Extragenic suppressor mutations in ΔripA disrupt stability and function of LpxA

Background: Francisella tularensis is a Gram-negative bacterium that infects hundreds of species including humans, and has evolved to grow efficiently within a plethora of cell types. RipA is a conserved membrane protein of F. tularensis, which is required for growth inside host cells. As a means to determine RipA function we isolated and mapped independent extragenic suppressor mutants in ΔripA that restored growth in host cells. Each suppressor mutation mapped to one of two essential genes, lpxA or glmU, which are involved in lipid A synthesis. We repaired the suppressor mutation in lpxA (S102, LpxA T36N) and the mutation in glmU (S103, GlmU E57D), and demonstrated that each mutation was responsible for the suppressor phenotype in their respective strains. We hypothesize that the mutation in S102 altered the stability of LpxA, which can provide a clue to RipA function. LpxA is an UDP-N-acetylglucosamine acyltransferase that catalyzes the transfer of an acyl chain from acyl carrier protein (ACP) to UDP-N-acetylglucosamine (UDP-GlcNAc) to begin lipid A synthesis. Results: LpxA was more abundant in the presence of RipA. Induced expression of lpxA in the ΔripA strain stopped bacterial division. The LpxA T36N S102 protein was less stable and therefore less abundant than wild type LpxA protein. Conclusion: These data suggest RipA functions to modulate lipid A synthesis in F. tularensis as a way to adapt to the host cell environment by interacting with LpxA

Spatiotemporal Stochastic Resonance in Fully Frustrated Josephson Ladders

We consider a Josephson-junction ladder in an external magnetic field with half flux quantum per plaquette. When driven by external currents, periodic in time and staggered in space, such a fully frustrated system is found to display spatiotemporal stochastic resonance under the influence of thermal noise. Such resonance behavior is investigated both numerically and analytically, which reveals significant effects of anisotropy and yields rich physics.Comment: 8 pages in two columns, 8 figures, to appear in Phys. Rev.

Potential of Notochordal Cells within Injectable Biomaterials to Promote Intervertebral Disc Regeneration

Low back pain is the leading cause of disability worldwide and is strongly associated with degeneration of the intervertebral disc (IVD).During degeneration the nucleus pulposus (NP) in the core of the IVD, is affected by altered matrix synthesis, increased degradation, andcell loss. Strategies combining regenerative cell sources with injectable biomaterials could provide a therapeutic approach to treatingIVD-degeneration related back pain. The juvenile cells of the NP, known as notochordal cells (NC), could provide both anabolic andanti-catabolic responses for disc regeneration. However, their behaviour within biomaterial delivery systems has not been investigated.Here, porcine NCs were incorporated into three injectable hydrogels: Albugel (an albumin/hyaluronan hydrogel), NPgel (a L-pNIPAMco-DMAc hydrogel) and NPgel with decellularized NC-matrix powder (dNCM). The NCs and biomaterial constructs were cultured for upto 4 weeks under 5% oxygen (n = 3 biological repeats). The ability of biomaterials to maintain NC viability, phenotype and extracellularmatrix synthesis and deposition was investigated through histological, immunohisto chemical and glycosaminogly cans analysis. NCs survived in all three biomaterials after 4 weeks, whilst phenotype and cell clustering were maintained to a greater extent in NPgel and Albugel. Thus, these biomaterials could facilitate maintenance of the NC phenotype, support matrix deposition and be a basis for future IVD regeneration strategies

The Structure and Dynamics of the Upper Chromosphere and Lower Transition Region as Revealed by the Subarcsecond VAULT Observations

The Very high Angular resolution ULtraviolet Telescope (VAULT) is a sounding rocket payload built to study the crucial interface between the solar chromosphere and the corona by observing the strongest line in the solar spectrum, the Ly-a line at 1216 {\AA}. In two flights, VAULT succeeded in obtaining the first ever sub-arcsecond (0.5") images of this region with high sensitivity and cadence. Detailed analyses of those observations have contributed significantly to new ideas about the nature of the transition region. Here, we present a broad overview of the Ly-a atmosphere as revealed by the VAULT observations, and bring together past results and new analyses from the second VAULT flight to create a synthesis of our current knowledge of the high-resolution Ly-a Sun. We hope that this work will serve as a good reference for the design of upcoming Ly-a telescopes and observing plans.Comment: 28 pages, 11 figure

Recent Advances in Understanding Particle Acceleration Processes in Solar Flares

We review basic theoretical concepts in particle acceleration, with particular emphasis on processes likely to occur in regions of magnetic reconnection. Several new developments are discussed, including detailed studies of reconnection in three-dimensional magnetic field configurations (e.g., current sheets, collapsing traps, separatrix regions) and stochastic acceleration in a turbulent environment. Fluid, test-particle, and particle-in-cell approaches are used and results compared. While these studies show considerable promise in accounting for the various observational manifestations of solar flares, they are limited by a number of factors, mostly relating to available computational power. Not the least of these issues is the need to explicitly incorporate the electrodynamic feedback of the accelerated particles themselves on the environment in which they are accelerated. A brief prognosis for future advancement is offered.Comment: This is a chapter in a monograph on the physics of solar flares, inspired by RHESSI observations. The individual articles are to appear in Space Science Reviews (2011

Strategic Fluorination to Achieve a Potent, Selective, Metabolically Stable, and Orally Bioavailable Inhibitor of CSNK2

The host kinase casein kinase 2 (CSNK2) has been proposed to be an antiviral target against β-coronaviral infection. To pharmacologically validate CSNK2 as a drug target in vivo, potent and selective CSNK2 inhibitors with good pharmacokinetic properties are required. Inhibitors based on the pyrazolo[1,5-a]pyrimidine scaffold possess outstanding potency and selectivity for CSNK2, but bioavailability and metabolic stability are often challenging. By strategically installing a fluorine atom on an electron-rich phenyl ring of a previously characterized inhibitor 1, we discovered compound 2 as a promising lead compound with improved in vivo metabolic stability. Compound 2 maintained excellent cellular potency against CSNK2, submicromolar antiviral potency, and favorable solubility, and was remarkably selective for CSNK2 when screened against 192 kinases across the human kinome. We additionally present a co-crystal structure to support its on-target binding mode. In vivo, compound 2 was orally bioavailable, and demonstrated modest and transient inhibition of CSNK2, although antiviral activity was not observed, possibly attributed to its lack of prolonged CSNK2 inhibition

More than an Amide Bioisostere: Discovery of 1,2,4-Triazole-containing Pyrazolo[1,5-a]pyrimidine Host CSNK2 Inhibitors for Combatting β-Coronavirus Replication

The pyrazolo[1,5-a]pyrimidine scaffold is a promising scaffold to develop potent and selective CSNK2 inhibitors with antiviral activity against β-coronaviruses. Herein, we describe the discovery of a 1,2,4-triazole group to substitute a key amide group for CSNK2 binding present in many potent pyrazolo[1,5-a]pyrimidine inhibitors. Crystallographic evidence demonstrates that the 1,2,4-triazole replaces the amide in forming key hydrogen bonds with Lys68 and a water molecule buried in the ATP-binding pocket. This isosteric replacement improves potency and metabolic stability at a cost of solubility. Optimization for potency, solubility, and metabolic stability led to the discovery of the potent and selective CSNK2 inhibitor 53. Despite excellent in vitro metabolic stability, rapid decline in plasma concentration of 53 in vivo was observed and may be attributed to lung accumulation, although in vivo pharmacological effect was not observed. Further optimization of this novel chemotype may validate CSNK2 as an antiviral target in vivo

An Integrated TCGA Pan-Cancer Clinical Data Resource to Drive High-Quality Survival Outcome Analytics

For a decade, The Cancer Genome Atlas (TCGA) program collected clinicopathologic annotation data along with multi-platform molecular profiles of more than 11,000 human tumors across 33 different cancer types. TCGA clinical data contain key features representing the democratized nature of the data collection process. To ensure proper use of this large clinical dataset associated with genomic features, we developed a standardized dataset named the TCGA Pan-Cancer Clinical Data Resource (TCGA-CDR), which includes four major clinical outcome endpoints. In addition to detailing major challenges and statistical limitations encountered during the effort of integrating the acquired clinical data, we present a summary that includes endpoint usage recommendations for each cancer type. These TCGA-CDR findings appear to be consistent with cancer genomics studies independent of the TCGA effort and provide opportunities for investigating cancer biology using clinical correlates at an unprecedented scale. Analysis of clinicopathologic annotations for over 11,000 cancer patients in the TCGA program leads to the generation of TCGA Clinical Data Resource, which provides recommendations of clinical outcome endpoint usage for 33 cancer types