The contribution of trees to ecosystem methane emissions in a temperate forested wetland

Wetland‐adapted trees are known to transport soil‐produced methane (CH₄), an important greenhouse gas to the atmosphere, yet seasonal variations and controls on the magnitude of tree‐mediated CH₄ emissions remain unknown for mature forests. We examined the spatial and temporal variability in stem CH₄ emissions in situ and their controls in two wetland‐adapted tree species (Alnus glutinosa and Betula pubescens) located in a temperate forested wetland. Soil and herbaceous plant‐mediated CH₄ emissions from hollows and hummocks also were measured, thus enabling an estimate of contributions from each pathway to total ecosystem flux. Stem CH₄ emissions varied significantly between the two tree species, with Alnus glutinosa displaying minimal seasonal variations, while substantial seasonal variations were observed in Betula pubescens. Trees from each species emitted similar quantities of CH₄ from their stems regardless of whether they were situated in hollows or hummocks. Soil temperature and pore‐water CH₄ concentrations best explained annual variability in stem emissions, while wood‐specific density and pore‐water CH₄ concentrations best accounted for between‐species variations in stem CH₄ emission. Our study demonstrates that tree‐mediated CH₄ emissions contribute up to 27% of seasonal ecosystem CH₄ flux in temperate forested wetland, with the largest relative contributions occurring in spring and winter. Tree‐mediated CH₄ emissions currently are not included in trace gas budgets of forested wetland. Further work is required to quantify and integrate this transport pathway into CH₄ inventories and process‐based models.     

MicroRNA signatures differentiate melanoma subtypes

Melanoma is an aggressive cancer that is highly resistance to therapies once metastasized. We studied microRNA (miRNA) expression in clinical melanoma subtypes and evaluated different miRNA signatures in the background of gain of function somatic and inherited mutations associated with melanoma. Total RNA from 42 patient derived primary melanoma cell lines and three independent normal primary melanocyte cell cultures was evaluated by miRNA array. MiRNA expression was then analyzed comparing subtypes and additional clinicopathologic criteria including somatic mutations. The prevalence and association of an inherited variant in a miRNA binding site in the 3′UTR of the KRAS oncogene, referred to as the KRAS-variant, was also evaluated. We show that seven miRNAs, miR-142-3p, miR-486, miR-214, miR-218, miR-362, miR-650 and miR-31, were significantly correlated with acral as compared to non-acral melanomas (p < 0.04). In addition, we discovered that the KRAS-variant was enriched in non-acral melanoma (25%), and that miR-137 under expression was significantly associated with melanomas with the KRAS-variant. Our findings indicate that miRNAs are differentially expressed in melanoma subtypes and that their misregulation can be impacted by inherited gene variants, supporting the hypothesis that miRNA misregulation reflects biological differences in melanoma.Key words: melanoma, microRNA profiling, biomarker, acral, KRAS-variant, SNP     

An overview on genus garcinia: phytochemical and therapeutical aspects

The genus Garcinia belongs to the family Clusiaceae and has been involved in ayurvedic preparations to medicate various pathophysiological disorders. The bioactive molecules like hydroxycitric acid (HCA), flavonoids, terpenes, polysaccharides, procyanidines and polyisoprenylated benzophenone derivatives like garcinol, xanthochymol and guttiferone isoforms have been isolated from the genus Garcinia. The genus has received the attention of pharmaceutical industries due to their immense remedial qualities. The HCA has been known for its hypolipidemic property. The polyisoprenylated benzophenone and xanthone derivatives are known for their antioxidant, apoptotic, anti-cancer, anti-inflammatory, anti-bacterial, anti-viral, anti-fungal, anti-ulcer, anti-protozoal, and HAT inhibiting properties. Future studies on the synthesis of therapeutically important products and their analogs and evaluation of their safety and efficacy would be of great interest. Though the genus includes more than 300 species, we have made an effort to conceive the curative qualities of bioactive compounds of selected plants to the best of our knowledge.     

Sequence determinants for the reaction specificity of murine (12R)-lipoxygenase: targeted substrate modification and site-directed mutagenesis

Mammalian lipoxygenases (LOXs) are categorized with respect to their positional specificity of arachidonic acid oxygenation. Site-directed mutagenesis identified sequence determinants for the positional specificity of these enzymes, and a critical amino acid for the stereoselectivity was recently discovered. To search for sequence determinants of murine (12R)-LOX, we carried out multiple amino acid sequence alignments and found that Phe(390), Gly(441), Ala(455), and Val(631) align with previously identified positional determinants of S-LOX isoforms. Multiple site-directed mutagenesis studies on Phe(390) and Ala(455) did not induce specific alterations in the reaction specificity, but yielded enzyme species with reduced specific activities and stereo random product patterns. Mutation of Gly(441) to Ala, which caused drastic alterations in the reaction specificity of other LOX isoforms, failed to induce major alterations in the positional specificity of mouse (12R)-LOX, but markedly modified the enantioselectivity of the enzyme. When Val(631), which aligns with the positional determinant Ile(593) of rabbit 15-LOX, was mutated to a less space-filling residue (Ala or Gly), we obtained an enzyme species with augmented catalytic activity and specifically altered reaction characteristics (major formation of chiral (11R)-hydroxyeicosatetraenoic acid methyl ester). The importance of Val(631) for the stereo control of murine (12R)-LOX was confirmed with other substrates such as methyl linoleate and 20-hydroxyeicosatetraenoic acid methyl ester. These data identify Val(631) as the major sequence determinant for the specificity of murine (12R)-LOX. Furthermore, we conclude that substrate fatty acids may adopt different catalytically productive arrangements at the active site of murine (12R)-LOX and that each of these arrangements may lead to the formation of chiral oxygenation products.     

TRPC1-mediated inhibition of 1-methyl-4-phenylpyridinium ion neurotoxicity in human SH-SY5Y neuroblastoma cells

Mammalian homologues of the Drosophila canonical transient receptor potential (TRP) proteins have been implicated to function as plasma membrane Ca(2+) channels. This study examined the role of TRPC1 in human neuroblastoma (SH-SY5Y) cells. SH-SY5Y cells treated with an exogenous neurotoxin, 1-methyl-4-phenylpyridinium ion (MPP(+)) significantly decreased TRPC1 protein levels. Confocal microscopy on SH-SY5Y cells treatment with MPP(+) showed decreased plasma membrane staining of TRPC1. Importantly, overexpression of TRPC1 reduced neurotoxicity induced by MPP(+). MPP(+)-induced alpha-synuclein expression was also suppressed by TRPC1 overexpression. Protection of SH-SY5Y cells against MPP(+) was significantly decreased upon the overexpression of antisense TRPC1 cDNA construct or the addition of a nonspecific transient receptor potential channel blocker lanthanum. Activation of TRPC1 by thapsigargin or carbachol decreased MPP(+) neurotoxicity, which was partially dependent on external Ca(2+). Staining of SH-SY5Y cells with an apoptotic marker (YO-PRO-1) showed that TRPC1 protects SH-SY5Y neuronal cells against apoptosis. Further, TRPC1 overexpression inhibited cytochrome c release and decreased Bax and Apaf-1 protein levels. Interpretation of the above data suggests that reduction in the cell surface expression of TRPC1 following MPP(+) treatment may be involved in dopaminergic neurodegeneration. Furthermore, TRPC1 may inhibit degenerative apoptotic signaling to provide neuroprotection against Parkinson's disease-inducing agents.     

Molecular epidemiology of ceftiofur-resistant Escherichia coli isolates from dairy calves

Healthy calves (n = 96, 1 to 9 weeks old) from a dairy herd in central Pennsylvania were examined each month over a five-month period for fecal shedding of ceftiofur-resistant gram-negative bacteria. Ceftiofur-resistant Escherichia coli isolates (n = 122) were characterized by antimicrobial resistance (disk diffusion and MIC), serotype, pulsed-field gel electrophoresis subtypes, beta-lactamase genes, and virulence genes. Antibiotic disk diffusion assays showed that the isolates were resistant to ampicillin (100%), ceftiofur (100%), chloramphenicol (94%), florfenicol (93%), gentamicin (89%), spectinomycin (72%), tetracycline (98%), ticarcillin (99%), and ticarcillin-clavulanic acid (99%). All isolates were multidrug resistant and displayed elevated MICs. The E. coli isolates belonged to 42 serotypes, of which O8:H25 was the predominant serotype (49.2%). Pulsed-field gel electrophoresis classified the E. coli isolates into 27 profiles. Cluster analysis showed that 77 isolates (63.1%) belonged to one unique group. The prevalence of pathogenic E. coli was low (8%). A total of 117 ceftiofur-resistant E. coli isolates (96%) possessed the bla(CMY2) gene. Based on phenotypic and genotypic characterization, the ceftiofur-resistant E. coli isolates belonged to 59 clonal types. There was no significant relationship between calf age and clonal type. The findings of this study revealed that healthy dairy calves were rapidly colonized by antibiotic-resistant strains of E. coli shortly after birth. The high prevalence of multidrug-resistant nonpathogenic E. coli in calves could be a significant source of resistance genes to other bacteria that share the same environment.     

Spatial event cluster detection using a compound Poisson distribution

Geographic disease surveillance methods identify regions that have higher disease rates than expected. These approaches are generally applied to incident or prevalent cases of disease. In some contexts, disease-related events rather than individuals are the appropriate units of analysis for geographic surveillance. We propose a compound Poisson approach that detects event clusters by testing individual areas that may be combined with their nearest neighbors. The method is applicable to situations where the population sizes are diverse and the population distribution by important strata may differ by area. For example, a geographical region might have sparse population in the northern areas, and other areas which are predominantly retirement communities. The approach requires a coarse geographical relationship and administrative data for the numbers of population, cases, and events in each area. Pediatric self-inflicted injuries requiring presentation to Alberta emergency departments provide an illustration.     

Refolding and purification of Zymomonas mobilis levansucrase produced as inclusion bodies in fed-batch culture of recombinant Escherichia coli

Zymomonas mobilis levansucrase was overproduced by the fed-batch culture of recombinant Escherichia coli harboring a novel expression system that is constitutively expressed by the promoter from the Rahnella aquatilis levansucrase gene. Most of the levansucrase was produced as inclusion bodies in the bacterial cytoplasm, accounting for approximately 20% of the total cellular protein. Refolding after complete denaturation by high concentrations of urea or guanidine hydrochloride was not successful, resulting in large amounts of insoluble aggregates. During the development of the refolding method, it was found that direct solubilization of the inclusion bodies with Triton X-100 reactivated the enzyme, with a considerable refolding efficiency. About 65% of inclusion body levansucrase was refolded into active levansucrase in the renaturation buffer containing 4% (v/v) Triton X-100. The in vitro refolded enzyme was purified to 95% purity by single-step DEAE-Sepharose ion exchange chromatography. Triton X-100 was removed by this ion exchange chromatography.     

Synthetic studies of cis-4-Amino-L-proline derivatives as novel lipid lowering agents

cis-4-Amino-L-proline derivatives 1 and 2 derived from quinazolinone and pyrazolo pyrimidone respectively were designed as novel lipid lowering agents. A preliminary in vivo screening indicates that 1b and 2a have moderate triglyceride lowering activity.     

The C(19) position of 25-hydroxyvitamin D(3) faces outward in the vitamin D sterol-binding pocket of vitamin D-binding protein

The radiolabeled affinity and photoaffinity analogues of 25-hydroxyvitamin D(3) (25-OH-D(3)) with probes at the C-19 position failed to specifically label the 25-OH-D(3)-binding pocket of vitamin D-binding protein (DBP). However, a hybrid analogue, with a bromoacetate affinity probe and a photoaffinity probe at C(3)-OH and C(19) positions, respectively, specifically labeled the ligand-binding pocket, suggesting that C(3)-OH points towards the 'inside' of the binding cavity while the C(19) position faces away from it.