N-Methylmesoporphyrin IX Fluorescence As A Reporter Of Strand Orientation In Guanine Quadruplexes

Guanine quadruplexes (GQ) are four-stranded DNA structures formed by guanine-rich DNA sequences. The formation of GQs inhibits cancer cell growth, although the detection of GQs invivo has proven difficult, in part because of their structural diversity. The development of GQ-selective fluorescent reporters would enhance our ability to quantify the number and location of GQs, ultimately advancing biological studies of quadruplex relevance and function. N-methylmesoporphyrin IX (NMM) interacts selectively with parallel-stranded GQs; in addition, its fluorescence is sensitive to the presence of DNA, making this ligand a possible candidate for a quadruplex probe. In the present study, we investigated the effect of DNA secondary structure on NMM fluorescence. We found that NMM fluorescence increases by about 60-fold in the presence of parallel-stranded GQs and by about 40-fold in the presence of hybrid GQs. Antiparallel GQs lead to lower than 10-fold increases in NMM fluorescence. Single-stranded DNA, duplex, or i-motif, induce no change in NMM fluorescence. We conclude that NMM shows promise as a turn-on\u27 fluorescent probe for detecting quadruplex structures, as well as for differentiating them on the basis of strand orientation

Cas9 interrogates DNA in discrete steps modulated by mismatches and supercoiling.

The CRISPR-Cas9 nuclease has been widely repurposed as a molecular and cell biology tool for its ability to programmably target and cleave DNA. Cas9 recognizes its target site by unwinding the DNA double helix and hybridizing a 20-nucleotide section of its associated guide RNA to one DNA strand, forming an R-loop structure. A dynamic and mechanical description of R-loop formation is needed to understand the biophysics of target searching and develop rational approaches for mitigating off-target activity while accounting for the influence of torsional strain in the genome. Here we investigate the dynamics of Cas9 R-loop formation and collapse using rotor bead tracking (RBT), a single-molecule technique that can simultaneously monitor DNA unwinding with base-pair resolution and binding of fluorescently labeled macromolecules in real time. By measuring changes in torque upon unwinding of the double helix, we find that R-loop formation and collapse proceed via a transient discrete intermediate, consistent with DNA:RNA hybridization within an initial seed region. Using systematic measurements of target and off-target sequences under controlled mechanical perturbations, we characterize position-dependent effects of sequence mismatches and show how DNA supercoiling modulates the energy landscape of R-loop formation and dictates access to states competent for stable binding and cleavage. Consistent with this energy landscape model, in bulk experiments we observe promiscuous cleavage under physiological negative supercoiling. The detailed description of DNA interrogation presented here suggests strategies for improving the specificity and kinetics of Cas9 as a genome engineering tool and may inspire expanded applications that exploit sensitivity to DNA supercoiling

Observations of Peak Strength Behavior in CSA Cement Mortars

The purpose of this study was to assess the mechanical property performance behavior of calcium sulfoaluminate (CSA) cement mortar when cured at ambient laboratory temperature of ~23°C (73°F) and constant 50% relative humidity for an extended period of time. Four CSA cement mortars were tested. Three CSA cement mortars contained equivalent mass amounts of calcium sulfate; whereas, the fourth mortar contained double the amount of calcium sulfate. The three CSA cement mortars containing constant mass amounts of calcium sulfate differed as the specific type of calcium sulfate varied across the three formulations—one mortar containing solely anhydrite, one mortar contained half anhydrite and half gypsum while the other mortar solely contained gypsum. The fourth mortar contained double the amount of calcium sulfate when compared with the others while having a 1/1 blend of anhydrite and gypsum. Specific mortars were either tested for direct tensile strength according to ASTM C307 or for compressive strength according to ASTM C109. All tested mortars displayed statistically significant strength loss trends versus time when cured at constant 50% relative humidity. Cement paste samples were analyzed with TGA/SDT and XRD in an effort to identify microstructure changes corresponding to observed strength loss. Cement paste analysis suggests strength loss within the tested CSA cement mortars occurred as a result of ettringite decomposition

The Cerebellar Fastigial Nucleus Contributes to CO\u3csub\u3e2\u3c/sub\u3e-H\u3csup\u3e+\u3c/sup\u3e Ventilatory Sensitivity in Awake Goats

The purpose of this study was to test the hypothesis that an intact cerebellar fastigial nucleus (CFN) is an important determinant of CO2-H+ sensitivity during wakefulness. Bilateral, stainless steel microtubules were implanted into the CFN (N = 9) for injection (0.5–10 μl) of the neurotoxin ibotenic acid. Two or more weeks after implantation of the microtubules, eupneic breathing and CO2-H+ sensitivity did not differ significantly (P \u3e 0.10) from pre-implantation conditions. Injection of ibotenic acid (50 mM) did not significantly alter eupneic PaCO2 (P \u3e 0.10). The coefficient of variation of eupneic PaCO2 was 4.0 ± 0.6 and 3.7 ± 0.4% over the 2 weeks before and after the lesion, respectively. CO2-H+ sensitivity expressed as inspired ventilation/PaCO2 decreased from 2.15 ± 0.17 pre-lesion to 1.58 ± 0.26 l/(min mmHg) 3–6 days post-lesion (P \u3c 0.02, −27%). There was no significant (P \u3e 0.10) recovery of sensitivity between 7 and 10 days post-lesion. The lesion also increased (P \u3c 0.05) the day-to-day variability of this index by nearly 100%. When CO2 sensitivity was expressed as elevated inspired CO2/room air VI, values at 7%, but not 3 and 5% inspired CO2, were reduced and more variable (P \u3c 0.05) after the ibotenic acid injections. We conclude that during wakefulness, the CFN contributes relatively more to overall ventilatory drive at high relative to low levels of hypercapnia

Networks and the epidemiology of infectious disease

The science of networks has revolutionised research into the dynamics of interacting elements. It could be argued that epidemiology in particular has embraced the potential of network theory more than any other discipline. Here we review the growing body of research concerning the spread of infectious diseases on networks, focusing on the interplay between network theory and epidemiology. The review is split into four main sections, which examine: the types of network relevant to epidemiology; the multitude of ways these networks can be characterised; the statistical methods that can be applied to infer the epidemiological parameters on a realised network; and finally simulation and analytical methods to determine epidemic dynamics on a given network. Given the breadth of areas covered and the ever-expanding number of publications, a comprehensive review of all work is impossible. Instead, we provide a personalised overview into the areas of network epidemiology that have seen the greatest progress in recent years or have the greatest potential to provide novel insights. As such, considerable importance is placed on analytical approaches and statistical methods which are both rapidly expanding fields. Throughout this review we restrict our attention to epidemiological issues

Effects of some anti-diabetic plants on the hepatic marker enzymes of diabetic rats

This study was embarked upon in order to evaluate the effects of the chloroform extracts of the leaves of Psidium guajava, Anacardium occidentale and Eucalyptus globulus and fruits of Xylopia aethiopica on hepatic marker enzymes of diabetic rats. The degree of hepatic damage caused by diabetes mellitus and the effects of the extracts were assessed using standard methods for assaying the activities of alanine aminotransferase (ALT), aspartate aminotransferase (AST) and alkaline phosphatase (ALP). All the extracts significantly (p<0.05) decreased the activities of ALT, AST and ALP with that of the P. guajava being the most protective. In addition, the P. guajava extract exerted more hepatoprotection than glibenclamide in terms of the AST and ALP activities. In conclusion, the chloroform extracts of the leaves of A. occidentale, E. globulus and P. guajava as well as the fruits of X. aethiopica exhibited remarkable protective effects on alloxan-induced acute liver damage and thus, may be used for treatment of some liver-associated disorders.Keyword: Alanine aminotransferase (ALT), aspartate aminotransferase (AST) and alkaline phosphatase (ALP), Psidium guajava, Anacardium occidentale, Eucalyptus globulus, Xylopia aethiopica, chloroform extracts, diabetes mellitus

2FGL J0846.0+2820: A new neutron star binary with a giant secondary and variable γ\gamma-ray emission

We present optical photometric and spectroscopic observations of the likely stellar counterpart to the unassociated \emph{Fermi}-Large Area Telescope (LAT) $\gamma$-ray source 2FGL J0846.0+2820, selected for study based on positional coincidences of optical variables with unassociated LAT sources. Using optical spectroscopy from the SOAR telescope, we have identified a late-G giant in an eccentric ($e$ = 0.06) 8.133 day orbit with an invisible primary. Modeling the spectroscopy and photometry together lead us to infer a heavy neutron star primary of $\sim 2 M_{\odot}$ and a partially stripped giant secondary of $\sim 0.8 M_{\odot}$. H$\alpha$ emission is observed in some of the spectra, perhaps consistent with the presence of a faint accretion disk. We find the $\gamma$-ray flux of 2FGL J0846.0+2820 dropped substantially in mid-2009, accompanied by an increased variation in the optical brightness, and since then it has not been detected by \emph{Fermi}. The long period and giant secondary are reminiscent of the $\gamma$-ray bright binary 1FGL J1417.7--4407, which hosts a millisecond pulsar apparently in the final stages of the pulsar recycling process. The discovery of 2FGL J0846.0+2820 suggests the identification of a new subclass of millisecond pulsar binaries that are the likely progenitors of typical field millisecond pulsars.Comment: 12 pages, 8 figures, 3 tables. Accepted for publication in Ap