A catalytic, practical and high-yielding procedure for the synthesis of indenes by direct C_sp3–H activation under gold(i) catalysis was developed.

Background: We previously showed that a VLDL- and LDL-rich mix of human native lipoproteins induces a set of repressive epigenetic marks,i.e. de novoDNA methylation, histone 4 hypoacetylation and histone 4 lysine 20 (H4K20) hypermethylation in THP-1 macrophages. Here, we: 1) ask what gene expression changes accompany these epigenetic responses; 2) test the involvement of candidate factors mediating the latter. We exploited genome expression arrays to identify target genes for lipoprotein-induced silencing, in addition to RNAi and expression studies to test the involvement of candidate mediating factors. The study was conducted in human THP-1 macrophages.Results: Native lipoprotein-inducedde novoDNA methylation was associated with a general repression of various critical genes for macrophage function, including pro-inflammatory genes. Lipoproteins showed differential effects on epigenetic marks, asde novoDNA methylation was induced by VLDL and to a lesser extent by LDL, but not by HDL, and VLDL induced H4K20 hypermethylation, while HDL caused H4 deacetylation. The analysis of candidate factors mediating VLDL-induced DNA hypermethylation revealed that this response was: 1) surprisingly, mediated exclusively by the canonical maintenance DNA methyltransferase DNMT1, and 2) independent of the Dicer/micro-RNA pathway.Conclusions: Our work provides novel insights into epigenetic gene regulation by native lipoproteins. Furthermore, we provide an example of DNMT1 acting as ade novoDNA methyltransferase independently of canonicalde novoenzymes, and show proof of principle thatde novoDNA methylation can occur independently of a functional Dicer/micro-RNA pathway in mammals.

Sporothrixschenckii is one of the etiological agents of sporotrichosis, a worldwide-distributed subcutaneous mycosis. Its cell wall contains a glycoconjugate composed of rhamnose, mannose, glucuronic acid, and proteins, named peptidorhamnomannan, which harbors important Sporothrix-specific immunogenic epitopes. Although the peptidorhamnomannan carbohydrate moiety has been extensively studied, thus far, little is known about the protein core. Here, using LC-MS/MS, we analyzed the S.schenckii peptidorhamnomannan peptide fraction and generated mass signals of 325 proteins, most of them likely to be moonlighting proteins. Among the identified proteins, chaperonin GroEL/Hsp60 and the uncharacterized protein Pap1 were selected for further analysis. Both proteins were heterologously expressed in bacteria, and they showed adhesive properties to the extracellular matrix proteins laminin, elastin, fibrinogen, and fibronectin, although Pap1 also was bound to type-I and type-II collagen. The inoculation of concentrations higher than 40 μg of these proteins, separately, increased immune effectors in the hemolymph of Galleriamellonella larvae and protected animals from an S.schenckii lethal challenge. These observations were confirmed when yeast-like cells, pre-incubated with anti-rHsp60 or anti-rPap1 antibodies were used to inoculate larvae. The animals inoculated with pretreated cells showed increased survival rates when compared to the control groups. In conclusion, we report that Hsp60 and Pap1 are part of the cell wall peptidorhamnomannan, can bind extracellular matrix components, and contribute to the S.schenckii virulence. To our knowledge, this is the first report about moonlighting protein in the S.schenckii cell wall with an important role during the pathogen–host interaction.

Selenium biofortified yeast is the most common dietary Se supplement in human nutrition and in farm animals. Therefore, the production and routine quality control of commercial products are highly demanded. In this work, a simple and cost-effective procedure is proposed for the determination of SeMet and Se(IV) in hydrolyzed yeast, consisting of ion-pair reversed phase separation, post-column hydride generation and Se quantification by atomic emission spectrometry with microwave plasma sustained by nitrogen (HPLC-HG-MP-AES). Freeze-dried biomass was hydrolyzed with methanesulfonic acid; chromatographic separation was performed with a mobile phase containing 0.08% v/v heptafluorobutyric and methanol (92 : 8) at a flow rate of 1 mL min⁻¹; the column effluent was on-line mixed with an alkaline solution of potassium persulfate (K₂S₂O₈ 6% m/v, NaOH 3% m/v), passed through a reaction coil submerged in a water bath at 60 °C, and then 10 M hydrochloric acid was added prior to hydride generation in the MP-AES multimode sample introduction system (NaBH₄ 2% m/v, NaOH 0.3% m/v). The total chromatographic run was accomplished in 5 min and the evaluated on-column quantification limits were 59 ng Se mL⁻¹ for Se(IV) and 0.52 μg mL⁻¹ for SeMet. The procedure was tested using standardized Seleno Excell® high selenium yeast and then applied for the analysis of Saccharomyces cerevisiae biofortified under different fermentation and exposure conditions. The procedure was capable of detecting differences in selenium concentration among cultures and the results were consistent with those obtained while coupling HPLC separation directly to ICP-MS detection.