Concerted EP2 and EP4 receptor signaling stimulates autocrine prostaglandin E(2) activation in human podocytes

Glomerular hyperfiltration is an important mechanism in the development of albuminuria. During hyperfiltration, podocytes are exposed to increased fluid flow shear stress (FFSS) in Bowman's space. Elevated Prostaglandin E2 (PGE(2)) synthesis and upregulated cyclooxygenase 2 (Cox2) are associated with podocyte injury by FFSS. We aimed to elucidate a PGE2 autocrine/paracrine pathway in human podocytes (hPC). We developed a modified liquid chromatography tandem mass spectrometry (LC/ESI-MS/MS) protocol to quantify cellular PGE(2), 15-keto-PGE(2), and 13,14-dihydro-15-keto-PGE(2) levels. hPC were treated with PGE(2) with or without separate or combined blockade of prostaglandin E receptors (EP), EP2, and EP4. Furthermore, the effect of FFSS on COX2, PTGER2, and PTGER4 expression in hPC was quantified. In hPC, stimulation with PGE(2) led to an EP2- and EP4-dependent increase in cyclic adenosine monophosphate (cAMP) and COX2, and induced cellular PGE(2). PTGER4 was downregulated after PGE(2) stimulation in hPC. In the corresponding LC/ESI-MS/MS in vivo analysis at the tissue level, increased PGE(2) and 15-keto-PGE(2) levels were observed in isolated glomeruli obtained from a well-established rat model with glomerular hyperfiltration, the Munich Wistar Frömter rat. COX2 and PTGER2 were upregulated by FFSS. Our data thus support an autocrine/paracrine COX2/PGE(2) pathway in hPC linked to concerted EP2 and EP4 signaling

Preparation, Structure and Spectroscopic Properties of NH 4 [Ln(S 2 CNH 2 ) 4 ] ⋅ H 2 O (Ln=La, Eu)

The title compounds were prepared under mild ambient conditions by a facile co-precipitation route. NH4[Eu(S2CNH2)4] ⋅ H2O (a) and NH4[La(S2CNH2)4] ⋅ H2O (b) crystallize isotypically in the monoclinic space group P21/c with a=8.4461(3), b=13.6367(3), c=16.2945(5) Å, β=103.759(2)° (for (a)), and a=8.50484(9), b=13.84476(16), c=16.20816(17) Å, β=103.7644(11)° for (b), respectively. The spectroscopic data reveal the presence of a ligand-to-metal charge transfer (LMCT) process of low energy in a and in the solid solutions NH4[La1−xEux(S2CNH2)4] ⋅ H2O (x=0.016 and 0.05). Despite of the consequent efficient luminescent quenching, it was possible to recorded excitation and emission spectra at room temperature. These spectra are characterized by narrow bands due to intraconfigurational-4f transitions of the Eu3+ ion. However, broad bands associated to the LMCT state were also observed, mainly for the solid solutions NH4[La1−xEux(S2CNH2)4] ⋅ H2O (x=0.016 and 0.05). Consequently, an intramolecular energy transfer mechanism is proposed, taking into account the role of the LMCT on the spectroscopic properties of dithiocarbamate complexes

Cu-Co/ZnAl2O4 Catalysts for CO Conversion to Higher Alcohols Synthesized from Co-Precipitated Hydrotalcite Precursors

The role of Cu:Co composition in bi-metallic Cu-Co/ZnAl2O4 catalysts on higher alcohol synthesis (HAS) was investigated at H2:CO = 4. The addition of Cu strongly facilitated Co reduction upon catalyst activation and suppressed coke deposition during HAS. Formation of predominantly hydrocarbons and higher alcohols was observed on the bi-metallic catalysts. Co/ZnAl2O4 produced mainly CH4 and Cu/ZnAl2O4 mainly CH3OH, while at Cu:Co = 0.6 the best ethanol selectivity of 4.5 % was reached. The microstructure of the spent catalysts confirmed a close interaction of Cu and Co

Alanine Zipper-Like Coiled-Coil Domains Are Necessary for Homotypic Dimerization of Plant GAGA-Factors in the Nucleus and Nucleolus

GAGA-motif binding proteins control transcriptional activation or repression of homeotic genes. Interestingly, there are no sequence similarities between animal and plant proteins. Plant BBR/BPC-proteins can be classified into two distinct groups: Previous studies have elaborated on group I members only and so little is known about group II proteins. Here, we focused on the initial characterization of AtBPC6, a group II protein from Arabidopsis thaliana. Comparison of orthologous BBR/BPC sequences disclosed two conserved signatures besides the DNA binding domain. A first peptide signature is essential and sufficient to target AtBPC6-GFP to the nucleus and nucleolus. A second domain is predicted to form a zipper-like coiled-coil structure. This novel type of domain is similar to Leucine zippers, but contains invariant alanine residues with a heptad spacing of 7 amino acids. By yeast-2-hybrid and BiFC-assays we could show that this Alanine zipper domain is essential for homotypic dimerization of group II proteins in vivo. Interhelical salt bridges and charge-stabilized hydrogen bonds between acidic and basic residues of the two monomers are predicted to form an interaction domain, which does not follow the classical knobs-into-holes zipper model. FRET-FLIM analysis of GFP/RFP-hybrid fusion proteins validates the formation of parallel dimers in planta. Sequence comparison uncovered that this type of domain is not restricted to BBR/BPC proteins, but is found in all kingdoms

Genome-wide analysis of WRKY gene family in Cucumis sativus

<p>Abstract</p> <p>Background</p> <p>WRKY proteins are a large family of transcriptional regulators in higher plant. They are involved in many biological processes, such as plant development, metabolism, and responses to biotic and abiotic stresses. Prior to the present study, only one full-length cucumber WRKY protein had been reported. The recent publication of the draft genome sequence of cucumber allowed us to conduct a genome-wide search for cucumber WRKY proteins, and to compare these positively identified proteins with their homologs in model plants, such as <it>Arabidopsis</it>.</p> <p>Results</p> <p>We identified a total of 55 WRKY genes in the cucumber genome. According to structural features of their encoded proteins, the cucumber WRKY (<it>CsWRKY</it>) genes were classified into three groups (group 1-3). Analysis of expression profiles of <it>CsWRKY </it>genes indicated that 48 WRKY genes display differential expression either in their transcript abundance or in their expression patterns under normal growth conditions, and 23 WRKY genes were differentially expressed in response to at least one abiotic stresses (cold, drought or salinity). The expression profile of stress-inducible <it>CsWRKY </it>genes were correlated with those of their putative <it>Arabidopsis WRKY (AtWRKY) </it>orthologs, except for the group 3 WRKY genes. Interestingly, duplicated group 3 <it>AtWRKY </it>genes appear to have been under positive selection pressure during evolution. In contrast, there was no evidence of recent gene duplication or positive selection pressure among <it>CsWRKY </it>group 3 genes, which may have led to the expressional divergence of group 3 orthologs.</p> <p>Conclusions</p> <p>Fifty-five WRKY genes were identified in cucumber and the structure of their encoded proteins, their expression, and their evolution were examined. Considering that there has been extensive expansion of group 3 WRKY genes in angiosperms, the occurrence of different evolutionary events could explain the functional divergence of these genes.</p

The CC-NB-LRR-Type Rdg2a Resistance Gene Confers Immunity to the Seed-Borne Barley Leaf Stripe Pathogen in the Absence of Hypersensitive Cell Death

BACKGROUND: Leaf stripe disease on barley (Hordeum vulgare) is caused by the seed-transmitted hemi-biotrophic fungus Pyrenophora graminea. Race-specific resistance to leaf stripe is controlled by two known Rdg (Resistance to Drechslera graminea) genes: the H. spontaneum-derived Rdg1a and Rdg2a, identified in H. vulgare. The aim of the present work was to isolate the Rdg2a leaf stripe resistance gene, to characterize the Rdg2a locus organization and evolution and to elucidate the histological bases of Rdg2a-based leaf stripe resistance. PRINCIPLE FINDINGS: We describe here the positional cloning and functional characterization of the leaf stripe resistance gene Rdg2a. At the Rdg2a locus, three sequence-related coiled-coil, nucleotide-binding site, and leucine-rich repeat (CC-NB-LRR) encoding genes were identified. Sequence comparisons suggested that paralogs of this resistance locus evolved through recent gene duplication, and were subjected to frequent sequence exchange. Transformation of the leaf stripe susceptible cv. Golden Promise with two Rdg2a-candidates under the control of their native 5′ regulatory sequences identified a member of the CC-NB-LRR gene family that conferred resistance against the Dg2 leaf stripe isolate, against which the Rdg2a-gene is effective. Histological analysis demonstrated that Rdg2a-mediated leaf stripe resistance involves autofluorescing cells and prevents pathogen colonization in the embryos without any detectable hypersensitive cell death response, supporting a cell wall reinforcement-based resistance mechanism. CONCLUSIONS: This work reports about the cloning of a resistance gene effective against a seed borne disease. We observed that Rdg2a was subjected to diversifying selection which is consistent with a model in which the R gene co-evolves with a pathogen effector(s) gene. We propose that inducible responses giving rise to physical and chemical barriers to infection in the cell walls and intercellular spaces of the barley embryo tissues represent mechanisms by which the CC-NB-LRR-encoding Rdg2a gene mediates resistance to leaf stripe in the absence of hypersensitive cell death.Davide Bulgarelli, Chiara Biselli, Nicholas C. Collins, Gabriella Consonni, Antonio M. Stanca, Paul Schulze-Lefert and Giampiero Val

The rice transcription factor OsWRKY47 is a positive regulator of the response to water deficit stress

OsWRKY47 is a divergent rice transcription factor belonging to the group II of the WRKY family. A transcriptomic analysis of the drought response of transgenic rice plants expressing PSARK::IPT, validated by qPCR, indicated that OsWRKY47 expression was induced under drought stress in PSARK::IPT plants. A PCR-assisted site selection assay (SELEX) of recombinant OsWRKY47 protein showed that the preferred sequence bound in vitro is (G/T)TTGACT. Bioinformatics analyses identified a number of gene targets of OsWRKY47; among these two genes encode a Calmodulin binding protein and a Cys-rich secretory protein. Using Oswrk47 knockout mutants and transgenic rice overexpressing OsWRKY47 we show that the transcription of these putative targets were regulated by OsWRKY47. Phenotypic analysis carried out with transgenic rice plants showed that Oswrky47 mutants displayed higher sensitivity to drought and reduced yield, while plants overexpressing OsWRKY47 were more tolerant.Fil: Raineri, Jesica. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe. Instituto de Agrobiotecnología del Litoral. Universidad Nacional del Litoral. Instituto de Agrobiotecnología del Litoral; ArgentinaFil: Wang, Songhu. Chinese Academy of Sciences; República de China. University of California at Davis; Estados UnidosFil: Peleg, Zvi. The Hebrew University of Jerusalem; Israel. University of California at Davis; Estados UnidosFil: Blumwald, Eduardo. University of California at Davis; Estados UnidosFil: Chan, Raquel Lia. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe. Instituto de Agrobiotecnología del Litoral. Universidad Nacional del Litoral. Instituto de Agrobiotecnología del Litoral; Argentin

New seed collections of North American pitseed goosefoot (Chenopodium berlandieri) and efforts to identify its diploid ancestors through whole-genome sequencing

Pitseed goosefoot (Chenopodium berlandieri) is an ecologically diverse wild/weedy North American species within the primary gene pool for improving South American quinoa (Chenopodium quinoa). Both taxa are 36-chromosome allotetraploids with subgenomes AA and BB. The A genome is found in a large number of diploids in the Americas, along with one Northeast Asian taxon, and was recently shown to be the maternal ancestor, while the paternal B genome is closely related to several extant Eurasian diploids. Two of our primary objectives were 1) to determine the extent of genetic diversity in the allotetraploid C. berlandieri-quinoahircinum complex and 2) to characterize the evolutionary path from polyploidization to domestication in these taxa. In an effort to survey genetic diversity, in 2018, we made seed collections of southern Texas, southern Great Plains, and New England coastal ecotypes of C. berlandieri as well as sympatric diploids. With respect to the second goal, we performed wholegenome sequencing of two Sonoran Desert Chenopodium A-genome diploids in subsection Cellulata and Andean cultivated C. pallidicaule in subsection Leiosperma. When paired reads were aligned to the whole-genome reference of C. quinoa strain ‘QQ74’, the match percentages were 99.31, 99.23, and 98.53 for C. watsonii, C. sonorense, and C. pallidicaule, respectively. These data strongly support C. watsonii as being the most closely related of these three species to the A-genome ancestor of quinoa. Ongoing sequencing efforts with a larger panel of diploids are aimed at identifying the maternal ancestor of C. quinoa and C. berlandieri, if extant

15-keto-Prostaglandin E(2) exhibits bioactive role by modulating glomerular cytoarchitecture through EP2/EP4 receptors

AIMS: Prostaglandins are important signaling lipids with prostaglandin E(2) (PGE(2)) known to be the most abundant prostaglandin across tissues. In kidney, PGE(2) plays an important role in the regulation of kidney homeostasis through its EP receptor signaling. Catabolism of PGE(2) yields the metabolic products that are widely considered biologically inactive. Although recent in vitro evidence suggested the ability of 15-keto-PGE(2) (a downstream metabolite of PGE(2)) to activate EP receptors, the question whether 15-keto-PGE(2) exhibits physiological roles remains unresolved. MATERIALS AND METHODS: Pharmacological treatment was performed in transgenic zebrafish embryos using 500 µM 15-keto-PGE(2) and 20 µM EP receptors antagonists' solutions during zebrafish embryonic development. After the exposure period, the embryos were fixed for confocal microscopy imaging and glomerular morphology analysis. KEY FINDINGS: Here, we show that 15-keto-PGE(2) can bind and stabilize EP2 and EP4 receptors on the plasma membrane in the yeast model. Using lipidomic analysis, we demonstrate both PGE(2) and 15-keto-PGE(2) are present at considerable levels in zebrafish embryos. Our high-resolution image analysis reveals the exogenous treatment with 15-keto-PGE(2) perturbs glomerular vascularization during zebrafish development. Specifically, we show that the increased levels of 15-keto-PGE(2) cause intercalation defects between podocytes and endothelial cells of glomerular capillaries effectively reducing the surface area of glomerular filtration barrier. Importantly, 15-keto-PGE(2)-dependent defects can be fully reversed by combined blockade of the EP2 and EP4 receptors. SIGNIFICANCE: Altogether, our results reveal 15-keto-PGE(2) to be a biologically active metabolite that modulates the EP receptor signaling in vivo, thus playing a potential role in kidney biology