Transbilayer movement of Glc-P-dolichol and its function as a glucosyl donor: protein-mediated transport of a water-soluble analog into sealed ER vesicles from pig brain

The results described in the accompanying article support the model in which glucosylphosphoryldolichol (Glc-P-Dol) is synthesized on the cytoplasmic face of the ER, and functions as a glucosyl donor for three Glc-P-Dol:Glc0-2Man9-GlcNAc2-P-P-Dol glucosyltransferases (GlcTases) in the lumenal compartment. In this study, the enzymatic synthesis and structural characterization by NMR and electrospray-ionization tandem mass spectrometry of a series of water-soluble beta-Glc-P-Dol analogs containing 2-4 isoprene units with either the cis - or trans - stereoconfiguration in the beta-position are described. The water- soluble analogs were (1) used to examine the stereospecificity of the Glc-P-Dol:Glc0-2Man9GlcNAc2-P-P-Dol glucosyltransferases (GlcTases) and (2) tested as potential substrates for a membrane protein(s) mediating the transbilayer movement of Glc-P-Dol in sealed ER vesicles from rat liver and pig brain. The Glc-P-Dol-mediated GlcTases in pig brain microsomes utilized [3H]Glc-labeled Glc-P-Dol10, Glc-P-(omega, c )Dol15, Glc-P(omega, t,t )Dol20, and Glc-P-(omega, t,c )Dol20as glucosyl donors with [3H]Glc3Man9GlcNAc2-P-P-Dol the major product labeled in vitro. A preference was exhibited for C15-20 substrates containing an internal cis -isoprene unit in the beta-position. In addition, the water-soluble analog, Glc-P-Dol10, was shown to enter the lumenal compartment of sealed microsomal vesicles from rat liver and pig brain via a protein-mediated transport system enriched in the ER. The properties of the ER transport system have been characterized. Glc- P-Dol10was not transported into or adsorbed by synthetic PC-liposomes or bovine erythrocytes. The results of these studies indicate that (1) the internal cis -isoprene units are important for the utilization of Glc-P-Dol as a glucosyl donor and (2) the transport of the water- soluble analog may provide an experimental approach to assay the hypothetical "flippase" proposed to mediate the transbilayer movement of Glc-P-Dol from the cytoplasmic face of the ER to the lumenal monolayer.      

Degradation of Alyssum murale biomass in soil

The Ni-hyperaccumulating plant Alyssum murale accumulates exceptionally high concentrations of nickel in its aboveground biomass. The reasons for hyperaccumulation remain unproven; however, it has been proposed that elemental alelopathy might be important. High-Ni leaves shed by the plant may create a "toxic zone" around the plant where germination or growth of competing plants is inhibited. The efficacy of this argument will partially depend upon the rate at which leaves degrade in soil and free metals are released, and the subsequent rate at which metals are bound to soil constituents. To test the degradation of biomass of hyperaccumulators, A. murale was grown on both high- and low-Ni soils to achieve high- (12.0 g Ni/kg) and low- (0.445 g Ni/kg) Ni biomass. Shredded leaf and stem biomass were added to a serpentine soil from Oregon that was originally used to grow high-Ni biomass and a low-Ni control soil from Maryland. Biomass Ni was readily soluble and extractable, suggesting near immediate release as biomass was added to soil Extractable nickel in soil amended with biomass declined rapidly over time due to Ni binding in soil These results suggest that Ni released from biomass of Ni hyperaccumulators may significantly affect their immediate niche only for short periods of time soon after leaf fall, but repeated application may create high Ni levels under and around hyperaccumulators.     

The effects of long-term endurance training on the immune and endocrine systems of elderly men: the role of cytokines and anabolic hormones

Background  

a decline in immune and endocrine function occurs with aging. The main purpose of this study was to investigate the impact of long-term endurance training on the immune and endocrine system of elderly men. The possible interaction between these systems was also analysed.     

MyD88-Dependent Signaling Influences Fibrosis and Alternative Macrophage Activation during Staphylococcus aureus Biofilm Infection

Bacterial biofilms represent a significant therapeutic challenge based on their ability to evade host immune and antibiotic-mediated clearance. Recent studies have implicated IL-1β in biofilm containment, whereas Toll-like receptors (TLRs) had no effect. This is intriguing, since both the IL-1 receptor (IL-1R) and most TLRs impinge on MyD88-dependent signaling pathways, yet the role of this key adaptor in modulating the host response to biofilm growth is unknown. Therefore, we examined the course of S. aureus catheter-associated biofilm infection in MyD88 knockout (KO) mice. MyD88 KO animals displayed significantly increased bacterial burdens on catheters and surrounding tissues during early infection, which coincided with enhanced dissemination to the heart and kidney compared to wild type (WT) mice. The expression of several proinflammatory mediators, including IL-6, IFN-γ, and CXCL1 was significantly reduced in MyD88 KO mice, primarily at the later stages of infection. Interestingly, immunofluorescence staining of biofilm-infected tissues revealed increased fibrosis in MyD88 KO mice concomitant with enhanced recruitment of alternatively activated M2 macrophages. Taken in the context of previous studies with IL-1β, TLR2, and TLR9 KO mice, the current report reveals that MyD88 signaling is a major effector pathway regulating fibrosis and macrophage polarization during biofilm formation. Together these findings represent a novel example of the divergence between TLR and MyD88 action in the context of S. aureus biofilm infection.     

Research Priorities for Conservation of Metallophyte Biodiversity and their Potential for Restoration and Site Remediation

Plants that have evolved to survive on metal‐rich soils—metallophytes—have key values that must drive research of their unique properties and ultimately their conservation. The ability of metallophytes to tolerate extreme metal concentrations commends them for revegetation of mines and metal‐contaminated sites. Metallophytes can also be exploited in environmental technologies, for example, phytostabilization, phytoremediation, and phytomining. Actions towards conserving metallophyte species are imperative, as metallophytes are increasingly under threat of extinction from mining activity. Although many hundreds of papers describe both the biology and applications of metallophytes, few have investigated the urgent need to conserve these unique species. This paper identifies the current state of metallophyte research, and advocates future research needs for the conservation of metallophyte biodiversity and the sustainable uses of metallophyte species in restoration, rehabilitation, contaminated site remediation, and other nascent phytotechnologies. Six fundamental questions are addressed: (1) Is enough known about the global status of metallophytes to ensure their conservation? (2) Are metallophytes threatened by the activities of the minerals industry, and can their potential for the restoration or rehabilitation of mined and disturbed land be realized? (3) What problems exist in gaining prior informed consent to access metallophyte genetic resources and how can the benefits arising from their uses be equitably shared? (4) What potential do metallophytes offer as a resource base for phytotechnologies? (5) Can genetic modification be used to “design” metallophytes to use in the remediation of contaminated land? (6) Does the prospect of using metallophytes in site remediation and restoration raise ethical issues?     

Leaching of Pseudomonas aeruginosa, and transconjugants containing pR68.45 through unsaturated, intact soil columns

Abstract Leaching of genetically engineered microbes (GEMs) through soil is a significant concern related to groundwater quality. The objective of this study was to examine the leaching, survival and gene transfer of a genetically engineered microbe and indigenous recipients of pR68.45 in nonsterile, undisturbed soil columns. Pseudomonas aeruginosa PAO25, containing the plasmid R68.45, was added to the surface of undisturbed soil columns (10 cm diameter × 80 cm length). Unsaturated flow conditions were maintained by 100 ml daily additions of 2 mM CaCl₂ for a period of 70 days. The population of the GEM exhibited a significant ( P = 0.05) linear decline with time. The GEM leached only to a depth of 30–40 cm in 70 days. Transfer of pR68.45 was shown to occur from P. aeruginosa into the indigenous bacterial population although relatively low numbers of transconjugants were observed (log 2 cfu g⁻¹ dry soil). The number of transconjugants also decreased with depth and time. Leaching of transconjugants, however, occured more readily than that of the GEM, probably as a result of plasmid transfer into smaller, more mobile bacteria. At 70 days incubation, no GEMs were detected in the columns, while transconjugants were observed at several depths. These results demonstrate the importance of examining both the survival and movement of GEMs and transconjugants in soil.     

An improved understanding of soil Cd risk to humans and low cost methods to phytoextract Cd from contaminated soils to prevent soil Cd risks

We believe greater consideration should be given the agronomic and nutritional/bioavailability factors that influence risk from Cd-contaminated soils. We have argued that the ability of rice to accumulate soil Cd in grain while excluding Fe, Zn and Ca (even though the soil contains 100-times more Zn than Cd) was important in adverse effects of soil Cd is farm families in Asia. Further, polished rice grain is deficient in Fe, Zn and Ca for humans, which promotes Cd absorption into duodenal cells. New kinetic studies clarified that dietary Cd is absorbed into duodenum enterocytes; 109Cd from a single meal remained in the duodenum for up to 16 days; part of the turnover pool 109Cd moved to the liver and kidneys by the end of the 64-day 'chase' period. Thus malnutrition induced by subsistence rice diets caused a higher absorption of dietary Cd and much higher potential risk from soil Cd than other crops. Because rice-induced Fe-Zn-Ca-malnutrition is so important in soil Cd risk, it seems evident that providing nutritional supplements to populations of exposed subsistence rice farmers could protect them against soil Cd during a period of soil remediation. In the long term, high Cd rice soils need to be remediated. Remediation by removal and replacement of contaminated soil is very expensive (on the order of $3 million/ha); while phytoextraction using the high Cd accumulating ecotypes of the Zn-Cd hyperaccumulator, Thlaspi caerulescens, should provide low cost soil Cd remediation.