Pyrrolopyrimidine inhibitors of DNA gyrase B (GyrB) and topoisomerase IV (ParE). Part I: Structure guided discovery and optimization of dual targeting agents with potent,broad-spectrum enzymatic activity

The bacterial topoisomerases DNA gyrase (GyrB) and topoisomerase IV (ParE) are essential enzymes that control the topological state of DNA during replication. The high degree of conservation in the ATP-binding pockets of these enzymes make them appealing targets for broad-spectrum inhibitor development. A pyrrolopyrimidine scaffold was identified from a pharmacophore-based fragment screen with optimization potential. Structural characterization of inhibitor complexes conducted using selected GyrB/ParE orthologs aided in the identification of important steric, dynamic and compositional differences in the ATP-binding pockets of the targets, enabling the design of highly potent pyrrolopyrimidine inhibitors with broad enzymatic spectrum and dual targeting activity.     

Coupling Demographic and Genetic Variability from Archived Collections of European Anchovy (Engraulis encrasicolus)

It is well known that temporal fluctuations in small populations deeply influence evolutionary potential. Less well known is whether fluctuations can influence the evolutionary potentials of species with large census sizes. Here, we estimated genetic population parameters from as survey of polymorphic microsatellite DNA loci in archived otoliths from Adriatic European anchovy (Engraulis encrasicolus), a fish with large census sizes that supports numerous local fisheries. Stocks have fluctuated greatly over the past few decades, and the Adriatic fishery collapsed in 1987. Our results show a significant reduction of mean genetic parameters as a consequence of the population collapse. In addition, estimates of effective population size (N_e) are much smaller than those expected in a fishes with large population census sizes (N_c). Estimates of N_e indicate low effective population sizes, even before the population collapse. The ratio N_e/N_e ranged between 10⁻⁶ and 10⁻⁸, indicating a large discrepancy between the anchovy gene pool and population census size. Therefore, anchovy populations may be more vulnerable to fishery effort and environmental change than previously thought.     

Fusing Data Mining,Machine Learning and Traditional Statistics to Detect Biomarkers Associated with Depression

Background

Atheoretical large-scale data mining techniques using machine learning algorithms have promise in the analysis of large epidemiological datasets. This study illustrates the use of a hybrid methodology for variable selection that took account of missing data and complex survey design to identify key biomarkers associated with depression from a large epidemiological study.

Methods

The study used a three-step methodology amalgamating multiple imputation, a machine learning boosted regression algorithm and logistic regression, to identify key biomarkers associated with depression in the National Health and Nutrition Examination Study (2009–2010). Depression was measured using the Patient Health Questionnaire-9 and 67 biomarkers were analysed. Covariates in this study included gender, age, race, smoking, food security, Poverty Income Ratio, Body Mass Index, physical activity, alcohol use, medical conditions and medications. The final imputed weighted multiple logistic regression model included possible confounders and moderators.

Results

After the creation of 20 imputation data sets from multiple chained regression sequences, machine learning boosted regression initially identified 21 biomarkers associated with depression. Using traditional logistic regression methods, including controlling for possible confounders and moderators, a final set of three biomarkers were selected. The final three biomarkers from the novel hybrid variable selection methodology were red cell distribution width (OR 1.15; 95% CI 1.01, 1.30), serum glucose (OR 1.01; 95% CI 1.00, 1.01) and total bilirubin (OR 0.12; 95% CI 0.05, 0.28). Significant interactions were found between total bilirubin with Mexican American/Hispanic group (p = 0.016), and current smokers (p<0.001).

Conclusion

The systematic use of a hybrid methodology for variable selection, fusing data mining techniques using a machine learning algorithm with traditional statistical modelling, accounted for missing data and complex survey sampling methodology and was demonstrated to be a useful tool for detecting three biomarkers associated with depression for future hypothesis generation: red cell distribution width, serum glucose and total bilirubin.     

Retinoic Acid Specifically Enhances Embryonic Stem Cell Metastate Marked by Zscan4

Pluripotency confers Embryonic Stem Cells (ESCs) the ability to differentiate in ectoderm, endoderm, and mesoderm derivatives, producing the majority of cell types. Although the majority of ESCs divide without losing pluripotency, it has become evident that ESCs culture consists of multiple cell populations with different degrees of potency that are spontaneously induced in regular ESC culture conditions. Zscan4, a key pluripotency factor, marks ESC subpopulation that is referred to as high-level of pluripotency metastate. Here, we report that in ESC cultures treated with retinoic acid (RA), Zscan4 ESCs metastate is strongly enhanced. In particular, we found that induction of Zscan4 metastate is mediated via RA receptors (RAR-alpha, RAR-beta, and RAR-gamma), and it is dependent on phosphoinositide-3-kinase (PI3K) signaling. Remarkably, Zscan4 metastate induced by RA lacks canonical pluripotency genes Oct3/4 and Nanog but retained both self-renewal and pluripotency capabilities. Finally we demonstrated that the conditional ablation of Zscan4 subpopulation is dispensable for both endoderm and mesoderm but is required for ectoderm lineage. In conclusion, our research provides new insights about the role of RA signaling during ESCs high pluripotency metastate fluctuation.     

Identification of a cDNA clone encoding DIP1-binding protein in Drosophila melanogaster

The Drosophila melanogaster L27a gene encodes a ribosomal protein which is a member of the L15 family of ribosomal proteins. D.m. L27a is closely related to the mammalian protein that has been found differentially expressed in lung cancer tissues and therefore could be involved in the control of cell proliferation such as the ribosomal protein S6. Our work elucidates the role of DIP1 which is a novel protein that we found in Drosophila. We performed a two-hybrid system assay and identified the L27a protein as an interactor of DIP1. The interaction was then validated by in vitro binding assays. DIP1, similar to other nuclear proteins in eukaryotes, is localized to the nuclear periphery and chromatin domain in all nuclei, but disappears at the metaphase. It is possible that in D.m. L27a protein, via interaction with DIP1, could be involved in protein synthesis as well as in cell cycle regulation.     

Targeted modification of homogalacturonan by transgenic expression of a fungal polygalacturonase alters plant growth

Pectins are a highly complex family of cell wall polysaccharides comprised of homogalacturonan (HGA), rhamnogalacturonan I and rhamnogalacturonan II. We have specifically modified HGA in both tobacco (Nicotiana tabacum) and Arabidopsis by expressing the endopolygalacturonase II of Aspergillus niger (AnPGII). Cell walls of transgenic tobacco plants showed a 25% reduction in GalUA content as compared with the wild type and a reduced content of deesterified HGA as detected by antibody labeling. Neutral sugars remained unchanged apart from a slight increase of Rha, Ara, and Gal. Both transgenic tobacco and Arabidopsis were dwarfed, indicating that unesterified HGA is a critical factor for plant cell growth. The dwarf phenotypes were associated with AnPGII activity as demonstrated by the observation that the mutant phenotype of tobacco was completely reverted by crossing the dwarfed plants with plants expressing PGIP2, a strong inhibitor of AnPGII. The mutant phenotype in Arabidopsis did not appear when transformation was performed with a gene encoding AnPGII inactivated by site directed mutagenesis.