Partial Resistance of Carrot to Alternaria dauci Correlates with In Vitro Cultured Carrot Cell Resistance to Fungal Exudates

Although different mechanisms have been proposed in the recent years, plant pathogen partial resistance is still poorly understood. Components of the chemical warfare, including the production of plant defense compounds and plant resistance to pathogen-produced toxins, are likely to play a role. Toxins are indeed recognized as important determinants of pathogenicity in necrotrophic fungi. Partial resistance based on quantitative resistance loci and linked to a pathogen-produced toxin has never been fully described. We tested this hypothesis using the Alternaria dauci – carrot pathosystem. Alternaria dauci, causing carrot leaf blight, is a necrotrophic fungus known to produce zinniol, a compound described as a non-host selective toxin. Embryogenic cellular cultures from carrot genotypes varying in resistance against A. dauci were confronted with zinniol at different concentrations or to fungal exudates (raw, organic or aqueous extracts). The plant response was analyzed through the measurement of cytoplasmic esterase activity, as a marker of cell viability, and the differentiation of somatic embryos in cellular cultures. A differential response to toxicity was demonstrated between susceptible and partially resistant genotypes, with a good correlation noted between the resistance to the fungus at the whole plant level and resistance at the cellular level to fungal exudates from raw and organic extracts. No toxic reaction of embryogenic cultures was observed after treatment with the aqueous extract or zinniol used at physiological concentration. Moreover, we did not detect zinniol in toxic fungal extracts by UHPLC analysis. These results suggest that strong phytotoxic compounds are present in the organic extract and remain to be characterized. Our results clearly show that carrot tolerance to A. dauci toxins is one component of its partial resistance.     

Design of new potent and selective secretory phospholipase A2 inhibitors. Part 5: synthesis and biological activity of 1-alkyl-4-[4,5-dihydro-1,2,4-[4H]-oxadiazol-5-one-3-ylmethylbenz-4'-yl(oyl)] piperazines

Among the different PLA(2)s identified to date, the group IIA secretory PLA(2) (sPLA(2) GIIA) is implied in diverse pathological conditions. In this work we describe the synthesis, inhibitory activities, and structure-activity relationships (SAR) of a new class of substituted piperazine derivatives. The in vitro fluorimetric assay using two groups of enzymes, GIB and GIIA, revealed several compounds as highly potent inhibitors (IC(50)=0.1 microM). The in vivo activity assessed by ip or per os administration in a carrageenan-induced edema test in rats showed that two compounds proved to be as potent as indomethacin (10 mg/kg).     

Immunoaffinity purification and characterization of glyceraldehyde-3-phosphate dehydrogenase from human erythrocytes

A new procedure utilizing immunoaffinity column chromatography has been used for the purification of glyceraldehyde-3-phosphate dehydrogenase （GAPDH, EC 1.2.1.12） from human erythrocytes. The comparison between this rapid method （one step） and the tra- ditional procedure including ammonium sulfate fractionation followed by Blue Sepharose CL-6B chromatography shows that the new method gives a highest specific activity with a highest yield in a short time. The characterization of the purified GAPDH reveals that the native enzyme is a homotetramer of -150 kDa with an absolute specificity for the oxidized form of nicotinamide adenine dinucleotide （NAD＋）. Western blot analysis using purified monospecific polyclonal antibodies raised against the purified GAPDH showed a single 36 kDa band corresponding to the enzyme subunit. Studies on the effect of temperature and pH on enzyme activity revealed optimal values of about 43℃ and 8.5, respectively. The kinetic parameters were also calculated： the Vmax was 4.3 U/mg and the Km values against G3P and NAD＋ were 20.7 and 17.8 μM, respectively. The new protocol described represents a simple, economic, and reproducible tool for the purification of GAPDH and can be used for other proteins.     

A 12 kb multi-allelic copy number variation encompassing a GC gene enhancer is associated with mastitis resistance in dairy cattle

Clinical mastitis (CM) is an inflammatory disease occurring in the mammary glands of lactating cows. CM is under genetic control, and a prominent CM resistance QTL located on chromosome 6 was reported in various dairy cattle breeds. Nevertheless, the biological mechanism underpinning this QTL has been lacking. Herein, we mapped, fine-mapped, and discovered the putative causal variant underlying this CM resistance QTL in the Dutch dairy cattle population. We identified a ~12 kb multi-allelic copy number variant (CNV), that is in perfect linkage disequilibrium with a lead SNP, as a promising candidate variant. By implementing a fine-mapping and through expression QTL mapping, we showed that the group-specific component gene (GC), a gene encoding a vitamin D binding protein, is an excellent candidate causal gene for the QTL. The multiplicated alleles are associated with increased GC expression and low CM resistance. Ample evidence from functional genomics data supports the presence of an enhancer within this CNV, which would exert cis-regulatory effect on GC. We observed that strong positive selection swept the region near the CNV, and haplotypes associated with the multiplicated allele were strongly selected for. Moreover, the multiplicated allele showed pleiotropic effects for increased milk yield and reduced fertility, hinting that a shared underlying biology for these effects may revolve around the vitamin D pathway. These findings together suggest a putative causal variant of a CM resistance QTL, where a cis-regulatory element located within a CNV can alter gene expression and affect multiple economically important traits.     

Characterization of fertility alteration and marker validation for male sterility genes in novel PTGMS lines hybrid rice

Photoperiod and thermosensitive genetic male sterile (PTGMS) lines have become one of the main sources of global rice production increasing. This study was conducted to evaluate the fertility alteration and validate the male sterility genes using validation markers in novel Egyptian Indica and Japonica PTGMS lines under natural conditions. The study revealed that the new genetic male sterile lines belong to the type of photo–thermosensitive genetic male sterility (PTGMS). The fertility alteration of these lines has influenced by photoperiod and temperature interaction. The new PTGMS lines have three sensitive periods of fertility alteration; transformation, sterility, and fertility period. Furthermore, the sensitive stage of fertility transformation might be from secondary branch primordial to pollen mother cells (PMC) meiosis. Under the natural Sakha condition, the new PTGMS lines were stable sterile under the condition of day length upper 13,75 h and temperature over 25 °C, while its convert to fertile under day length under 13 h, and temperature lower than 24 °C. The co-dominant markers identified the pms3 and tms5 genes in the new PTGMS lines, indicated that the fertility alteration in these lines controlled by photoperiod and thermosensitive stages.     

Cellular and Subcellular Aquaporin-4 Distribution in the Mouse Neurohypophysis and the Effects of Osmotic Stimulation

Water channel aquaporin-4 (AQP4) is the most abundant water channel in the rodent brain and is mainly expressed in cerebral areas involved in central osmoreception and osmoregulation. The neurohypophysis is the release site of hypothalamic neurohormones vasopressin and oxytocin, which are involved in the regulation of the water balance. The authors investigated the cellular and subcellular distribution of AQP4 in the mouse neurohypophysis before and after chronic osmotic stimulation, using immunofluorescence microscopy and immunoperoxidase electron microscopy. They showed that AQP4 was abundant in the mouse hypophysis, mainly in the neural lobe. AQP4 was discontinuously distributed along pituicytes plasma membranes, in the dense neurosecretory granules and microvesicles of nerve endings and fibers, and along the luminal and abluminal membranes of fenestrated capillary endothelial cells. After chronic osmotic stimulation, AQP4 immunolabeling was enhanced. Taken together, these results suggest that AQP4 could be involved in the pituicyte sensor effect during osmoregulation, the modification and/or maturation mechanism of neurosecretory granules during neurohormone release, and the blood perfusion of the hypophysis.     

Microorganisms and Maillard reaction products: a review of the literature and recent findings

Research on the impact of Maillard reaction products (MRPs) on microorganisms has been reported in the literature for the last 60 years. In the current study, the impact of an MRP-rich medium on the growth of three strains of Escherichia coli was measured by comparing two classic methods for studying the growth of bacteria (plate counting and optical density at 600 nm) and by tracing MRP utilisation. Early stage and advanced MRPs in the culture media were assessed by quantifying furosine and N ^ε -carboxymethyllysine (CML) levels, respectively, using chromatographic methods. These measures were performed prior to and during bacterial growth to estimate the potential use of these MRPs by Escherichia coli CIP 54.8. Glucose and lysine, the two MRP precursors used in the MRP-rich medium, were also quantified by chromatographic means. Compared to control media, increased lag phases and decreased growth rates were observed in the MRP-rich medium for two out of the three Escherichia coli strains tested. In contrast, one strain isolated from the faeces of a piglet fed on a MRP-rich diet was not influenced by the presence of MRPs in the medium. Overall, CML as well as the products obtained by the thermal degradation of glucose and lysine, regardless of the Maillard reaction, did not affect the growth of the three strains tested. In addition, no degradation of fructoselysine or CML was found in the presence of Escherichia coli CIP 54.8.     

A 660-Kb Deletion with Antagonistic Effects on Fertility and Milk Production Segregates at High Frequency in Nordic Red Cattle: Additional Evidence for the Common Occurrence of Balancing Selection in Livestock

In dairy cattle, the widespread use of artificial insemination has resulted in increased selection intensity, which has led to spectacular increase in productivity. However, cow fertility has concomitantly severely declined. It is generally assumed that this reduction is primarily due to the negative energy balance of high-producing cows at the peak of lactation. We herein describe the fine-mapping of a major fertility QTL in Nordic Red cattle, and identify a 660-kb deletion encompassing four genes as the causative variant. We show that the deletion is a recessive embryonically lethal mutation. This probably results from the loss of RNASEH2B, which is known to cause embryonic death in mice. Despite its dramatic effect on fertility, 13%, 23% and 32% of the animals carry the deletion in Danish, Swedish and Finnish Red Cattle, respectively. To explain this, we searched for favorable effects on other traits and found that the deletion has strong positive effects on milk yield. This study demonstrates that embryonic lethal mutations account for a non-negligible fraction of the decline in fertility of domestic cattle, and that associated positive effects on milk yield may account for part of the negative genetic correlation. Our study adds to the evidence that structural variants contribute to animal phenotypic variation, and that balancing selection might be more common in livestock species than previously appreciated.     

1H, 13C and 15N backbone and side-chain chemical shift assignments for reduced unusual thioredoxin Patrx2 of Pseudomonas aeruginosa

The gram-negative organism Pseudomonas aeruginosa is an opportunistic human pathogen and a leading cause of hospital-acquired infections. In P. aeruginosa PAO1, three cytoplasmic thioredoxins have been identified. An unusual thioredoxin (Patrx2) (108 amino acids) encoded by the PA2694 gene, is identified as a new thioredoxin-like protein based on sequence homology. Thioredoxin is a ubiquitous protein, which serves as a general protein disulfide oxidoreductase. Patrx2 present an atypical active site CGHC. We report the nearly complete ¹H, ¹³C and ¹⁵N resonance assignments of reduced Patrx2. 2D and 3D heteronuclear NMR experiments were performed with uniformly ¹⁵N-, ¹³C-labelled Patrx2, resulting in 97.2 % backbone and 92.5 % side-chain ¹H, ¹³C and ¹⁵N resonance assignments for the reduced form. (BMRB deposits with accession number 18130).