A study on nitrate reductase activity (NRA) of geophytes from Mediterranean environment

Nitrate reductase activity (NRA) in different compartments of 14 Mediterranean geophytes (bulbous, tuberous and rhizomatous) and actual mineral nitrogen (NO₃⁻ and NH₄⁺) in their soils were investigated. The nitrate reduction capacities of each species were determined as NRA per total plant material. Differences among compartments for NRA were significant in all species. The highest NRA was found in leaves of tuberous species (Anemone coronaria, Cyclamen coum) and of most bulbous species (Allium flavum, Allium guttatum, Bellevelia sarmatica, Galanthus plicatus, Leucojum aestivum, Ornithogalum nutans, Tulipa sylvestris). Therefore, in this group of species the contribution of the leaves to total plant NRA was the highest. The other bulbous species (Allium scorodoprasum, Crocus chrysanthus, Fritillaria bithynica, Muscari neglectum) and one rhizomatous taxon (Iris suaveolens) have a different NRA distribution within the plants. In these species the highest values of NRA were found in different organs. For example, in Allium scorodoprasum the highest NRA was in tunics, and in flowers in M. neglectum. Although leaves are the main compartments reducing nitrate in most of the studied geophytes, other compartments also contribute to total plant nitrate reduction.Our results show that the nitrate reduction capacity is different among geophyte species. Even if it roughly reflects the nitrogen supply in a habitat, differences in nitrate reduction capacities of different species collected from same sites indicate that the nitrate reducing capacity is species-specific.     

Association of Y chromosome haplogroup I with HIV progression, and HAART outcome

The host genetic basis of differential outcomes in HIV infection, progression, viral load set point and highly active retroviral therapy (HAART) responses was examined for the common Y haplogroups in European Americans and African Americans. Accelerated progression to acquired immune deficiency syndrome (AIDS) and related death in European Americans among Y chromosome haplogroup I (Y-I) subjects was discovered. Additionally, Y-I haplogroup subjects on HAART took a longer time to HIV-1 viral suppression and were more likely to fail HAART. Both the accelerated progression and longer time to viral suppression results observed in haplogroup Y-I were significant after false-discovery-rate corrections. A higher frequency of AIDS-defining illnesses was also observed in haplogroup Y-I. These effects were independent of the previously identified autosomal AIDS restriction genes. When the Y-I haplogroup subjects were further subdivided into six I subhaplogroups, no one subhaplogroup accounted for the effects on HIV progression, viral load or HAART response. Adjustment of the analyses for population stratification found significant and concordant haplogroup Y-I results. The Y chromosome haplogroup analyses of HIV infection and progression in African Americans were not significant. Our results suggest that one or more loci on the Y chromosome found on haplogroup Y-I have an effect on AIDS progression and treatment responses in European Americans. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Biodegradation performance of an anaerobic hybrid reactor treating olive mill effluent under various organic loading rates

The primary objective of this study was to evaluate the performance of a 20 l lab scale anaerobic hybrid reactor (AHR) combining sludge blanket in the lower part and filter in the upper part under varying organic loading rates (OLRs) in order to study biodegradation of olive mill effluent (OME). For this purpose, some parameters, such as total phenols, effluent chemical oxygen demand (COD), suspended solids (SS), volatile fatty acids (VFAs), and pH in the influent and effluent, and removal efficiencies for those parameters (except pH) were continuously monitored throughout the experimental period of 477 days. Eleven different organic loadings between 0.45 and 32 kg COD m⁻³ day⁻¹ were imposed by either varying influent COD or hydraulic retention time (HRT). The results demonstrated that the AHR reactor could tolerate high influent COD concentrations. Removal efficiencies for the studied pollution parameters were found to be as follows: COD, 50–94%; total phenol, 39–80%; color, 0–54%; and suspended solids, 19–87%. The levels of VFAs in the effluent, which was principally acetate, butyrate, iso-butyrate, and propionate, varied between 10 and 2005 mg l⁻¹ depending upon OLRs. A COD removal efficiency of 90% could be achieved as long as OLR is kept at a level of less than 10 kg COD m⁻³ day⁻¹. However, a secondary treatment unit for polishing purposes is necessary to comply with receiving media discharge standards.     

Quantifying Interactions Within the NADP(H) Enzyme Network in Drosophila melanogaster

In this report, we use synthetic, activity-variant alleles in Drosophila melanogaster to quantify interactions across the enzyme network that reduces nicotinamide adenine dinucleotide phosphate (NADP) to NADPH. We examine the effects of large-scale variation in isocitrate dehydrogenase (IDH) or glucose-6-phosphate dehydrogenase (G6PD) activity in a single genetic background and of smaller-scale variation in IDH, G6PD, and malic enzyme across 10 different genetic backgrounds. We find significant interactions among all three enzymes in adults; changes in the activity of any one source of a reduced cofactor generally result in changes in the other two, although the magnitude and directionality of change differs depending on the gene and the genetic background. Observed interactions are presumably through cellular mechanisms that maintain a homeostatic balance of NADPH/NADP, and the magnitude of change in response to modification of one source of reduced cofactor likely reflects the relative contribution of that enzyme to the cofactor pool. Our results suggest that malic enzyme makes the largest single contribution to the NADPH pool, consistent with the results from earlier experiments in larval D. melanogaster using naturally occurring alleles. The interactions between all three enzymes indicate functional interdependence and underscore the importance of examining enzymes as components of a network.IN traits determined by a network of gene products, the phenotype is a function of the alleles present and of the relative contributions of individual network member genes. Since selection is on phenotype, the total composite genotype, not just individual loci, determines the fitness of an organism. In establishing the connection between genotype and phenotype for such networks, the first challenge is to quantify the relative contribution of each member of the network to the endpoint phenotype. By addressing function on a network-wide basis, interactions and interconnections that may not be apparent in individual gene examinations can be determined (Proulx et al. 2005).In most organisms, reduction of the cofactor nicotinamide adenine dinucleotide phosphate, or NADP, to NADPH is primarily the function of four enzymes: cytosolic malic enzyme (MEN), cytosolic isocitrate dehydrogenase (IDH), and the two oxidative enzymes of the pentose shunt, glucose-6-phosphate dehydrogenase (G6PD) and 6-phosphogluconate (6PGD; recently reviewed in Ying 2008). In larval Drosophila melanogaster, MEN produces ∼30% of the available NADPH, IDH ∼20%, and G6PD and 6PGD the remaining ∼40% (Geer et al. 1979a,b). It is believed that these four enzymes interact to maintain the NADP/NADPH balance and supply of reducing power for lipogenesis and antioxidation (Geer et al. 1976, 1978, 1981; Wilton et al. 1982; Bentley et al. 1983; Geer and Laurie-Ahlberg 1984; Merritt et al. 2005; Pollak et al. 2007; Singh et al. 2007; Ying 2008). Dietary induction studies and observations of natural genetic variation have found connections between MEN activity and the activities of the pentose shunt enzymes to be generally straightforward and compensatory; reductions in one lead to increases in the other. The interactions involving IDH activity, however, have been found to be more complicated and at times counterintuitive; reductions in reducing power sometimes lead to decreases in IDH activity.In an earlier study (Merritt et al. 2005), we quantified the impact of genetic variation in Men activity on IDH and G6PD activities and triglyceride (a strong correlate with total lipid; Clark and Keith 1989) concentration. 6PGD was not independently assayed because earlier works suggest that G6PD and 6PGD activities are highly correlated, likely because of their coupled function in the pentose shunt (Wilton et al. 1982). We examined both naturally occurring Men alleles and synthetic alleles created by P-element excision and found significant associations between MEN activity and induction of the activities of both IDH and G6PD. The apparent interactions between MEN and IDH and G6PD across these 10 different third chromosome lines were quantified as mean elasticity coefficients: = −0.76 ± 0.236 and = −0.88 ± 0.208. Because MEN activity was reduced by 20%, both IDH and G6PD activity varied in a compensatory direction, increasing almost 1:1 with the decrease in MEN.The significant change in enzyme activity of two members of the NADPH network in response to our genetic reduction of the activity of a third strongly suggests that a physiological mechanism coregulates the three enzymes. Such functional interdependence would mean that individual members of the network do not act in isolation and should be examined collectively, not as isolated units. In this study, we characterize the effects of the independently varying activity levels of IDH, G6PD, and MEN on the activity of each other and triglyceride concentration in adult flies. We found significant responses to changes in all three enzymes, although the responses to genetic changes in IDH and G6PD were generally small; variation in MEN caused the greatest changes in the other enzymes.     

Theoretical studies on the molecular structure and vibrational spectra of some dimethyl substituted pyridine derivatives

The molecular geometries, normal mode frequencies, intensities and corresponding infrared assignments of monomeric and dimeric 2,3-dimethylpyridine, 2,4-dimethylpyridine, 3,4-dimethylpyridine, 3,5-dimethylpyridine and monomeric 2,6-dimethylpyridine in the ground state were investigated at the density functional theory (DFT)-B3LYP level using the 6-311+G(d, p) basis set. The vibrational frequencies and geometric parameters of C–H stretching and bending in the fundamental region were calculated and compared to the Fourier transform infrared (FT-IR) data obtained. In the studied monomeric and dimeric dimethyl substituted pyridine derivatives, the C–H stretching and bending frequency shifts that occur between the dimer and the monomer may be diagnostic of the magnitude of dimerization energy. As supported by data in the literature, the most stable dimeric form was obtained for the 3,4-dimethylpyridine molecule. Figure Molecular model and numbering scheme of the studied dimeric dimethylpyridinederivatives     

The basic domain of TRF2 directs binding to DNA junctions irrespective of the presence of TTAGGG repeats

The replication of long tracts of telomeric repeats may require specific factors to avoid fork regression (Fouché, N., Ozgür, S., Roy, D., and Griffith, J. (2006) Nucleic Acids Res., in press). Here we show that TRF2 binds to model replication forks and four-way junctions in vitro in a structure-specific but sequence-independent manner. A synthetic peptide encompassing the TRF2 basic domain also binds to DNA four-way junctions, whereas the TRF2 truncation mutant (TRF2(DeltaB)) and a mutant basic domain peptide do not. In the absence of the basic domain, the ability of TRF2 to localize to model telomere ends and facilitate t-loop formation in vitro is diminished. We propose that TRF2 plays a key role during telomere replication in binding chickenfoot intermediates of telomere replication fork regression. Junction-specific binding would also allow TRF2 to stabilize a strand invasion structure that is thought to exist at the strand invasion site of the t-loop.     

Role of G proteins and ERK activation in hemin-induced erythroid differentiation of K562 cells

Heterotrimeric G proteins which couple extracellular signals to intracellular effectors play a central role in cell growth and differentiation. The pluripotent erythroleukemic cell line K562 that acquires the capability to synthesize hemoglobin in response to a variety of agents can be used as a model system for erythroid differentiation. Using Western blot analysis and RT-PCR, we studied alterations in G protein expression accompanying hemin-induced differentiation of K562 cells. We demonstrated the presence of G(alpha s), G(alpha i2) and G(alpha q) and the absence of G(alpha i1), G(alpha o) and G(alpha 16) in K562 cells. We observed the short form of G(alpha s) to be expressed predominantly in these cells. Treatment of K562 cells with hemin resulted in an increase in the levels of G(alpha s) and G(alpha q). On the other hand, the level of G(alpha i2) was found to increase on the third day after induction with hemin, followed by a decrease to levels lower of those of uninduced cells. The mitogen-activated protein kinase ERK1/2 pathway is crucial in the control of cell proliferation and differentiation. Both Gi- and Gq-coupled receptors stimulate MAPK activation. We therefore examined the phosphorylation of ERK1/2 during hemin-induced differentiation of K562 cells. Using anti-ERK1/2 antibodies, we observed that ERK2 was primarily phosphorylated in K562 cells. ERK2 phosphorylation increased gradually until 48 h and returned to basal values by 96 h following hemin treatment. Our results suggest that changes in G protein expression and ERK2 activity are involved in hemin-induced differentiation of K562 cells.     

Exosome secreted by MSC reduces myocardial ischemia/reperfusion injury

Human ESC-derived mesenchymal stem cell (MSC)-conditioned medium (CM) was previously shown to mediate cardioprotection during myocardial ischemia/reperfusion injury through large complexes of 50–100 nm. Here we show that these MSCs secreted 50- to 100-nm particles. These particles could be visualized by electron microscopy and were shown to be phospholipid vesicles consisting of cholesterol, sphingomyelin, and phosphatidylcholine. They contained coimmunoprecipitating exosome-associated proteins, e.g., CD81, CD9, and Alix. These particles were purified as a homogeneous population of particles with a hydrodynamic radius of 55–65 nm by size-exclusion fractionation on a HPLC. Together these observations indicated that these particles are exosomes. These purified exosomes reduced infarct size in a mouse model of myocardial ischemia/reperfusion injury. Therefore, MSC mediated its cardioprotective paracrine effect by secreting exosomes. This novel role of exosomes highlights a new perspective into intercellular mediation of tissue injury and repair, and engenders novel approaches to the development of biologics for tissue repair.     

Persistent organic pollutants in Pakistan: Potential threat to ecological integrities in terms of genotoxicity and oxidative stress

As a consequence of both increasing population and industrialization in agro-economic sector, Pakistan has inevitably been confronted by multicomplex environmental challenges. Owing in part to poor regulatory framework, pollution due to persistent organic pollutants (POPs) has caused serious problems throughout the country. Resultantly, extensive use of POPs is causing vigorous deterioration of environment and human health. The current study addresses: (1) the general information on associated ecological effects and toxicity assessment by meta-analysis for local fauna and flora (2) their respective occurrence in living organisms; and (3) sources and distribution patterns of various POPs classes in environmental compartments of Pakistan. Based on the study, it can be concluded that the environment of Pakistan is highly contaminated with organochlorine pesticides (OCPs), polychlorinated biphenyls (PCBs), polybrominated diphenyl ethers (PBDEs), dechlorane plus (DP), and polychlorinated naphthalenes (PCNs), which is further supported with the meta-analysis. Nevertheless, unavailability of environmental quality standards and food safety for POPs render it a forthcoming challenge of multicompartment toxicity exposure. Therefore, strategies must be planned for risk assessment of biologically active POPs, while the POP waste inventory should be elevated, along with the necessary measures to promote appropriate handling and treatment of POP as a matter of prime importance.