based on these results, we propose that although CAA1 is not essential for nodule development and symbiotic nitrogen fixation, it may participate in an auxiliary mechanism that buffers the bacteroid periplasm, creating an environment favorable for NH(3) protonation, thus facilitating its diffusion and transport to the plant. In addition, changes in the nodule delta(13)C abundance suggest the recycling of at least part of the HCO(3)(-) produced by CAA1.

Cessation of photosynthesis in Lotus japonicus leaves leads to reprogramming of nodule metabolism

Symbiotic nitrogen fixation (SNF) involves global changes in gene expression and metabolite accumulation in both rhizobia and the host plant. In order to study the metabolic changes mediated by leaf–root interaction, photosynthesis was limited in leaves by exposure of plants to darkness, and subsequently gene expression was profiled by real-time reverse transcription–PCR (RT–PCR) and metabolite levels by gas chromatography–mass spectrometry in the nodules of the model legume Lotus japonicus. Photosynthetic carbon deficiency caused by prolonged darkness affected many metabolic processes in L. japonicus nodules. Most of the metabolic genes analysed were down-regulated during the extended dark period. In addition to that, the levels of most metabolites decreased or remained unaltered, although accumulation of amino acids was observed. Reduced glycolysis and carbon fixation resulted in lower organic acid levels, especially of malate, the primary source of carbon for bacteroid metabolism and SNF. The high amino acid concentrations together with a reduction in total protein concentration indicate possible protein degradation in nodules under these conditions. Interestingly, comparisons between amino acid and protein content in various organs indicated systemic changes in response to prolonged darkness between nodulated and non-nodulated plants, rendering the nodule a source organ for both C and N under these conditions.

Nitrogen-Fixing Nodules Are an Important Source of Reduced Sulfur, Which Triggers Global Changes in Sulfur Metabolism in Lotus japonicus

In addition to providing biologically available nitrogen, nitrogen-fixing nodules represent an important site of thiol biosynthesis, which reprograms sulfur partitioning and metabolism in the plant. We combined transcriptomic and biochemical approaches to study rhizobial and plant sulfur (S) metabolism in nitrogen (N) fixing nodules (Fix+) of Lotus japonicus, as well as the link of S-metabolism to symbiotic nitrogen fixation and the effect of nodules on whole-plant S-partitioning and metabolism. Our data reveal that N-fixing nodules are thiol-rich organs. Their high adenosine 5′-phosphosulfate reductase activity and strong 35S-flux into cysteine and its metabolites, in combination with the transcriptional upregulation of several rhizobial and plant genes involved in S-assimilation, highlight the function of nodules as an important site of S-assimilation. The higher thiol content observed in nonsymbiotic organs of N-fixing plants in comparison to uninoculated plants could not be attributed to local biosynthesis, indicating that nodules are an important source of reduced S for the plant, which triggers whole-plant reprogramming of S-metabolism. Enhanced thiol biosynthesis in nodules and their impact on the whole-plant S-economy are dampened in plants nodulated by Fix− mutant rhizobia, which in most respects metabolically resemble uninoculated plants, indicating a strong interdependency between N-fixation and S-assimilation.

Maintenance of metabolic homeostasis and induction of cytoprotectants and secondary metabolites in alachlor-treated GmGSTU4-overexpressing tobacco plants, as resolved by metabolomics

Herbicides are an invaluable tool for agricultural production scaling up. However, their continuous and intensive use has led to an increased incidence of herbicide resistant weeds and environmental pollution. Plant glutathione transferases (GSTs) are tightly connected with crop and weed herbicide tolerance capacitating their efficient metabolic detoxification, thus GSTs can be biotechnologically exploited towards addressing those issues. However, information on their effects at a “systems” level in response to herbicides is lacking. Here, we aimed to study the effects of the chloroacetanilide herbicide alachlor on the metabolome of wild-type and tobacco plants overexpressing the soybean tau class glutathione transferase GmGSTU4. Alachlor-treated wild-type plants This system, naturally serving the detoxification of endogenous exhibited an abiotic stress-like response with increased abundance of compatible solutes, decrease in TCA cycle intermediates and decreased sugar and amino acid content. Transgenic plants responded distinctly, exhibiting an increased induction of abiotic stress responsive metabolites, accumulation of secondary metabolites and its precursors, and metabolic detoxification by-products compared to wild-type plants. These results suggest that the increased metabolic capacity of GmGSTU4 overexpressing plants is accompanied by pleiotropic metabolic alterations, which could be the target for further manipulation in order to develop herbicide resistant crops, plants with increased phytoremediation potential, as well as efficient management of non-target site, GST induced, herbicide resistance in weeds.

Global metabolomics analysis reveals distinctive tolerance mechanisms in different plant organs of lentil (Lens culinaris) upon salinity stress

Background and AimsOmic technologies in the past years have provided a variety of data in model plants. In legumes, results οn Lotus japonicus and Medicago truncatula have highlighted the biochemistry which takes place inside cells under a variety of abiotic stresses. Here we conducted metabolomics in the forage legume lentil (Lens culinaris) upon salinity stress on acclimated and non-acclimated plants and compared results from leaf and root analyses.MethodsWe used two lentil varieties, originated from different geographical locations and studied differences in their global metabolite profile i) using gradual or initial application of salt stress, ii) between leaves and roots, and iii) between the varieties.ResultsMost important differences were noted in salinity induced diminished abundance of organic acids in both varieties’ leaves and roots, accumulation of sugars and polyols in leaves, and accumulation of other key-metabolites, such as L-asparagine, D-trehalose, allantoin and urea in the roots. We also demonstrated the driver of deleterious Cl− accumulation in leaves for potential compartmentalization in the vacuole, a defensive mechanism for withstanding salinity stress in plants. Finally, a model is suggested of how legumes upregulate a metabolic pathway, which involves purines catabolism in order to assimilate carbon and nitrogen, which are limited during salinity stress.ConclusionsFuture omics works with lentil can help understanding the regulation of the biochemical “arsenal” against abiotic stresses such as salinity and render the selection of better crops.