OPT3 Is a Phloem-Specific Iron Transporter That Is Essential for Systemic Iron Signaling and Redistribution of Iron and Cadmium in Arabidopsis

Iron is essential for both plant growth and human health and nutrition. Knowledge of the signaling mechanisms that communicate iron demand from shoots to roots to regulate iron uptake as well as the transport systems mediating iron partitioning into edible plant tissues is critical for the development of crop biofortification strategies. Here, we report that OPT3, previously classified as an oligopeptide transporter, is a plasma membrane transporter capable of transporting transition ions in vitro. Studies in Arabidopsis thaliana show that OPT3 loads iron into the phloem, facilitates iron recirculation from the xylem to the phloem, and regulates both shoot-to-root iron signaling and iron redistribution from mature to developing tissues. We also uncovered an aspect of crosstalk between iron homeostasis and cadmium partitioning that is mediated by OPT3. Together, these discoveries provide promising avenues for targeted strategies directed at increasing iron while decreasing cadmium density in the edible portions of crops and improving agricultural productivity in iron deficient soils.     

Glucose Homeostasis Is Important for Immune Cell Viability during Candida Challenge and Host Survival of Systemic Fungal Infection

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Whole-Genome Analysis of a Rare Human Korean G3P Rotavirus Strain Suggests a Complex Evolutionary Origin Potentially Involving Reassortment Events between Feline and Bovine Rotaviruses

A rare human rotavirus, G3P[9] strain RVA/Human-tc/KOR/CAU12-2-51/2013/G3P[9], was isolated from the stool of a 9-year-old female hospitalized with acute watery diarrhea in August 2012 in South Korea using a cell culture system, and its genome was analyzed. The complete genomic constellation of the CAU12-2-51 strain revealed a novel genotype constellation for human rotavirus, G3-P[9]-I2-R2-C2-M2-A3-N2-T3-E3-H3. Phylogenetic analysis revealed that the CAU12-2-51 strain originated from feline- and bovine-like reassortment strains. The genes encoding VP4, VP7, NSP1, NSP3, NSP4, and NSP5 were related to human/feline-like and feline rotavirus strains, whereas the remaining five genes encoding VP1, VP2, VP3, VP6, and NSP2 were related to the human/bovine-like and bovine rotavirus strains. This novel strain was identified for the first time, providing evidence of feline/bovine-to-human transmission of rotavirus. The data presented herein provide information regarding rotavirus diversity and evolution.     

Local and Systemic Regulation of Plant Root System Architecture and Symbiotic Nodulation by a Receptor-Like Kinase

In plants, root system architecture is determined by the activity of root apical meristems, which control the root growth rate, and by the formation of lateral roots. In legumes, an additional root lateral organ can develop: the symbiotic nitrogen-fixing nodule. We identified in Medicago truncatula ten allelic mutants showing a compact root architecture phenotype (cra2) independent of any major shoot phenotype, and that consisted of shorter roots, an increased number of lateral roots, and a reduced number of nodules. The CRA2 gene encodes a Leucine-Rich Repeat Receptor-Like Kinase (LRR-RLK) that primarily negatively regulates lateral root formation and positively regulates symbiotic nodulation. Grafting experiments revealed that CRA2 acts through different pathways to regulate these lateral organs originating from the roots, locally controlling the lateral root development and nodule formation systemically from the shoots. The CRA2 LRR-RLK therefore integrates short- and long-distance regulations to control root system architecture under non-symbiotic and symbiotic conditions.     

Macromolecules Released by a Plant Growth-promoting Rhizobacterium as Elicitors of Systemic Resistance in Tomato to Bacterial and Fungal Pathogens

One of 500 rhizobacteria isolated from soil, rhizosphere and rhizoplane of healthy tomato plants was previously selected in laboratory, greenhouse and field tests as a good inducer of systemic resistance. This plant growth‐promoting rhizobacterium (PGPR) was identified as Bacillus cereus by fatty‐acid analysis. Bacillus cereus bacterial cells were removed from liquid culture by centrifugation and the supernatant repeatedly dialyzed (cut‐off = 12 000 daltons) against distilled water. Dialysates applied to roots protected tomato plants against leaf fungal and bacterial pathogens, evidence that macromolecules synthesized by the PGPR and released into the environment act as elicitors of systemic resistance.     

Gcn4-Mediator Specificity Is Mediated by a Large and Dynamic Fuzzy Protein-Protein Complex

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Intra-Operative Tissue Oxygen Tension Is Increased by Local Insufflation of Humidified-Warm CO2 during Open Abdominal Surgery in a Rat Model

Introduction

Maintenance of high tissue oxygenation (PtO₂) is recommended during surgery because PtO₂ is highly predictive of surgical site infection and colonic anastomotic leakage. However, surgical site perfusion is often sub-optimal, creating an obstructive hurdle for traditional, systemically applied therapies to maintain or increase surgical site PtO_2. This research tested the hypothesis that insufflation of humidified-warm CO₂ into the abdominal cavity would increase sub-peritoneal PtO₂ during open abdominal surgery.

Materials and Methods

15 Wistar rats underwent laparotomy under general anesthesia. Three sets of randomized cross-over experiments were conducted in which the abdominal cavity was subjected to alternating exposure to 1) humidified-warm CO₂ & ambient air; 2) humidified-warm CO₂ & dry-cold CO₂; and 3) dry-cold CO₂ & ambient air. Sub-peritoneal PtO₂ and tissue temperature were measured with a polarographic oxygen probe.

Results

Upon insufflation of humidified-warm CO₂, PtO₂ increased by 29.8 mmHg (SD 13.3; p<0.001), or 96.6% (SD 51.9), and tissue temperature by 3.0°C (SD 1.7 p<0.001), in comparison with exposure to ambient air. Smaller, but significant, increases in PtO₂ were seen in experiments 2 and 3. Tissue temperature decreased upon exposure to dry-cold CO₂ compared with ambient air (-1.4°C, SD 0.5, p = 0.001).

Conclusions

In a rat model, insufflation of humidified-warm CO₂ into the abdominal cavity during open abdominal surgery causes an immediate and potentially clinically significant increase in PtO₂. The effect is an additive result of the delivery of CO₂ and avoidance of evaporative cooling via the delivery of the CO₂ gas humidified at body temperature.     

Trypanosoma cruzi Infection Is a Potent Risk Factor for Non-alcoholic Steatohepatitis Enhancing Local and Systemic Inflammation Associated with Strong Oxidative Stress and Metabolic Disorders

Background

The immune mechanisms underlying experimental non-alcoholic steatohepatitis (NASH), and more interestingly, the effect of T. cruzi chronic infection on the pathogenesis of this metabolic disorder are not completely understood.

Methodology/Principal Findings

We evaluated immunological parameters in male C57BL/6 wild type and TLR4 deficient mice fed with a standard, low fat diet, LFD (3% fat) as control group, or a medium fat diet, MFD (14% fat) in order to induce NASH, or mice infected intraperitoneally with 100 blood-derived trypomastigotes of Tulahuen strain and also fed with LFD (I+LFD) or MFD (I+MFD) for 24 weeks. We demonstrated that MFD by itself was able to induce NASH in WT mice and that parasitic infection induced marked metabolic changes with reduction of body weight and steatosis revealed by histological studies. The I+MFD group also improved insulin resistance, demonstrated by homeostasis model assessment of insulin resistance (HOMA-IR) analysis; although parasitic infection increased the triglycerides and cholesterol plasma levels. In addition, hepatic M1 inflammatory macrophages and cytotoxic T cells showed intracellular inflammatory cytokines which were associated with high levels of IL6, IFNγ and IL17 plasmatic cytokines and CCL2 chemokine. These findings correlated with an increase in hepatic parasite load in I+MFD group demonstrated by qPCR assays. The recruitment of hepatic B lymphocytes, NK and dendritic cells was enhanced by MFD, and it was intensified by parasitic infection. These results were TLR4 signaling dependent. Flow cytometry and confocal microscopy analysis demonstrated that the reactive oxygen species and peroxinitrites produced by liver inflammatory leukocytes of MFD group were also exacerbated by parasitic infection in our NASH model.

Conclusions

We highlight that a medium fat diet by itself is able to induce steatohepatitis. Our results also suggest a synergic effect between damage associated with molecular patterns generated during NASH and parasitic infection, revealing an intense cross-talk between metabolically active tissues, such as the liver, and the immune system. Thus, T. cruzi infection must be considered as an additional risk factor since exacerbates the inflammation and accelerates the development of hepatic injury.     

Inactivation of Escherichia coli by Polychromatic Simulated Sunlight: Evidence for and Implications of a Fenton Mechanism Involving Iron,Hydrogen Peroxide,and Superoxide

Sunlight inactivation of Escherichia coli has previously been shown to accelerate in the presence of oxygen, exogenously added hydrogen peroxide, and bioavailable forms of exogenously added iron. In this study, mutants unable to effectively scavenge hydrogen peroxide or superoxide were found to be more sensitive to polychromatic simulated sunlight (without UVB wavelengths) than wild-type cells, while wild-type cells grown under low-iron conditions were less sensitive than cells grown in the presence of abundant iron. Furthermore, prior exposure to simulated sunlight was found to sensitize cells to subsequent hydrogen peroxide exposure in the dark, but this effect was attenuated for cells grown with low iron. Mutants deficient in recombination DNA repair were sensitized to simulated sunlight (without UVB wavelengths), but growth in the presence of iron chelators reduced the degree of sensitization conferred by this mutation. These findings support the hypothesis that hydrogen peroxide, superoxide, and intracellular iron all participate in the photoinactivation of E. coli and further suggest that the inactivation rate of enteric bacteria in the environment may be strongly dependent on iron availability and growth conditions.     

Accumulation of Salicylic Acid and 4-Hydroxybenzoic Acid in Phloem Fluids of Cucumber during Systemic Acquired Resistance Is Preceded by a Transient Increase in Phenylalanine Ammonia-Lyase Activity in Petioles and Stems

Cucumber (Cucumis sativa) leaves infiltrated with Pseudomonas syringae pv. syringae cells produced a mobile signal for systemic acquired resistance between 3 and 6 h after inoculation. The production of a mobile signal by inoculated leaves was followed by a transient increase in phenylalanine ammonia-lyase (PAL) activity in the petioles of inoculated leaves and in stems above inoculated leaves; with peaks in activity at 9 and 12 h, respectively, after inoculation. In contrast, PAL activity in inoculated leaves continued to rise slowly for at least 18 h. No increases in PAL activity were detected in healthy leaves of inoculated plants. Two benzoic acid derivatives, salicylic acid (SA) and 4-hydroxybenzoic acid (4HBA), began to accumulate in phloem fluids at about the time PAL activity began to increase, reaching maximum concentrations 15 h after inoculation. The accumulation of SA and 4HBA in phloem fluids was unaffected by the removal of all leaves 6 h after inoculation, and seedlings excised from roots prior to inoculation still accumulated high levels of SA and 4HBA. These results suggest that SA and 4HBA are synthesized de novo in stems and petioles in response to a mobile signal from the inoculated leaf.     

Planktonic Replication Is Essential for Biofilm Formation by Legionella pneumophila in a Complex Medium under Static and Dynamic Flow Conditions

Legionella pneumophila persists for a long time in aquatic habitats, where the bacteria associate with biofilms and replicate within protozoan predators. While L. pneumophila serves as a paradigm for intracellular growth within protozoa, it is less clear whether the bacteria form or replicate within biofilms in the absence of protozoa. In this study, we analyzed surface adherence of and biofilm formation by L. pneumophila in a rich medium that supported axenic replication. Biofilm formation by the virulent L. pneumophila strain JR32 and by clinical and environmental isolates was analyzed by confocal microscopy and crystal violet staining. Strain JR32 formed biofilms on glass surfaces and upright polystyrene wells, as well as on pins of “inverse” microtiter plates, indicating that biofilm formation was not simply due to sedimentation of the bacteria. Biofilm formation by an L. pneumophila fliA mutant lacking the alternative sigma factor σ²⁸ was reduced, which demonstrated that bacterial factors are required. Accumulation of biomass coincided with an increase in the optical density at 600 nm and ceased when the bacteria reached the stationary growth phase. L. pneumophila neither grew nor formed biofilms in the inverse system if the medium was exchanged twice a day. However, after addition of Acanthamoeba castellanii, the bacteria proliferated and adhered to surfaces. Sessile (surface-attached) and planktonic (free-swimming) L. pneumophila expressed β-galactosidase activity to similar extents, and therefore, the observed lack of proliferation of surface-attached bacteria was not due to impaired protein synthesis or metabolic activity. Cocultivation of green fluorescent protein (GFP)- and DsRed-labeled L. pneumophila led to randomly interspersed cells on the substratum and in aggregates, and no sizeable patches of clonally growing bacteria were observed. Our findings indicate that biofilm formation by L. pneumophila in a rich medium is due to growth of planktonic bacteria rather than to growth of sessile bacteria. In agreement with this conclusion, GFP-labeled L. pneumophila initially adhered in a continuous-flow chamber system but detached over time; the detachment correlated with the flow rate, and there was no accumulation of biomass. Under these conditions, L. pneumophila persisted in biofilms formed by Empedobacter breve or Microbacterium sp. but not in biofilms formed by Klebsiella pneumoniae or other environmental bacteria, suggesting that specific interactions between the bacteria modulate adherence.     

Early Signaling in Primary T Cells Activated by Antigen Presenting Cells Is Associated with a Deep and Transient Lamellal Actin Network

Cellular signaling transduction critically depends on molecular interactions that are in turn governed by dynamic subcellular distributions of the signaling system components. Comprehensive insight into signal transduction requires an understanding of such distributions and cellular structures driving them. To investigate the activation of primary murine T cells by antigen presenting cells (APC) we have imaged more than 60 signaling intermediates during T cell stimulation with microscopy across resolution limits. A substantial number of signaling intermediates associated with a transient, wide, and actin-associated lamellum extending from an interdigitated T cell:APC interface several micrometers into the T cell, as characterized in detail here. By mapping the more than 60 signaling intermediates onto the spatiotemporal features of cell biological structures, the lamellum and other ones previously described, we also define distinct spatial and temporal characteristics of T cell signal initiation, amplification, and core signaling in the activation of primary T cells by APCs. These characteristics differ substantially from ones seen when T cells are activated using common reductionist approaches.