Active Transport of Phosphorylated Carbohydrates Promotes Intestinal Colonization and Transmission of a Bacterial Pathogen

Efficient acquisition of extracellular nutrients is essential for bacterial pathogenesis, however the identities and mechanisms for transport of many of these substrates remain unclear. Here, we investigate the predicted iron-binding transporter AfuABC and its role in bacterial pathogenesis in vivo. By crystallographic, biophysical and in vivo approaches, we show that AfuABC is in fact a cyclic hexose/heptose-phosphate transporter with high selectivity and specificity for a set of ubiquitous metabolites (glucose-6-phosphate, fructose-6-phosphate and sedoheptulose-7-phosphate). AfuABC is conserved across a wide range of bacterial genera, including the enteric pathogens EHEC O157:H7 and its murine-specific relative Citrobacter rodentium, where it lies adjacent to genes implicated in sugar sensing and acquisition. C. rodentium ΔafuA was significantly impaired in an in vivo murine competitive assay as well as its ability to transmit infection from an afflicted to a naïve murine host. Sugar-phosphates were present in normal and infected intestinal mucus and stool samples, indicating that these metabolites are available within the intestinal lumen for enteric bacteria to import during infection. Our study shows that AfuABC-dependent uptake of sugar-phosphates plays a critical role during enteric bacterial infection and uncovers previously unrecognized roles for these metabolites as important contributors to successful pathogenesis.     

Protective effect of N-acetylcysteine supplementation on mitochondrial oxidative stress and mitochondrial enzymes in cerebral cortex of streptozotocin-treated diabetic rats

Diabetic encephalopathy, characterized by cognitive deficits involves hyperglycemia-induced oxidative stress. Impaired mitochondrial functions might play an important role in accelerated oxidative damage observed in diabetic brain. The aim of the present study was to examine the role of mitochondrial oxidative stress and dysfunctions in the development of diabetic encephalopathy along with the neuroprotective potential of N-acetylcysteine (NAC). Chronic hyperglycemia accentuated mitochondrial oxidative stress in terms of increased ROS production and lipid peroxidation. Significant decrease in Mn-SOD activity along with protein and non-protein thiols was observed in the mitochondria from diabetic brain. The activities of mitochondrial enzymes; NADH dehydrogenase, succinate dehydrogenase and cytochrome oxidase were decreased in the diabetic brain. Increased mitochondrial oxidative stress and dysfunctions were associated with increased cytochrome c and active caspase-3 levels in cytosol. Electron microscopy revealed mitochondrial swelling and chromatin condensation in neurons of diabetic animals. NAC administration, on the other hand was found to significantly improve diabetes-induced biochemical and morphological changes, bringing them closer to the controls. The results from the study provide evidence for the role of mitochondrial oxidative stress and dysfunctions in the development of diabetic encephalopathy and point towards the clinical potential of NAC as an adjuvant therapy to conventional anti-hyperglycemic regimens for the prevention and/or delaying the progression of CNS complications.     

Carbon footprint of spring barley in relation to preceding oilseeds and N fertilization

Purpose

Carbon footprint of field crops can be lowered through improved cropping practices. The objective of this study was to determine the carbon footprint of spring barley (Hordeum vulgare L.) in relation to various preceding oilseed crops that were fertilized at various rates of inorganic N the previous years. System boundary was from cradle-to-farm gate.

Materials and methods

Canola-quality mustard (Brassica juncea L.), canola (Brassica napus L.), sunflower (Helianthus annuus L.), and flax (Linum usitatissimum L.) were grown under the N fertilizer rates of 10, 30, 70, 90, 110, 150, and 200?kg?N?ha^?1 the previous year, and spring barley was grown on the field of standing oilseed stubble the following year. The study was conducted at six environmental sites; they were at Indian Head in 2005, 2006 and 2007, and at Swift Current in 2004, 2005 and 2006, Saskatchewan, Canada.

Results and discussion

On average, barley grown at humid Indian Head emitted greenhouse gases (GHGs) of 1,003?kg?CO₂eq?ha^?1, or 53% greater than that at the drier Swift Current site. Production and delivery of fertilizer N to farm gate accounted for 26% of the total GHG emissions, followed by direct and indirect emissions of 28% due to the application of N fertilizers to barley crop. Emissions due to N fertilization were 26.6 times the emission from the use of phosphorous, 5.2 times the emission from pesticides, and 4.2 times the emission from various farming operations. Decomposition of crop residues contributed emissions of 173?kg?CO₂eq?ha^?1, or 19% of the total emission. Indian Head-produced barley had significantly greater grain yield, resulting in about 11% lower carbon footprint than Swift Current-produced barley (0.28 vs. 0.32?kg?CO₂eq?kg^?1 of grain). Emissions in the barley production was a linear function of the rate of fertilizer N applied to the previous oilseed crops due to increased emissions from crop residue decomposition coupled with higher residual soil mineral N.

Conclusions

The key to lower the carbon footprint of barley is to increase grain yield, make a wise choice of crop types, reduce N inputs to crops grown in the previous and current growing seasons, and improved N use efficiency.     

Mitochondrial DNA-depleted A549 cells are resistant to bleomycin

Alveolar epithelial cells are considered to be the primary target of bleomycin-induced lung injury, leading to interstitial fibrosis. The molecular mechanisms by which bleomycin causes this damage are poorly understood but are suspected to involve generation of reactive oxygen species and DNA damage. We studied the effect of bleomycin on mitochondrial DNA (mtDNA) and nuclear DNA (nDNA) in human alveolar epithelial A549 cells. Bleomycin caused an increase in reactive oxygen species production, DNA damage, and apoptosis in A549 cells; however, bleomycin induced more mtDNA than nDNA damage. DNA damage was associated with activation of caspase-3, cleavage of poly(ADP-ribose) polymerase, and cleavage and activation of protein kinase D1 (PKD1), a newly identified mitochondrial oxidative stress sensor. These effects appear to be mtDNA-dependent, because no caspase-3 or PKD1 activation was observed in mtDNA-depleted (ρ(0)) A549 cells. Survival rate after bleomycin treatment was higher for A549 ρ(0) than A549 cells. These results suggest that A549 ρ(0) cells are more resistant to bleomycin toxicity than are parent A549 cells, likely in part due to the depletion of mtDNA and impairment of mitochondria-dependent apoptotic pathways.     

Targeting intracellular transport combined with efficient uptake and storage significantly increases grain iron and zinc levels in rice

Rice, a staple food for more than half of the world population, is an important target for iron and zinc biofortification. Current strategies mainly focus on the expression of genes for efficient uptake, long‐distance transport and storage. Targeting intracellular iron mobilization to increase grain iron levels has not been reported. Vacuole is an important cell compartment for iron storage and the NATURAL RESISTANCE ASSOCIATED MACROPHAGE PROTEIN (NRAMP) family of transporters export iron from vacuoles to cytosol when needed. We developed transgenic Nipponbare rice lines expressing AtNRAMP3 under the control of the UBIQUITIN or rice embryo/aleurone‐specific 18‐kDa Oleosin (Ole18) promoter together with NICOTIANAMINE SYNTHASE (AtNAS1) and FERRITIN (PvFER), or expressing only AtNRAMP3 and PvFER together. Iron and zinc were increased close to recommended levels in polished grains of the transformed lines, with maximum levels when AtNRAMP3, AtNAS1 and PvFER were expressed together (12.67 μg/g DW iron and 45.60 μg/g DW zinc in polished grains of line NFON16). Similar high iron and zinc levels were obtained in transgenic Indica IR64 lines expressing the AtNRAMP3, AtNAS1 and PvFER cassette (13.65 μg/g DW iron and 48.18 μg/g DW zinc in polished grains of line IR64_1), equalling more than 90% of the recommended iron increase in rice endosperm. Our results demonstrate that targeting intracellular iron stores in combination with iron and zinc transport and endosperm storage is an effective strategy for iron biofortification. The increases achieved in polished IR64 grains are of dietary relevance for human health and a valuable nutrition trait for breeding programmes.     

N-Benzyl-1-heteroaryl-3-(trifluoromethyl)-1H-pyrazole-5-carboxamides as inhibitors of co-activator associated arginine methyltransferase 1 (CARM1)

A series of N-benzyl-1-heteroaryl-3-(trifluoromethyl)-1H-pyrazole-5-carboxamides targeting co-activator associated arginine methyltransferase 1 (CARM1) have been designed and synthesized. The potency of these inhibitors was influenced by the nature of the heteroaryl fragment with the thiophene analogues being superior to thiazole, pyridine, isoindoline and benzofuran based inhibitors.     

Sulfamides as novel histone deacetylase inhibitors

The sulfamide moiety has been utilized to design novel HDAC inhibitors. The potency and selectivity of these inhibitors were influenced both by the nature of the scaffold, and the capping group. Linear long-chain-based analogs were primarily HDAC6-selective, while analogs based on the lysine scaffold resulted in potent HDAC1 and HDAC6 inhibitors.     

Massive colonization of protein-coding exons by selfish genetic elements in Paramecium germline genomes

Ciliates are unicellular eukaryotes with both a germline genome and a somatic genome in the same cytoplasm. The somatic macronucleus (MAC), responsible for gene expression, is not sexually transmitted but develops from a copy of the germline micronucleus (MIC) at each sexual generation. In the MIC genome of Paramecium tetraurelia, genes are interrupted by tens of thousands of unique intervening sequences called internal eliminated sequences (IESs), which have to be precisely excised during the development of the new MAC to restore functional genes. To understand the evolutionary origin of this peculiar genomic architecture, we sequenced the MIC genomes of 9 Paramecium species (from approximately 100 Mb in Paramecium aurelia species to >1.5 Gb in Paramecium caudatum). We detected several waves of IES gains, both in ancestral and in more recent lineages. While the vast majority of IESs are single copy in present-day genomes, we identified several families of mobile IESs, including nonautonomous elements acquired via horizontal transfer, which generated tens to thousands of new copies. These observations provide the first direct evidence that transposable elements can account for the massive proliferation of IESs in Paramecium. The comparison of IESs of different evolutionary ages indicates that, over time, IESs shorten and diverge rapidly in sequence while they acquire features that allow them to be more efficiently excised. We nevertheless identified rare cases of IESs that are under strong purifying selection across the aurelia clade. The cases examined contain or overlap cellular genes that are inactivated by excision during development, suggesting conserved regulatory mechanisms. Similar to the evolution of introns in eukaryotes, the evolution of Paramecium IESs highlights the major role played by selfish genetic elements in shaping the complexity of genome architecture and gene expression.

A comparative genomics study of nine Paramecium species reveals successful invasion of genes by transposable elements in their germline genomes, showing that the internal eliminated sequences (IESs) followed an evolutionary trajectory remarkably similar to that of spliceosomal introns.