Chemical Composition and Sulfur Speciation in Bulk Tissue by X-Ray Spectroscopy and X-Ray Microscopy: Corneal Development during Embryogenesis

The chemical composition and sulfur (S) speciation of developing chick corneas at embryonic days 12, 14, and 16 were investigated using synchrotron scanning x-ray fluorescence microscopy and x-ray absorption near-edge structure spectroscopy. The aim was to develop techniques for the analysis of bulk tissue and identify critical physicochemical variations that correlate with changes in corneal structure-function relationships. Derived data were subjected to principal component analysis and linear discriminant analysis, which highlighted differences in the elemental and S species composition at different stages of embryonic growth. Notably, distinct elemental compositions of chlorine, potassium, calcium, phosphorus, and S altered with development during the transition of the immature opaque cornea to a mature transparent tissue. S  structure spectroscopy revealed developmentally regulated alterations in thiols, organic monosulfides, ester sulfate, and inorganic sulfate species. The transient molecular structures and compositional changes reported here provide a deeper understanding of the underlying basis of corneal development during the acquisition of transparency. The experimental and analytical approach is new, to our knowledge, and has wide potential applicability in the life sciences.     

Lipid organization regulates annexin A2 Ca2 +-sensitivity for membrane bridging and its modulator effects on membrane fluidity

Annexin A2 (AnxA2) is a phospholipid binding protein that has been implicated in many membrane-related cellular functions. AnxA2 is able to bind different acidic phospholipids such as phosphatidylserine (PS) and phosphatidylinositol-4,5-bisphosphate (PI2P). This binding is mediated by Ca^2 +-dependent and Ca^2 +-independent mechanisms. The specific functions of annexin A2 related to these two phospholipids and the molecular mechanisms involved in their interaction remain obscure. Herein we studied the influence of lipid composition on the Ca^2 +-dependency of AnxA2-mediated membrane bridging and on membrane fluidity. Membrane models of ten different lipid compositions and detergent-resistant membranes from two cellular sources were investigated. The results show that the AnxA2-mediated membrane bridging requires 3 to 50 times less calcium for PS-membranes than for PI2P-membranes. Membrane fluidity was measured by the ratiometric fluorescence parameter generalized polarization method with two fluorescent probes. Compared to controls containing low phospholipid ligand, AnxA2 was found to reduce the membrane fluidity of PI2P-membranes twice as much as the PS-membranes in the presence of calcium. On the contrary, at mild acidic pH in the absence of calcium AnxA2 reduces the fluidity of the PS-membranes more than the PI2P-membranes. The presence of cholesterol on the bilayer reduced the AnxA2 capacity to reduce membrane fluidity. The presented data shed light on the specific roles of PI2P, PS and cholesterol present on membranes related to the action of annexin A2 as a membrane bridging molecule during exocytosis and endocytosis events and as a plasma membrane domain phospholipid packing regulator.     

BRCA1 promoter methylation in peripheral blood DNA was identified in sporadic breast cancer and controls

Objective Epigenetics, particularly DNA methylation, has recently been shown to be important in breast cancer initiation. We investigated the clinical and prognostic importance of whole blood breast cancer early onset gene 1 (BRCA1) DNA methylation in sporadic breast cancer. Methods Genomic DNA was extracted from the peripheral blood cells (PBCs) of 902 breast cancer patients at diagnosis, with no BRCA1 mutation, and 990 control women. DNA methylation was measured by quantitative analysis of methylated alleles (QAMA) to estimate the extent of methylation of 2 CpG sites in the promoter region of BRCA1 oncosuppressor. Results BRCA1 promoter methylation rate in PBCs was 47.1% with a 95% confidence interval [46.1; 48.1] in breast cancer patients, and 45.9% with a 95% confidence interval [45.0; 46.8] in controls. We found a trend toward BRCA1 promoter hypermethylation in PBCs of sporadic breast cancer patients compared with controls. Association between methylation and clinicopathological features was evaluated using statistical tests. BRCA1 promoter methylation in PBCs increased significantly in breast cancer patients compared with controls, for age over 70 years (p = 0.022), in post-menopausal status (p = 0.013), for a body mass index (BMI) <20 (p = 0.0095) or a waist-to-hip ratio (WHR) ≤76.8 (p = 0.0027). We also found an association of increased BRCA1 promoter methylation in PBCs with ACA/ACA genotype for the SNP Thr594Thr in ESR (estrogen receptor gene), known to be associated with breast cancer risk (p = 0.092), reflecting the reduced presence of this genotype in this breast cancer case-control study. Conclusion Analysis of site-specific DNA methylation in PBCs by QAMA provides quantitative DNA methylation values that may serve as important prognostic indicators.     

Uncovering the origin of the black stains in Lascaux Cave in France

Lascaux Cave in France was discovered in 1940. Since being opened to visitors the cave has suffered three major microbial outbreaks. The current problem is the fast dissemination of black stains which are threatening the Palaeolithic paintings. Previous data pointed to the involvement of new fungal species in the formation of black stains on the rock walls and ceiling. However, it appears that there could be other reasons for the formation of different and extensive black stains coating the surface of the clayey sediments. Our analyses reveal that black stains on clayey sediments are mainly produced by Acremonium nepalense, a manganese oxide‐depositing fungus, widely distributed in the cave. Thus, in Lascaux Cave, the black stains have a dual origin: on limestone rocks they are mainly produced by the accumulation of fungal melanins, and on clayey sediments by the biogenic deposition of black manganese oxides.     

How to Handle Speciose Clades? Mass Taxon-Sampling as a Strategy towards Illuminating the Natural History of Campanula (Campanuloideae)

Background

Speciose clades usually harbor species with a broad spectrum of adaptive strategies and complex distribution patterns, and thus constitute ideal systems to disentangle biotic and abiotic causes underlying species diversification. The delimitation of such study systems to test evolutionary hypotheses is difficult because they often rely on artificial genus concepts as starting points. One of the most prominent examples is the bellflower genus Campanula with some 420 species, but up to 600 species when including all lineages to which Campanula is paraphyletic. We generated a large alignment of petD group II intron sequences to include more than 70% of described species as a reference. By comparison with partial data sets we could then assess the impact of selective taxon sampling strategies on phylogenetic reconstruction and subsequent evolutionary conclusions.

Methodology/Principal Findings

Phylogenetic analyses based on maximum parsimony (PAUP, PRAP), Bayesian inference (MrBayes), and maximum likelihood (RAxML) were first carried out on the large reference data set (D680). Parameters including tree topology, branch support, and age estimates, were then compared to those obtained from smaller data sets resulting from “classification-guided” (D088) and “phylogeny-guided sampling” (D101). Analyses of D088 failed to fully recover the phylogenetic diversity in Campanula, whereas D101 inferred significantly different branch support and age estimates.

Conclusions/Significance

A short genomic region with high phylogenetic utility allowed us to easily generate a comprehensive phylogenetic framework for the speciose Campanula clade. Our approach recovered 17 well-supported and circumscribed sub-lineages. Knowing these will be instrumental for developing more specific evolutionary hypotheses and guide future research, we highlight the predictive value of a mass taxon-sampling strategy as a first essential step towards illuminating the detailed evolutionary history of diverse clades.     

Wild mouse lemurs revisit artificial feeding platforms: implications for field experiments on sensory and cognitive abilities in small primates

Dealing effectively with space to find important resources in a natural environment is a fundamental ability necessary for survival. Evidence has already been provided that wild gray mouse lemurs revisit stationary feeding sites regularly. In this study, we explore to what extent two sympatric mouse lemur species, Microcebus murinus and M. ravelobensis, revisited artificial feeding sites during a period of food scarcity. As the tested populations are marked with individual transponders, we built up artificial feeding platforms equipped with a transponder reader at nine different locations where mouse lemurs had been previously caught. We baited them with a liquid reward and recorded the visitors' ID, the time and frequency of their visits, as well as all encounters that occurred on the platforms. Only mouse lemurs visited platforms and a total of sixteen individuals across both species were identified. Mouse lemurs visited a platform with a frequency of 2.02 (+/-0.95, range: 1-3.4) times in a night and they revisited it on several consecutive nights following their first visit (percentage of revisits 90.6%+/-11.7, range: 73.3-100%). First visits on a platform occurred on average 44 min (+/-35; range: 13-131) after sunset. We identified encounters between mouse lemurs on platforms: all of them were agonistic and within a species. Within a dyad, chasers were significantly heavier than chasees (N=7 dyads). Our design of platform experiments offers the advantage of observing wild individually known small primates in their natural environment and of setting up controlled experiments to gain insight into their sensory and cognitive abilities.