CHO genome mining for synthetic promoter design

Synthetic promoters are an attractive alternative for use in mammalian hosts such as CHO cells as they can be designed de novo with user-defined functionalities. In this study, we describe and validate a method for bioprocess-directed design of synthetic promoters utilizing CHO genomic sequence information. We designed promoters with two objective features, (i) constitutive high-level recombinant gene transcription, and (ii) upregulated transcription under mild hypothermia or late-stage culture. CHO genes varying in transcriptional activity were selected based on a comparative analysis of RNA-Seq transcript levels in normal and biphasic cultures in combination with estimates of mRNA half-life from published genome scale datasets. Discrete transcription factor regulatory elements (TFREs) upstream of these genes were informatically identified and functionally screened in vitro to identify a subset of TFREs with the potential to support high activity recombinant gene transcription during biphasic cell culture processes. Two libraries of heterotypic synthetic promoters with varying TFRE combinations were then designed in silico that exhibited a maximal 2.5-fold increase in transcriptional strength over the CMV-IE promoter after transient transfection into host CHO-K1 cells. A subset of synthetic promoters was then used to create stable transfectant pools using CHO-K1 cells under glutamine synthetase selection. Whilst not achieving the maximal 2.5-fold increase in productivity over stable pools harboring the CMV promoter, all stably transfected cells utilizing synthetic promoters exhibited increased reporter production - up to 1.6-fold that of cells employing CMV, both in the presence or absence of intron A immediately downstream of the promoter. The increased productivity of stably transfected cells harboring synthetic promoters was maintained during fed-batch culture, with or without a transition to mild hypothermia at the onset of stationary phase. Our data exemplify that it is important to consider both host cell and intended bioprocess contexts as design criteria in the de novo construction of synthetic genetic parts for mammalian cell engineering

The effects of a 12-Month weight loss intervention on cognitive outcomes in adults with overweight and obesity

Obesity is associated with poorer executive functioning and reward sensitivity. Yet, we know very little about whether weight loss through diet and/or increased exercise engagement improves cognitive function. This study evaluated whether weight loss following a dietary and exercise intervention was associated with improved cognitive performance. We enrolled 125 middle-aged adults with overweight and obesity (98 female) into a 12-month behavioral weight loss intervention. Participants were assigned to one of three groups: energy-restricted diet alone, an energy-restricted diet plus 150 min of moderate intensity exercise per week or an energy restricted diet plus 250 min of exercise per week. All participants completed tests measuring executive functioning and/or reward sensitivity, including the Iowa Gambling Task (IGT). Following the intervention, weight significantly decreased in all groups. A MANCOVA controlling for age, sex and race revealed a significant multivariate effect of group on cognitive changes. Post-hoc ANCOVAs revealed a Group x Time interaction only on IGT reward sensitivity, such that the high exercise group improved their performance relative to the other two intervention groups. Post-hoc ANCOVAs also revealed a main effect of Time, independent of intervention group, on IGT net payoff score. Changes in weight were not associated with other changes in cognitive performance. Engaging in a high amount of exercise improved reward sensitivity above and beyond weight loss alone. This suggests that there is additional benefit to adding exercise into behavioral weight loss regimens on executive functioning, even without additional benefit to weight loss

Entangling power and operator entanglement in qudit systems

We establish the entangling power of a unitary operator on a general finite-dimensional bipartite quantum system with and without ancillas, and give relations between the entangling power based on the von Neumann entropy and the entangling power based on the linear entropy. Significantly, we demonstrate that the entangling power of a general controlled unitary operator acting on two equal-dimensional qudits is proportional to the corresponding operator entanglement if linear entropy is adopted as the quantity representing the degree of entanglement. We discuss the entangling power and operator entanglement of three representative quantum gates on qudits: the SUM, double SUM, and SWAP gates.Comment: 8 pages, 1 figure. Version 3: Figure was improved and the MS was a bit shortene

Deactivation study of the hydrodeoxygenation of p-methylguaiacol over silica supported rhodium and platinum catalysts

Hydrodeoxygenation of para-methylguaiacol using silica supported Rh or Pt catalysts was investigated using a fixed-bed reactor at 300 °C, under 4 barg hydrogen and a WHSV of 2.5 h−1. The activity, selectivity and deactivation of the catalysts were examined in relation to time on stream. Three catalysts were tested: 2.5% Rh/silica supplied by Johnson Matthey (JM), 2.5% Rh/silica and 1.55% Pt/silica both prepared in-house. The Rh/silica (JM) showed the best stability with steady-state reached after 6 h on stream and a constant activity over 3 days of reaction. In contrast the other two catalysts did not reach steady state within the timeframe of the tests, with continuous deactivation over the time on stream. Nevertheless higher coking was observed on the Rh/silica (JM) catalyst, while all three catalysts showed evidence of sintering. The Pt catalyst (A) showed higher selectivity for the production of 4-methylcatechol while the Rh catalysts were more selective toward the cresols. In all cases, complete hydrodeoxygenation of the methylguaiacol to methylcyclohexane was not observed

Fibulin-1 Is Increased in Asthma – A Novel Mediator of Airway Remodeling?

Background: The extracellular matrix is a dynamic and complex network of macromolecules responsible for maintaining and influencing cellular functions of the airway. The role of fibronectin, an extracellular matrix protein, is well documented in asthma. However, the expression and function of fibulin-1, a secreted glycoprotein which interacts with fibronectin, has not been reported. Fibulin-1 is widely expressed in basement membranes in many organs including the lung. There are four isoforms in humans (A-D) of which fibulin-1C and 1D predominate. The objective of this study was to study the expression of fibulin-1 in volunteers with and without asthma, and to examine its function in vitro. Methodology/Principal Findings: We used immunohistochemistry and dot-blots to examine fibulin-1 levels in bronchial biopsies, bronchoalveolar lavage fluid and serum. Real-time PCR for fibulin-1C and 1D, and ELISA and western blotting for fibulin-1 were used to study the levels in airway smooth muscle cells. The function of fibulin-1C was determined by assessing its role, using an antisense oligonucleotide, in cell proliferation, migration and wound healing. A murine model of airway hyperresponsiveness (AHR) was used to explore the biological significance of fibulin-1. Levels of fibulin-1 were significantly increased in the serum and bronchoalveolar lavage fluid of 21 asthmatics compared with 11 healthy volunteers. In addition fibulin-1 was increased in asthma derived airway smooth muscle cells and fibulin-1C contributed to the enhanced proliferation and wound repair in these cells. These features were reversed when fibulin-1C was suppressed using an antisense oligomer. In a mouse model of AHR, treatment with an AO inhibited the development of AHR to methacholine. Conclusions: Our data collectively suggest fibulin-1C may be worthy of further investigation as a target for airway remodeling in asthma

Cold Nuclear Matter Effects on Dijet Productions in Relativistic Heavy-ion Reactions at LHC

We investigate the cold nuclear matter(CNM) effects on dijet productions in high-energy nuclear collisions at LHC with the next-to-leading order perturbative QCD. The nuclear modifications for dijet angular distributions, dijet invariant mass spectra, dijet transverse momentum spectra and dijet momentum imbalance due to CNM effects are calculated by incorporating EPS, EKS, HKN and DS param-etrization sets of parton distributions in nucleus . It is found that dijet angular distributions and dijet momentum imbalance are insensitive to the initial-state CNM effects and thus provide optimal tools to study the final-state hot QGP effects such as jet quenching. On the other hand, the invariant mass spectra and the transverse momentum spectra of dijet are generally enhanced in a wide region of the invariant mass or transverse momentum due to CNM effects with a feature opposite to the expected suppression because of the final-state parton energy loss effect in the QGP. The difference of EPS, EKS, HKN and DS parametrization sets of nuclear parton distribution functions is appreciable for dijet invariant mass spectra and transverse momentum spectra at p+Pb collisions, and becomes more pronounced for those at Pb+Pb reactions.Comment: 10 pages, 11 figure

A genomic portrait of the emergence, evolution, and global spread of a methicillin-resistant staphylococcus aureus pandemic

The widespread use of antibiotics in association with high-density clinical care has driven the emergence of drug-resistant bacteria that are adapted to thrive in hospitalized patients. Of particular concern are globally disseminated methicillin-resistant Staphylococcus aureus (MRSA) clones that cause outbreaks and epidemics associated with health care. The most rapidly spreading and tenacious health-care-associated clone in Europe currently is EMRSA-15, which was first detected in the UK in the early 1990s and subsequently spread throughout Europe and beyond. Using phylogenomic methods to analyze the genome sequences for 193 S. aureus isolates, we were able to show that the current pandemic population of EMRSA-15 descends from a health-care-associated MRSA epidemic that spread throughout England in the 1980s, which had itself previously emerged from a primarily community-associated methicillin-sensitive population. The emergence of fluoroquinolone resistance in this EMRSA-15 subclone in the English Midlands during the mid-1980s appears to have played a key role in triggering pandemic spread, and occurred shortly after the first clinical trials of this drug. Genome-based coalescence analysis estimated that the population of this subclone over the last 20 yr has grown four times faster than its progenitor. Using comparative genomic analysis we identified the molecular genetic basis of 99.8% of the antimicrobial resistance phenotypes of the isolates, highlighting the potential of pathogen genome sequencing as a diagnostic tool. We document the genetic changes associated with adaptation to the hospital environment and with increasing drug resistance over time, and how MRSA evolution likely has been influenced by country-specific drug use regimens

Superconducting correlations in metallic nanoparticles: exact solution of the BCS model by the algebraic Bethe ansatz

Superconducting pairing of electrons in nanoscale metallic particles with discrete energy levels and a fixed number of electrons is described by the reduced BCS model Hamiltonian. We show that this model is integrable by the algebraic Bethe ansatz. The eigenstates, spectrum, conserved operators, integrals of motion, and norms of wave functions are obtained. Furthermore, the quantum inverse problem is solved, meaning that form factors and correlation functions can be explicitly evaluated. Closed form expressions are given for the form factors that describe superconducting pairing.Comment: revised version, 5 pages, revtex, no figure

Protein dynamics influence the enzymatic activity of phospholipase a/acyltransferases 3 and 4

Phospholipase A/acyltransferase 3 (PLAAT3) and PLAAT4 are enzymes involved in the synthesis of bioactive lipids. Despite sequential and structural similarities, the two enzymes differ in activity and specificity. The relation between the activity and dynamics of the N-terminal domains of PLAAT3 and PLAAT4 was studied. PLAAT3 has a much higher melting temperature and exhibits less nanosecond and millisecond dynamics in the active site, in particular in loop L2(B6), as shown by NMR spectroscopy and molecular dynamics calculations. Swapping the L2(B6) loops between the two PLAAT enzymes results in strongly increased phospholipase activity in PLAAT3 but no reduction in PLAAT4 activity, indicating that this loop contributes to the low activity of PLAAT3. The results show that, despite structural similarity, protein dynamics differ substantially between the PLAAT variants, which can help to explain the activity and specificity differences.Macromolecular BiochemistryMolecular Physiolog

Direct Measurements of the Branching Fractions for D0→K−e+νeD^0 \to K^-e^+\nu_e and D0→π−e+νeD^0 \to \pi^-e^+\nu_e and Determinations of the Form Factors f+K(0)f_{+}^{K}(0) and f+π(0)f^{\pi}_{+}(0)

The absolute branching fractions for the decays $D^0 \to K^-e ^+\nu_e$ and $D^0 \to \pi^-e^+\nu_e$ are determined using $7584\pm 198 \pm 341$ singly tagged $\bar D^0$ sample from the data collected around 3.773 GeV with the BES-II detector at the BEPC. In the system recoiling against the singly tagged $\bar D^0$ meson, $104.0\pm 10.9$ events for $D^0 \to K^-e ^+\nu_e$ and $9.0 \pm 3.6$ events for $D^0 \to \pi^-e^+\nu_e$ decays are observed. Those yield the absolute branching fractions to be $BF(D^0 \to K^-e^+\nu_e)=(3.82 \pm 0.40\pm 0.27)$ and $BF(D^0 \to \pi^-e^+\nu_e)=(0.33 \pm 0.13\pm 0.03)$. The vector form factors are determined to be $|f^K_+(0)| = 0.78 \pm 0.04 \pm 0.03$ and $|f^{\pi}_+(0)| = 0.73 \pm 0.14 \pm 0.06$. The ratio of the two form factors is measured to be $|f^{\pi}_+(0)/f^K_+(0)|= 0.93 \pm 0.19 \pm 0.07$.Comment: 6 pages, 5 figure