Cell organisation,sulphur metabolism and ion transport-related genes are differentially expressed in Paracoccidioides brasiliensis mycelium and yeast cells

Background

Bread wheat (Triticum aestivum) is an important staple food. However, wheat gluten proteins cause celiac disease (CD) in 0.5 to 1% of the general population. Among these proteins, the α-gliadins contain several peptides that are associated to the disease.

Results

We obtained 230 distinct α-gliadin gene sequences from severaldiploid wheat species representing the ancestral A, B, and D genomes of the hexaploid bread wheat. The large majority of these sequences (87%) contained an internal stop codon. All α-gliadin sequences could be distinguished according to the genome of origin on the basis of sequence similarity, of the average length of the polyglutamine repeats, and of the differences in the presence of four peptides that have been identified as T cell stimulatory epitopes in CD patients through binding to HLA-DQ2/8. By sequence similarity, α-gliadins from the public database of hexaploid T. aestivum could be assigned directly to chromosome 6A, 6B, or 6D. T. monococcum (A genome) sequences, as well as those from chromosome 6A of bread wheat, almost invariably contained epitope glia-α9 and glia-α20, but never the intact epitopes glia-α and glia-α2. A number of sequences from T. speltoides, as well as a number of sequences fromchromosome 6B of bread wheat, did not contain any of the four T cell epitopes screened for. The sequences from T. tauschii (D genome), as well as those from chromosome 6D of bread wheat, were found to contain all of these T cell epitopes in variable combinations per gene. The differences in epitope composition resulted mainly from point mutations. These substitutions appeared to be genome specific.

Conclusion

Our analysis shows that α-gliadin sequences from the three genomes of bread wheat form distinct groups. The four known T cell stimulatory epitopes are distributed non-randomly across the sequences, indicating that the three genomes contribute differently to epitope content. A systematic analysis of all known epitopes in gliadins and glutenins will lead to better understanding of the differences in toxiCity among wheat varieties. On the basis of such insight, breeding strategies can be designed to generate less toxic varieties of wheat which may be tolerated by at least part of the CD patient population.     

Light-to-dark transitions trigger a transient increase in intracellular Ca2+ modulated by the redox state of the photosynthetic electron transport chain in the cyanobacterium Anabaena sp. PCC7120

Light‐to‐dark transitions represent one of the most crucial environmental stresses that photosynthetic organisms must cope with, since substantial metabolism adaptations are required in order to utilize alternative energy and carbon sources. Although signal transduction systems for changing light regimes are not sufficiently understood, calcium has been implicated in plants as a second messenger in light‐on and light‐off events. Much less is known about light signalling in cyanobacteria, but it has been shown that calcium probably performs similar signalling roles in these organisms and other prokaryotes. Herein it is reported that light‐to‐dark transitions trigger a calcium transient in aequorin expressing Anabaena sp. PCC7120. The magnitude of this transient depends on the fluence rate previously irradiated and can reach a peak height over 2 µm free calcium when the fluence rate of light is around 400 µmol photons s^?1 m^?2. The use of increasing calcium concentration, ethylene glycol‐bis (β‐aminoethylether) N,N,N′,N′‐tetraacetic acid (EGTA), verapamil and trifluoperazine indicated that these transients are originated by a calcium influx probably through verapamil‐sensitive Ca²⁺ channels and are probably modulated by calcium‐binding proteins. Experiments with different light spectral qualities and the photosynthetic inhibitors 3‐(3,4 dichlorophenyl)1,1,dimelthylurea (DCMU) and 3,5‐dibromo‐3‐methyl‐b‐isopropyl‐p‐benzoquinone (DBMIB) indicate that the calcium transient triggered by the light‐to‐dark transition is not coupled to a specific photoreceptor but rather to changes in the redox state of photosynthetic electron transport chain components other than the plastoquinone pool.     

The function of TLR4 in interferon gamma or interleukin-13 exposed and lipopolysaccharide stimulated gingival epithelial cell cultures

Gingival epithelial cells are part of the first line of host defense against infection. Toll-like receptors (TLRs) serve important immune and nonimmune functions. We investigated how interferon gamma (INF-γ) and interleukin 13 (IL-13) are involved in the TLR4 ligand-induced regulation of interleukin-8 (IL-8) effects on gingival epithelial cells. We used immunohistochemistry to localize TLR4 in ten healthy and ten periodontitis tissue specimens. Gingival epithelial cells then were primed with Th1 cytokine (INF-γ) or Th2 cytokine (IL-13) before stimulation with Escherichia coli-derived lipopolysaccharide (LPS) and enzyme-linked immunosorbent assay (ELISA) was performed to detect the level of IL-8 secretion in cell culture supernatants. Although both healthy and periodontitis gingival tissue samples expressed TLR4, the periodontitis samples showed more intense expression on gingival epithelial cells. Gingival epithelial cell cultures were primed with either INF-γ or IL-13 before stimulation with TLR4 ligand. Supernatants from co-stimulated epithelial cells exhibited IL-8 production in opposite directions, i.e., as one stimulates the release, the other reduces the release. INF-γ significantly increased TLR4 function, whereas IL-13 significantly decreased TLR4 function, i.e., production of IL-8. Pathogen associated molecular pattern-LPS, shared by many different periodonto-pathogenic bacteria, activates the gingival epithelial cells in a TLR-dependent manner. Diminished or increased TLR function in gingival epithelial cells under the influence of different Th cell types may protect or be harmful due to the altered TLR signaling.     

Effects of contrasting demographic histories on selection at major histocompatibility complex loci in two sympatric species of tuco‐tucos (Rodentia: Ctenomyidae)

To explore the impact of history on selection and genetic structure at functional loci, we compared patterns of major histocompatibility complex (MHC) variability in two sympatric species of ctenomyid rodents with different demographic backgrounds. Although Ctenomys talarum has experienced a stable demographic history, Ctenomys australis has undergone a recent demographic expansion. Accordingly, we predicted that MHC allele frequency distributions should be more skewed, differences between coding and noncoding regions should be less pronounced, and evidence of current selection on MHC loci should be reduced in C. australis relative to C. talarum. To test these predictions, we compared variation at the MHC class II DRB and DQA genes with that at multiple neutral markers, including DQA intron 2, the mitochondrial control region, and 8–12 microsatellite loci. These analyses supported the first two of our predictions but indicated that estimates of selection (based on ω‐values) were greater for C. australis. Further exploration of these data, however, revealed differences in the time frames over which selection appears to have acted on each species, with evidence of contemporary selection on MHC loci being limited to C. talarum. Collectively, these findings indicate that demographic history can substantially influence genetic structure at functional loci and that the effects of history on selection may be temporally complex and dynamic. © 2010 The Linnean Society of London, Biological Journal of the Linnean Society, 2010, 99 , 260–277.     

Proteomic analysis of tomato (Lycopersicon esculentum) pollen

In flowering plants, pollen grains are produced in the anther and released to the external environment with the primary function of delivering sperm cells to the female gametophyte. This study was conducted to identify proteins in tomato pollen and to analyse their roles in relation to pollen function. Tomato is an important crop which is grown worldwide and is an excellent experimental system. Proteins were extracted from pollen, separated by two-dimensional gel electrophoresis (2-DE), and identified by matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF MS) and peptide mass fingerprinting. Of the 960 spots observed on Colloidal Coomassie Blue (CCB)-stained 2-DE gels, 190 were selected for analysis. Of these, 158 spots, representing 133 distinct proteins, were identified by searching the NCBInr and Expressed Sequence Tag databases. The identified proteins were classified based on designated functions and the majority included those involved in defence mechanisms, energy conversions, protein synthesis and processing, cytoskeleton formation, Ca(2+) signalling, and as allergens. A number of proteins in tomato pollen were similar to those reported in the pollen of other species; however, several additional proteins with roles in defence mechanisms, metabolic processes, and hormone signalling were identified. The potential roles of the identified proteins in the survival strategy of the small, independent, two-celled pollen grain of tomato, and subsequently in pollen germination and tube growth are discussed.     

Broad-spectrum antimicrobial activity of volatile organic compounds from endophytic Pseudomonas putida BP25 against diverse plant pathogens

ABSTRACT

Black pepper endophytic Pseudomonas putida BP25 produced diverse antimicrobial volatile organic compounds having potential for plant disease management. Chemically synthesised volatiles such as 2, 5-dimethyl pyrazine; 2-methyl pyrazine; dimethyl trisulphide; 2-ethyl 5-methyl pyrazine; and 2-ethyl 3, 6-dimethyl pyrazine showed inhibitory activity against oomycete pathogens, Phytophthora capsici & Pythium myriotylum; fungal pathogens, Rhizoctonia solani, Colletotrichum gloeosporioides, Athelia rolfsii, Gibberella moniliformis & Magnaporthe oryzae; bacterial pathogen, Ralstonia pseudosolanacearum and plant parasitic nematode, Radopholus similis. Among them, dimethyl trisulphide completely inhibited oomycete and fungal pathogens as well as R. similis at a concentration of 2.65?µg?cm^?3 under in vitro conditions. Pyrazines suppressed Phytophthora lesions on shoot cuttings of black pepper upon in planta volatile treatment. Dimethyl trisulphide was the only compound that exhibited soil fumigant activity against P. capsici, R. solani and A. rolfsii (6.25?µg?cm^?3), C. gloeosporioides and G. moniliformis (12.5?µg?cm^?3), and R. similis (50?µg?cm^?3). Altogether, endophytic Pseudomonas putida BP25 and its volatile organic compounds offer an alternative strategy for eco-friendly disease management in agriculture.     

Protection of HepG2 cells against acrolein toxicity by 2-cyano-3,12-dioxooleana-1,9-dien-28-imidazolide via glutathione-mediated mechanism

Acrolein is an environmental toxicant, mainly found in smoke released from incomplete combustion of organic matter. Several studies showed that exposure to acrolein can lead to liver damage. The mechanisms involved in acrolein-induced hepatocellular toxicity, however, are not completely understood. This study examined the cytotoxic mechanisms of acrolein on HepG2 cells. Acrolein at pathophysiological concentrations was shown to cause apoptotic cell death and an increase in levels of protein carbonyl and thiobarbituric acid reactive acid substances. Acrolein also rapidly depleted intracellular glutathione (GSH), GSH-linked glutathione-S-transferases, and aldose reductase, three critical cellular defenses that detoxify reactive aldehydes. Results further showed that depletion of cellular GSH by acrolein preceded the loss of cell viability. To further determine the role of cellular GSH in acrolein-mediated cytotoxicity, buthionine sulfoximine (BSO) was used to inhibit cellular GSH biosynthesis. It was observed that depletion of cellular GSH by BSO led to a marked potentiation of acrolein-mediated cytotoxicity in HepG2 cells. To further assess the contribution of these events to acrolein-induced cytotoxicity, triterpenoid compound 2-cyano-3,12-dioxooleana-1,9-dien-28-imidazolide (CDDO-Im) was used for induction of GSH. Induction of GSH by CDDO-Im afforded cytoprotection against acrolein toxicity in HepG2 cells. Furthermore, BSO significantly inhibited CDDO-Im-mediated induction in cellular GSH levels and also reversed cytoprotective effects of CDDO-Im in HepG2 cells. These results suggest that GSH is a predominant mechanism underlying acrolein-induced cytotoxicity as well as CDDO-Im-mediated cytoprotection. This study may provide understanding on the molecular action of acrolein which may be important to develop novel strategies for the prevention of acrolein-mediated toxicity.     

Leaf anatomical variation in Alchornea triplinervia (Spreng) Mull. Arg. (Euphorbiaceae) under distinct light and soil water regimes

Alchornea triplinervia (Spreng.) Müll. Arg. (Euphorbiaceae) is a tree which occurs in a broad range of habitats in Brazil. In the State of Rio de Janeiro, it occurs from montane forests to swamplands at sea level. A quantitative approach was used to examine the role of light and soil water regime on the variations found in anatomical traits of the palisade and spongy parenchyma, outer epidermal cell wall of the abaxial and adaxial surfaces, the percentage of sclerenchymatous area in relation to the total midrib area and the ratio of palisade to spongy parenchyma for five distinct ecological populations: M—late secondary montane forest (shaded, unflooded); M2—early secondary montane forest (semi-exposed, unflooded); SI—primary swamp forest (semi-exposed, flooded); S2—secondary swamp forest (exposed, flooded); and D—deforestation area (exposed, unflooded). Tukey tests were carried out for multiple comparisons, while one-way factor variance analysis was used to test for differences among ecological populations. A principal component analysis was used to order the populations as well as to find the higher variance component. These populations developed different response levels to the environmental factors studied, namely light and soil water regime. Light accounted for the variations found in palisade and spongy parenchyma while the combination of light and soil water determined the variations found regarding the outer epidermal cell wall of the abaxial surface, the percentage of sclerenchymatous area in relation to the total midrib area and the compaction of the spongy parenchyma. The separation of S1/M2 and S2/D populations into two groups was due to similar light regimes, which suggested that light was affecting the leaf anatomical variation of A. triplinervia more than the interaction of light and soil water regime.