Adsorption at cell surface and cellular uptake of silica nanoparticles with different surface chemical functionalizations: impact on cytotoxicity

International audienceSilica nanoparticles are particularly interesting for medical applications because of the high inertness and chemical stability of silica material. However, at the nanoscale their innocuousness must be carefully verified before clinical use. The aim of this study was to investigate the in vitro biological toxicity of silica nanoparticles depending on their surface chemical functionalization. To that purpose, three kinds of 50 nm fluorescent silica-based nanoparticles were synthesized: 1) sterically stabilized silica nanoparticles coated with neutral polyethylene glycol (PEG) molecules, 2) positively charged silica nanoparticles coated with amine groups and 3) negatively charged silica nanoparticles coated with carboxylic acid groups. RAW 264.7 murine macrophages were incubated for 20 hours with each kind of nanoparticles. Their cellular uptake and adsorption at the cell membrane were assessed by a fluorimetric assay and cellular responses were evaluated in terms of cytotoxicity, pro-inflammatory factor production and oxidative stress. Results showed that the highly positive charged nanoparticle, were the most adsorbed at cell surface and triggered more cytotoxicity than other nanoparticles types. To conclude, this study clearly demonstrated that silica nanoparticles surface functionalization represents a key parameter in their cellular uptake and biological toxicity

Analysis of complete genome sequence of Neorickettsia risticii: causative agent of Potomac horse fever

Neorickettsia risticii is an obligate intracellular bacterium of the trematodes and mammals. Horses develop Potomac horse fever (PHF) when they ingest aquatic insects containing encysted N. risticii-infected trematodes. The complete genome sequence of N. risticii Illinois consists of a single circular chromosome of 879 977 bp and encodes 38 RNA species and 898 proteins. Although N. risticii has limited ability to synthesize amino acids and lacks many metabolic pathways, it is capable of making major vitamins, cofactors and nucleotides. Comparison with its closely related human pathogen N. sennetsu showed that 758 (88.2%) of protein-coding genes are conserved between N. risticii and N. sennetsu. Four-way comparison of genes among N. risticii and other Anaplasmataceae showed that most genes are either shared among Anaplasmataceae (525 orthologs that generally associated with housekeeping functions), or specific to each genome (>200 genes that are mostly hypothetical proteins). Genes potentially involved in the pathogenesis of N. risticii were identified, including those encoding putative outer membrane proteins, two-component systems and a type IV secretion system (T4SS). The bipolar localization of T4SS pilus protein VirB2 on the bacterial surface was demonstrated for the first time in obligate intracellular bacteria. These data provide insights toward genomic potential of N. risticii and intracellular parasitism, and facilitate our understanding of PHF pathogenesis

Nuclear Receptor HNF4α Binding Sequences are Widespread in Alu Repeats

<p>Abstract</p> <p>Background</p> <p>Alu repeats, which account for ~10% of the human genome, were originally considered to be junk DNA. Recent studies, however, suggest that they may contain transcription factor binding sites and hence possibly play a role in regulating gene expression.</p> <p>Results</p> <p>Here, we show that binding sites for a highly conserved member of the nuclear receptor superfamily of ligand-dependent transcription factors, hepatocyte nuclear factor 4alpha (HNF4α, NR2A1), are highly prevalent in Alu repeats. We employ high throughput protein binding microarrays (PBMs) to show that HNF4α binds > 66 unique sequences in Alu repeats that are present in ~1.2 million locations in the human genome. We use chromatin immunoprecipitation (ChIP) to demonstrate that HNF4α binds Alu elements in the promoters of target genes (<it>ABCC3, APOA4, APOM, ATPIF1, CANX, FEMT1A, GSTM4, IL32, IP6K2, PRLR, PRODH2, SOCS2, TTR</it>) and luciferase assays to show that at least some of those Alu elements can modulate HNF4α-mediated transactivation <it>in vivo </it>(<it>APOM, PRODH2, TTR, APOA4</it>). HNF4α-Alu elements are enriched in promoters of genes involved in RNA processing and a sizeable fraction are in regions of accessible chromatin. Comparative genomics analysis suggests that there may have been a gain in HNF4α binding sites in Alu elements during evolution and that non Alu repeats, such as Tiggers, also contain HNF4α sites.</p> <p>Conclusions</p> <p>Our findings suggest that HNF4α, in addition to regulating gene expression via high affinity binding sites, may also modulate transcription via low affinity sites in Alu repeats.</p

The transcriptional activity of hepatocyte nuclear factor 4 alpha is inhibited via phosphorylation by ERK1/2

Hepatocyte nuclear factor 4 alpha (HNF4alpha) nuclear receptor is a master regulator of hepatocyte development, nutrient transport and metabolism. HNF4alpha is regulated both at the transcriptional and post-transcriptional levels by different mechanisms. Several kinases (PKA, PKC, AMPK) were shown to phosphorylate and decrease the activity of HNF4alpha. Activation of the ERK1/2 signalling pathway, inducing proliferation and survival, inhibits the expression of HNF4alpha. However, based on our previous results we hypothesized that HNF4alpha is also regulated at the post-transcriptional level by ERK1/2. Here we show that ERK1/2 is capable of directly phosphorylating HNF4alpha in vitro at several phosphorylation sites including residues previously shown to be targeted by other kinases, as well. Furthermore, we also demonstrate that phosphorylation of HNF4alpha leads to a reduced trans-activational capacity of the nuclear receptor in luciferase reporter gene assay. We confirm the functional relevance of these findings by demonstrating with ChIP-qPCR experiments that 30-minute activation of ERK1/2 leads to reduced chromatin binding of HNF4alpha. Accordingly, we have observed decreasing but not disappearing binding of HNF4alpha to the target genes. In addition, 24-hour activation of the pathway further decreased HNF4alpha chromatin binding to specific loci in ChIP-qPCR experiments, which confirms the previous reports on the decreased expression of the HNF4a gene due to ERK1/2 activation. Our data suggest that the ERK1/2 pathway plays an important role in the regulation of HNF4alpha-dependent hepatic gene expression

Bagging Statistical Network Inference from Large-Scale Gene Expression Data

Modern biology and medicine aim at hunting molecular and cellular causes of biological functions and diseases. Gene regulatory networks (GRN) inferred from gene expression data are considered an important aid for this research by providing a map of molecular interactions. Hence, GRNs have the potential enabling and enhancing basic as well as applied research in the life sciences. In this paper, we introduce a new method called BC3NET for inferring causal gene regulatory networks from large-scale gene expression data. BC3NET is an ensemble method that is based on bagging the C3NET algorithm, which means it corresponds to a Bayesian approach with noninformative priors. In this study we demonstrate for a variety of simulated and biological gene expression data from S. cerevisiae that BC3NET is an important enhancement over other inference methods that is capable of capturing biochemical interactions from transcription regulation and protein-protein interaction sensibly. An implementation of BC3NET is freely available as an R package from the CRAN repository

Regulation of Clock-Controlled Genes in Mammals

The complexity of tissue- and day time-specific regulation of thousands of clock-controlled genes (CCGs) suggests that many regulatory mechanisms contribute to the transcriptional output of the circadian clock. We aim to predict these mechanisms using a large scale promoter analysis of CCGs

Supersymmetric QCD corrections to e+e−→tbˉH−e^+e^-\to t\bar{b}H^- and the Bernstein-Tkachov method of loop integration

The discovery of charged Higgs bosons is of particular importance, since their existence is predicted by supersymmetry and they are absent in the Standard Model (SM). If the charged Higgs bosons are too heavy to be produced in pairs at future linear colliders, single production associated with a top and a bottom quark is enhanced in parts of the parameter space. We present the next-to-leading-order calculation in supersymmetric QCD within the minimal supersymmetric SM (MSSM), completing a previous calculation of the SM-QCD corrections. In addition to the usual approach to perform the loop integration analytically, we apply a numerical approach based on the Bernstein-Tkachov theorem. In this framework, we avoid some of the generic problems connected with the analytical method.Comment: 14 pages, 6 figures, accepted for publication in Phys. Rev.

Phylogenomic analyses of malaria parasites and evolution of their exported proteins

<p>Abstract</p> <p>Background</p> <p><it>Plasmodium falciparum </it>is the most malignant agent of human malaria. It belongs to the taxon Laverania, which includes other ape-infecting <it>Plasmodium </it>species. The origin of the Laverania is still debated. <it>P. falciparum </it>exports pathogenicity-related proteins into the host cell using the <it>Plasmodium </it>export element (PEXEL). Predictions based on the presence of a PEXEL motif suggest that more than 300 proteins are exported by <it>P. falciparum</it>, while there are many fewer exported proteins in non-Laverania.</p> <p>Results</p> <p>A whole-genome approach was applied to resolve the phylogeny of eight <it>Plasmodium </it>species and four outgroup taxa. By using 218 orthologous proteins we received unanimous support for a sister group position of Laverania and avian malaria parasites. This observation was corroborated by the analyses of 28 exported proteins with orthologs present in all <it>Plasmodium </it>species. Most interestingly, several deviations from the <it>P. falciparum </it>PEXEL motif were found to be present in the orthologous sequences of non-Laverania.</p> <p>Conclusion</p> <p>Our phylogenomic analyses strongly support the hypotheses that the Laverania have been founded by a single <it>Plasmodium </it>species switching from birds to African great apes or <it>vice versa</it>. The deviations from the canonical PEXEL motif in orthologs may explain the comparably low number of exported proteins that have been predicted in non-Laverania.</p

Maintaining Integrity Under Stress:Envelope Stress Response Regulation of Pathogenesis in Gram-Negative Bacteria

The Gram-negative bacterial envelope is an essential interface between the intracellular and harsh extracellular environment. Envelope stress responses (ESRs) are crucial to the maintenance of this barrier and function to detect and respond to perturbations in the envelope, caused by environmental stresses. Pathogenic bacteria are exposed to an array of challenging and stressful conditions during their lifecycle and, in particular, during infection of a host. As such, maintenance of envelope homeostasis is essential to their ability to successfully cause infection. This review will discuss our current understanding of the σE- and Cpx-regulated ESRs, with a specific focus on their role in the virulence of a number of model pathogens