Functional interplay between NTP leaving group and base pair recognition during RNA polymerase II nucleotide incorporation revealed by methylene substitution.

RNA polymerase II (pol II) utilizes a complex interaction network to select and incorporate correct nucleoside triphosphate (NTP) substrates with high efficiency and fidelity. Our previous 'synthetic nucleic acid substitution' strategy has been successfully applied in dissecting the function of nucleic acid moieties in pol II transcription. However, how the triphosphate moiety of substrate influences the rate of P-O bond cleavage and formation during nucleotide incorporation is still unclear. Here, by employing β,γ-bridging atom-'substituted' NTPs, we elucidate how the methylene substitution in the pyrophosphate leaving group affects cognate and non-cognate nucleotide incorporation. Intriguingly, the effect of the β,γ-methylene substitution on the non-cognate UTP/dT scaffold (∼3-fold decrease in kpol) is significantly different from that of the cognate ATP/dT scaffold (∼130-fold decrease in kpol). Removal of the wobble hydrogen bonds in U:dT recovers a strong response to methylene substitution of UTP. Our kinetic and modeling studies are consistent with a unique altered transition state for bond formation and cleavage for UTP/dT incorporation compared with ATP/dT incorporation. Collectively, our data reveals the functional interplay between NTP triphosphate moiety and base pair hydrogen bonding recognition during nucleotide incorporation

The Pothole Hydrology-Linked Systems Simulator (PHyLiSS)—Development and Application of a Systems Model for Prairie-Pothole Wetlands

The North American Prairie Pothole Region covers about 770,000 square kilometers of the United States and Canada (including parts of 5 States and 3 provinces: North Dakota, South Dakota, Montana, Minnesota, Iowa, Saskatchewan, Manitoba, and Alberta). The Laurentide Ice Sheet shaped the landscape of the region about 12,000 to 14,000 years ago. The retreat of the ice sheet left behind low-permeability glacial till and a landscape dotted with millions of depressions known today as prairie potholes. The wetlands that subsequently formed in these depressions, prairie-pothole wetlands, provide critical migratory-bird habitat and support dynamic aquatic communities. Extensive grasslands and productive agricultural systems surround these wetland ecosystems. In prairie-pothole wetlands, the compositions of plant, invertebrate, and vertebrate communities are highly dependent on hydrogeochemical conditions. Regional climate shifts between wet and dry periods affect the length of time that wetlands contain ponded surface water and the chemistry of that ponded water. Land-use change can exacerbate or reduce the effects of climate on wetland hydrology and water chemistry. A mechanistic understanding of the relation among climate, land use, hydrology, chemistry, and biota in prairie-pothole wetlands is needed to better understand the complex, and often interacting, effects of climate and land use on prairie-pothole wetland systems and to facilitate climate and land-use change adaptation efforts. The Pothole Hydrology-Linked Systems Simulator (PHyLiSS) model was developed to address this need. The model simulates water-surface elevation dynamics in prairie-pothole wetlands and quantifies changes in salinity. The PHyLiSS model is unique among other wetland models because it accommodates differing sizes and morphometries of wetland basins, is not dependent on a priori designations of wetland class, and allows for functional changes associated with dynamic shifts in ecohydrological states. The PHyLiSS model also has the capability to simulate wetland salinity, and potential future iterations will also simulate the effects of changing hydrology and geochemical conditions on biota. This report documents the development of the hydrological and geochemical components of the PHyLiSS model and provides example applications

Markov models and the ensemble Kalman filter for estimation of sorption rates.

Non-equilibrium sorption of contaminants in ground water systems is examined from the perspective of sorption rate estimation. A previously developed Markov transition probability model for solute transport is used in conjunction with a new conditional probability-based model of the sorption and desorption rates based on breakthrough curve data. Two models for prediction of spatially varying sorption and desorption rates along a one-dimensional streamline are developed. These models are a Markov model that utilizes conditional probabilities to determine the rates and an ensemble Kalman filter (EnKF) applied to the conditional probability method. Both approaches rely on a previously developed Markov-model of mass transfer, and both models assimilate the observed concentration data into the rate estimation at each observation time. Initial values of the rates are perturbed from the true values to form ensembles of rates and the ability of both estimation approaches to recover the true rates is examined over three different sets of perturbations. The models accurately estimate the rates when the mean of the perturbations are zero, the unbiased case. For the cases containing some bias, addition of the ensemble Kalman filter is shown to improve accuracy of the rate estimation by as much as an order of magnitude

NVP-BEZ235 and NVP-BGT226, dual phosphatidylinositol 3-kinase/mammalian target of rapamycin inhibitors, enhance tumor and endothelial cell radiosensitivity

<p>Abstract</p> <p>Background</p> <p>The phosphatidylinositol 3-kinase (PI3K)/Akt pathway is activated in tumor cells and promotes tumor cell survival after radiation-induced DNA damage. Because the pathway may not be completely inhibited after blockade of PI3K itself, due to feedback through mammalian target of rapamycin (mTOR), more effective inhibition might be expected by targeting both PI3K and mTOR inhibition.</p> <p>Materials and methods</p> <p>We investigated the effect of two dual PI3K/mTOR (both mTORC1 and mTORC2) inhibitors, NVP-BEZ235 and NVP-BGT226, on SQ20B laryngeal and FaDu hypopharyngeal cancer cells characterised by EGFR overexpression, on T24 bladder tumor cell lines with H-Ras mutation and on endothelial cells. Analysis of target protein phosphorylation, clonogenic survival, number of residual γH2AX foci, cell cycle and apoptosis after radiation was performed in both tumor and endothelial cells. In vitro angiogenesis assays were conducted as well.</p> <p>Results</p> <p>Both compounds effectively inhibited phosphorylation of Akt, mTOR and S6 target proteins and reduced clonogenic survival in irradiated tumor cells. Persistence of DNA damage, as evidenced by increased number of γH2AX foci, was detected after irradiation in the presence of PI3K/mTOR inhibition, together with enhanced G2 cell cycle delay. Treatment with one of the inhibitors, NVP-BEZ235, also resulted in decreased clonogenicity after irradiation of tumor cells under hypoxic conditions. In addition, NVP-BEZ235 blocked VEGF- and IR-induced Akt phosphorylation and increased radiation killing in human umbilical venous endothelial cells (HUVEC) and human dermal microvascular dermal cells (HDMVC). NVP-BEZ235 inhibited VEGF-induced cell migration and capillary tube formation in vitro and enhanced the antivascular effect of irradiation. Treatment with NVP-BEZ235 moderately increased apoptosis in SQ20B and HUVEC cells but not in FaDu cells, and increased necrosis in both tumor and endothelial all cells tumor.</p> <p>Conclusions</p> <p>The results of this study demonstrate that PI3K/mTOR inhibitors can enhance radiation-induced killing in tumor and endothelial cells and may be of benefit when combined with radiotherapy.</p

Effect of Maternal Exercise on Adverse Cardio-Pulmonary Responses in Offspring Exposed to Post-Natal Chronic Hypoxia

Emerging evidence shows exercise by women throughout pregnancy (i.e., maternal exercise) endows the offspring with positive health benefits and resistance to many diseases later in life. PURPOSE: To determine if maternal exercise reduces unfavorable cardiopulmonary development and improves exercise capacity in offspring after chronic hypoxic exposure throughout early life. METHODS: Female adult C57BL/6J mice were randomly assigned to one of the following groups: high altitude, sedentary (HS), high altitude, active (HE), or the respective, low-altitude conditions (LS or LE). Pregnant females in the exercise groups performed voluntary wheel running during mating and the perinatal period (about 4 weeks). Three days after birth, the pups in the high-altitude groups were exposed to chronic hypobaric hypoxia (450mmHg). When pups were eight weeks old, body size (weight and tail length) was measured and exercise capacity was assessed via a Rotarod test. Terminal procedures were performed to measure right ventricular systolic pressure (RVSP), hematocrit, and heart remodeling (Fulton’s Index: right ventricular weight ratio to bodyweight (RV/BW) and to the left ventricle plus septum (RV/LV+S)). RESULTS: All variables measured had a main effect for altitude (pCONCLUSION: Chronic hypoxia decreases exercise capacity and attenuates growth. This stress also induces potentially harmful cardiovascular changes, such as high blood viscosity, RVSP, and right ventricular hypertrophy. Our results indicate that maternal exercise may attenuate some of the negative effects of chronic hypoxia in the offspring, but does not prevent significant development impairments

Linear stability analysis of transverse dunes

Sand-moving winds blowing from a constant direction in an area of high sand availability form transverse dunes, which have a fixed profile in the direction orthogonal to the wind. Here we show, by means of a linear stability analysis, that transverse dunes are intrinsically unstable. Any along-axis perturbation on a transverse dune amplify in the course of dune migration due to the combined effect of two main factors, namely: the lateral transport through avalanches along the dune's slip-face, and the scaling of dune migration velocity with the inverse of the dune height. Our calculations provide a quantitative explanation for recent observations from experiments and numerical simulations, which showed that transverse dunes moving on the bedrock cannot exist in a stable form and decay into a chain of crescent-shaped barchans.Comment: 8 pages, 4 figure

Randomized trial of l-serine in patients with hereditary sensory and autonomic neuropathy type 1

OBJECTIVE: To evaluate the safety and efficacy of l-serine in humans with hereditary sensory autonomic neuropathy type I (HSAN1). METHODS: In this randomized, placebo-controlled, parallel-group trial with open-label extension, patients aged 18-70 years with symptomatic HSAN1 were randomized to l-serine (400 mg/kg/day) or placebo for 1 year. All participants received l-serine during the second year. The primary outcome measure was the Charcot-Marie-Tooth Neuropathy Score version 2 (CMTNS). Secondary outcomes included plasma sphingolipid levels, epidermal nerve fiber density, electrophysiologic measurements, patient-reported measures, and adverse events. RESULTS: Between August 2013 and April 2014, we enrolled and randomized 18 participants, 16 of whom completed the study. After 1 year, the l-serine group experienced improvement in CMTNS relative to the placebo group (-1.5 units, 95% CI -2.8 to -0.1, p = 0.03), with evidence of continued improvement in the second year of treatment (-0.77, 95% CI -1.67 to 0.13, p = 0.09). Concomitantly, deoxysphinganine levels dropped in l-serine-treated but not placebo-treated participants (59% decrease vs 11% increase; p \u3c 0.001). There were no serious adverse effects related to l-serine. CONCLUSION: High-dose oral l-serine supplementation appears safe in patients with HSAN1 and is potentially effective at slowing disease progression. CLINICALTRIALSGOV IDENTIFIER: NCT01733407. CLASSIFICATION OF EVIDENCE: This study provides Class I evidence that high-dose oral l-serine supplementation significantly slows disease progression in patients with HSAN1

Emergency department and hospital crowding: causes, consequences, and cures

Overcrowding with associated delays in patient care is a problem faced by emergency departments (EDs) worldwide. ED overcrowding can be the result of poor ED department design and prolonged throughput due to staffing, ancillary service performance, and flow processes. As such, the problem may be addressed by process improvements within the ED. A broad body of literature demonstrates that ED overcrowding can be a function of hospital capacity rather than an ED specific issue. Lack of institutional capacity leads to boarding in the ED with resultant ED crowding. This is a problem not solvable by the ED and must be addressed as an institution-wide problem. This paper discusses the causes of ED overcrowding, provides a brief overview of the drastic consequences, and discusses possible cures that have been successfully implemented

Rapidly Decaying Supernova 2010X: A Candidate ".Ia" Explosion

We present the discovery, photometric and spectroscopic follow-up observations of SN 2010X (PTF 10bhp). This supernova decays exponentially with tau_d=5 days, and rivals the current recordholder in speed, SN 2002bj. SN 2010X peaks at M_r=-17mag and has mean velocities of 10,000 km/s. Our light curve modeling suggests a radioactivity powered event and an ejecta mass of 0.16 Msun. If powered by Nickel, we show that the Nickel mass must be very small (0.02 Msun) and that the supernova quickly becomes optically thin to gamma-rays. Our spectral modeling suggests that SN 2010X and SN 2002bj have similar chemical compositions and that one of Aluminum or Helium is present. If Aluminum is present, we speculate that this may be an accretion induced collapse of an O-Ne-Mg white dwarf. If Helium is present, all observables of SN 2010X are consistent with being a thermonuclear Helium shell detonation on a white dwarf, a ".Ia" explosion. With the 1-day dynamic-cadence experiment on the Palomar Transient Factory, we expect to annually discover a few such events.Comment: 6 pages, 5 figures; Minor Changes; Note correction in Fig 4 caption; published by ApJ