Low-dose exposure to bisphenols A, F and S of human primary adipocyte impacts coding and non-coding RNA profiles.

Bisphenol A (BPA) exposure has been suspected to be associated with deleterious effects on health including obesity and metabolically-linked diseases. Although bisphenols F (BPF) and S (BPS) are BPA structural analogs commonly used in many marketed products as a replacement for BPA, only sparse toxicological data are available yet. Our objective was to comprehensively characterize bisphenols gene targets in a human primary adipocyte model, in order to determine whether they may induce cellular dysfunction, using chronic exposure at two concentrations: a "low-dose" similar to the dose usually encountered in human biological fluids and a higher dose. Therefore, BPA, BPF and BPS have been added at 10 nM or 10 μM during the differentiation of human primary adipocytes from subcutaneous fat of three non-diabetic Caucasian female patients. Gene expression (mRNA/lncRNA) arrays and microRNA arrays, have been used to assess coding and non-coding RNA changes. We detected significantly deregulated mRNA/lncRNA and miRNA at low and high doses. Enrichment in "cancer" and "organismal injury and abnormalities" related pathways was found in response to the three products. Some long intergenic non-coding RNAs and small nucleolar RNAs were differentially expressed suggesting that bisphenols may also activate multiple cellular processes and epigenetic modifications. The analysis of upstream regulators of deregulated genes highlighted hormones or hormone-like chemicals suggesting that BPS and BPF can be suspected to interfere, just like BPA, with hormonal regulation and have to be considered as endocrine disruptors. All these results suggest that as BPA, its substitutes BPS and BPF should be used with the same restrictions

Characterization of a Bvg-regulated fatty acid methyl-transferase in Bordetella pertussis.

The whooping cough agent Bordetella pertussis controls the expression of its large virulence regulon in a coordinated manner through the two-component signal transduction system BvgAS. In addition to the genes coding for bona fide virulence factors, the Bvg regulon comprises genes of unknown function. In this work, we characterized a new Bvg-activated gene called BP2936. Homologs of BP2936 are found in other pathogenic Bordetellae and in several other species, including plant pathogens and environmental bacteria. We showed that the gene product of BP2936 is a membrane-associated methyl-transferase of free fatty acids. We thus propose to name it FmtB, for fatty acid methyl-transferase of Bordetella. The role of this protein was tested in cellular and animal models of infection, but the loss of BP2936 did not appear to affect host-pathogen interactions in those assays. The high level of conservation of BP2936 among B. pertussis isolates nevertheless argues that it probably plays a role in the life cycle of this pathogen

What Is the Best NGS Enrichment Method for the Molecular Diagnosis of Monogenic Diabetes and Obesity?

Molecular diagnosis of monogenic diabetes and obesity is of paramount importance for both the patient and society, as it can result in personalized medicine associated with a better life and it eventually saves health care spending. Genetic clinical laboratories are currently switching from Sanger sequencing to next-generation sequencing (NGS) approaches but choosing the optimal protocols is not easy. Here, we compared the sequencing coverage of 43 genes involved in monogenic forms of diabetes and obesity, and variant detection rates, resulting from four enrichment methods based on the sonication of DNA (Agilent SureSelect, RainDance technologies), or using enzymes for DNA fragmentation (Illumina Nextera, Agilent HaloPlex). We analyzed coding exons and untranslated regions of the 43 genes involved in monogenic diabetes and obesity. We found that none of the methods achieves yet full sequencing of the gene targets. Nonetheless, the RainDance, SureSelect and HaloPlex enrichment methods led to the best sequencing coverage of the targets; while the Nextera method resulted in the poorest sequencing coverage. Although the sequencing coverage was high, we unexpectedly found that the HaloPlex method missed 20% of variants detected by the three other methods and Nextera missed 10%. The question of which NGS technique for genetic diagnosis yields the highest diagnosis rate is frequently discussed in the literature and the response is still unclear. Here, we showed that the RainDance enrichment method as well as SureSelect, which are both based on the sonication of DNA, resulted in a good sequencing quality and variant detection, while the use of enzymes to fragment DNA (HaloPlex or Nextera) might not be the best strategy to get an accurate sequencing

Experimental design and analysis workflow.

<p>BPA, BPS, BPF and DMSO: respectively Bisphenol A, S and F and dimethyl sulfoxide.</p

Numbers of differentially down and up-regulated miRNA probes amongst the 483 probes after the exposition of the pre-adipocytes to 10 nM or 10 μM of bisphenol shared between the concentrations for each bisphenol.

<p>Numbers of differentially down and up-regulated miRNA probes amongst the 483 probes after the exposition of the pre-adipocytes to 10 nM or 10 μM of bisphenol shared between the concentrations for each bisphenol.</p

List of differentially expressed probes deregulated for the three bisphenols A, F and S at 10 nM.

<p>List of differentially expressed probes deregulated for the three bisphenols A, F and S at 10 nM.</p

Enrichment analysis using Ingenuity Pathway Analysis on the lists of mRNA probes differentially expressed after exposition to the three bisphenols A, F and S at 10 nM and 10 μM.

<p>Enrichment analysis using Ingenuity Pathway Analysis on the lists of mRNA probes differentially expressed after exposition to the three bisphenols A, F and S at 10 nM and 10 μM.</p

Upstream regulators provided by Ingenuity Pathway Analysis in the 10 genes differentially expressed at both concentrations (10 nM and 10 μM) for the three bisphenols (A, F and S).

<p>Upstream regulators provided by Ingenuity Pathway Analysis in the 10 genes differentially expressed at both concentrations (10 nM and 10 μM) for the three bisphenols (A, F and S).</p

Heatmap representation of significantly differentially expressed genes in the three bisphenols at 10 nM and 10 μM.

<p>The represented values are log₂ fold changes of the ratio of the expression in a given bisphenol and concentration over the control condition which is differentiated without any bisphenol. Genes (probes) highlighted in pink are significant for all concentrations of all bisphenols.</p

Number of differentially down and up-regulated miRNA probes amongst the 483 probes after the exposition of the pre-adipocytes to 10 nM or 10 μM of bisphenol (A, F and S) during differentiation shared between BPA and BPS or BPF.

<p>Number of differentially down and up-regulated miRNA probes amongst the 483 probes after the exposition of the pre-adipocytes to 10 nM or 10 μM of bisphenol (A, F and S) during differentiation shared between BPA and BPS or BPF.</p