THE DNA POLYMERASE GENE OF A BROWN ALGAL VIRUS: STRUCTURE AND PHYLOGENY

We have reported the complete sequence of the DNA polymerase gene from the virus that infected a filamentous brown alga, Feldmannia sp. (FsV). The DNA polymerase gene from FsV encoded 986 amino acids and contained all the conserved motifs of 3'-5' exonuclease domains and catalytic domains found in B-family (α-like) DNA polymerases. The codons for the FsV DNA polymerase appeared to have some bias toward guanine/cytosine (G/C) in the third position. The phylogenetic analysis of the FsV DNA polymerase gene and other viral DNA polymerase genes indicated that FsV belongs to a family of algal viruses recently defined as Phycodnaviridae.     

Gadolinium prevents high airway pressure-induced permeability increases in isolated rat lungs

To determine theinitial signaling event in the vascular permeability increase afterhigh airway pressure injury, we compared groups of lungs ventilated atdifferent peak inflation pressures (PIPs) with (gadolinium group) andwithout (control group) infusion of 20 µM gadolinium chloride, aninhibitor of endothelial stretch-activated cationchannels. Microvascular permeability was assessed by using the capillary filtration coefficient(K_fc), ameasure of capillary hydraulic conductivity.K_fc was measuredafter ventilation for 30-min periods with 7, 20, and 30 cmH₂O PIP with 3 cmH₂O positive end-expiratorypressure and with 35 cmH₂O PIPwith 8 cmH₂O positive end-expiratory pressure. In control lungs,K_fc increasedsignificantly to 1.8 and 3.7 times baseline after 30 and 35 cmH₂O PIP, respectively. In thegadolinium group,K_fc was unchangedfrom baseline (0.060 ± 0.010 ml · min¹ · cmH₂O¹ · 100 g¹) after any PIPventilation period. Pulmonary vascular resistance increasedsignificantly from baseline in both groups before the lastK_fc measurementbut was not different between groups. These results suggest thatmicrovascular permeability is actively modulated by a cellular responseto mechanical injury and that stretch-activated cation channels mayinitiate this response through increases in intracellular calciumconcentration.

     

miR-126 regulates angiogenic signaling and vascular integrity

Precise regulation of the formation, maintenance, and remodeling of the vasculature is required for normal development, tissue response to injury, and tumor progression. How specific microRNAs intersect with and modulate angiogenic signaling cascades is unknown. Here, we identified microRNAs that were enriched in endothelial cells derived from mouse embryonic stem (ES) cells and in developing mouse embryos. We found that miR-126 regulated the response of endothelial cells to VEGF. Additionally, knockdown of miR-126 in zebrafish resulted in loss of vascular integrity and hemorrhage during embryonic development. miR-126 functioned in part by directly repressing negative regulators of the VEGF pathway, including the Sprouty-related protein SPRED1 and phosphoinositol-3 kinase regulatory subunit 2 (PIK3R2/p85-beta). Increased expression of Spred1 or inhibition of VEGF signaling in zebrafish resulted in defects similar to miR-126 knockdown. These findings illustrate that a single miRNA can regulate vascular integrity and angiogenesis, providing a new target for modulating vascular formation and function.     

Carbon monoxide-driven electron transport in Clostridium thermoautotrophicum membranes.

Membrane vesicles of Clostridium thermoautotrophicum prepared by osmotic lysis after lysozyme treatment contained carbon monoxide dehydrogenase and methylenetetrahydrofolate dehydrogenase with specific activities three- to fourfold higher than the specific activity of the cytoplasm. The membrane-associated carbon monoxide dehydrogenase mediated the reduction with CO or the oxidation with CO2 of b-type cytochromes and other electron carriers in the membrane.     

Mobilization Assessment and Possibility of Increased Availability of PAHs in Contaminated Soil Using Column Tests

Surfactants are well known to increase solubility/mobility of hydrocarbons and can be used to remediate contaminated water and soil. We wanted to explore if Ivey sol® 106 used at less than the critical micelle concentration (CMC) could effectively mobilize PAH (polycyclic aromatic hydrocarbons) from contaminated soil. The first step was to establish a measurement technique. Hence, a column leaching method was undertaken to investigate mobility of PAH-contaminated soil from a former gaswork facility. The methodology was based on a recycled flow of aqueous solution containing CaCl₂ 0.01M through two different soil columns. In the first column test, the free desorption of hydrocarbons was studied by recycling the solution through the soil column with a peristaltic pump and with a liquid/solid ratio of 2, based on ISO/DIS 18772. The solution was replaced with new solution every three days to aid desorption.

In the second column test, the set-up was similar with the exception of the aforementioned recycling solution. In this case, a second column was filled with a resin, Amberlite XAD-2, which captures PAHs entering the solution through the soil column, cleaning it of hydrocarbons (induced desorption). The results obtained for induced desorption and free desorption with reposition showed that liberation of PAHs in the presence of resins was higher (7%) as opposed to free desorption (below 0.3%). These two experiments demonstrated low mobilization of PAHs.

A third column test was performed using a non ionic surfactant, Ivey-sol® 106, 100 μg g^?1 of soil below the CMC in the recycling solution. The introduction of Ivey-sol® 106 at 0.005%w/v increased desorption of PAHs to 32%, thus demonstrating the potential for increased bioavailability of the PAHs for bioremediation of the soil.     

Detection and identification of microorganisms in wine: a review of molecular techniques

The microbial ecology of wine is complex. Microbes can play both positive and negative roles in the quality of the final product. Due to this impact, the microbial ecology of wine has been well studied. Traditional indirect methods, such as plating, have largely been replaced by a number of molecular methods. These methods are typically either indirect methods used for identification of cultured organisms, or direct methods used to profile whole populations or identify specific microbes in a mixed population. These molecular methods offer a number of advantages over traditional methods including speed and precision. This review will examine both direct and indirect molecular methods, provide examples of their impact on the study of the microbial ecology of wine, and also discuss their strengths and limitations.     

Multiplexed quantification of estrogen receptor and HER2/Neu in tissue and cell lysates by peptide immunoaffinity enrichment mass spectrometry

Access to a wider range of quantitative protein assays would significantly impact the number and use of tissue markers in guiding disease treatment. Quantitative mass spectrometry-based peptide and protein assays, such as immuno-SRM assays, have seen tremendous growth in recent years in application to protein quantification in biological fluids such as plasma or urine. Here, we extend the capability of the technique by demonstrating the application of a multiplexed immuno-SRM assay for quantification of estrogen receptor (ER) and human epidermal growth factor receptor 2 (HER2) levels in cell line lysates and human surgical specimens. The performance of the assay was characterized using peptide response curves, with linear ranges covering approximately four orders of magnitude and limits of detection in the low fmol/mg lysate range. Reproducibility was acceptable with median coefficients of variation of approximately 10%. We applied the assay to measurements of ER and HER2 in well-characterized cell line lysates with good discernment based on ER/HER2 status. Finally, the proteins were measured in surgically resected breast cancers, and the results showed good correlation with ER/HER2 status determined by clinical assays. This is the first implementation of the peptide-based immuno-SRM assay technology in cell lysates and human surgical specimens.