Biotransformation of Natural and Synthetic Isoflavonoids by Two Recombinant Microbial Enzymes

Isolation and synthesis of isoflavonoids has become a frequent endeavor, due to their interesting biological activities. The introduction of hydroxyl groups into isoflavonoids by the use of enzymes represents an attractive alternative to conventional chemical synthesis. In this study, the capabilities of biphenyl-2,3-dioxygenase (BphA) and biphenyl-2,3-dihydrodiol 2,3-dehydrogenase (BphB) of Burkholderia sp. strain LB400 to biotransform 14 isoflavonoids synthesized in the laboratory were investigated by using recombinant Escherichia coli strains containing plasmid vectors expressing the bphA1A2A3A4 or bphA1A2A3A4B genes of strain LB400. The use of BphA and BphB allowed us to biotransform 7-hydroxy-8-methylisoflavone and 7-hydroxyisoflavone into 7,2′,3′-trihydroxy-8-methylisoflavone and 7,3′,4′-trihydroxyisoflavone, respectively. The compound 2′-fluoro-7-hydroxy-8-methylisoflavone was dihydroxylated by BphA at ortho-fluorinated and meta positions of ring B, with concomitant dehalogenation leading to 7,2′,3′,-trihydroxy-8-methylisoflavone. Daidzein (7,4′-dihydroxyisoflavone) was biotransformed by BphA, generating 7,2′,4′-trihydroxyisoflavone after dehydration. Biotransformation products were analyzed by gas chromatography-mass spectrometry and nuclear magnetic resonance techniques.     

Cancer du testicule: BEP et spermatogenèse

The sperm characteristics of 44 men treated with two or more courses of BEP chemotherapy for non seminomatous germ cell testicular tumours were investigated before and 25.2 ± 19.4 months after chemotherapy. Before treatment, 54.5% of patients were oligozoospermic. The mean sperm characteristics did not differ before and after chemotherapy. However, following chemotherapy, the recovery of initial sperm count was more frequent after one year than before. During the first year, recovery was more frequent in patients treated with two than in those treated with more than two BEP cycles. In patients with good pre-treatment sperm count, sperm production was reduced by half after chemotherapy. In a subgroup of men who provided two sperm samples after chemotherapy, sperm production was better in the second sample than in the first. Our data suggest that sperm recovery is related to testicular function prior to therapy, to the time elapsed after chemotherapy and in the first year to the number of chemotherapy cycles. In conclusion, our study is reassuring concerning the long-term male reproductive toxicity of BEP. However, further studies are required to analyse the possible effects on sperm genetic material during the recovery period.     

Effects of age and distance on the composition of mixed deciduous forest fragments in an agricultural landscape

Abstract. Forest patches in central Belgium were inventoried twice for the presence or absence of forest plant species to study the effects of age and distance on species composition. All forests in the study area were subdivided based on their land use history. To avoid effects of autocorrelated environmental characteristics on species composition, habitat homogeneity was indirectly investigated using a TWINSPAN classification of the vegetation data. Two major habitats (alluvial and non‐alluvial forests) were distinguished and analysed separately. Patterns of species composition were investigated at the landscape level using Mantel tests. Species composition similarity measures were calculated between all pairs of fragments based on the floristic data. A highly significant correlation was found between species composition similarity and inter‐patch distance. Difference in age, which we used as a measure for habitat quality, was less important in explaining species composition patterns. The effects of spatial configuration became significant when difference in age was accounted for, and a partial correlation was calculated between inter‐patch distance and species composition similarity. Different results were found for alluvial and non‐alluvial forest types. Alluvial forests were more influenced by the spatial configuration than the non‐alluvial. For the non‐alluvial forest type effects measured with the difference in age between forest fragments obscured the effects of inter‐patch distance. Based on our findings we suggest that species composition is not only internally generated, but external processes such as differential colonization caused by varying degrees of isolation may be of overriding importance.     

Partial replacement of sodium in meat and fish products by using magnesium salts. A review

Sodium intake exceeds the nutritional recommendations in most industrialized countries becoming one concern for public health. This elimination or reduction is not simple due to its role in final food sensory, quality and safety. The aim of this work is to review the possibilities of magnesium ion, due to its healthy properties, to become a partial substitute of sodium in the production of fish and meat products, and a particular case for Spanish dry-cured ham and loin. Magnesium diffusion into different muscle based foods such as ham or loin, and its effect in the most important characteristics of the final product (microbiology, physico-chemical and sensory properties) has been analyzed. Results show that magnesium has more difficulty to penetrate inside the muscle and slightly modifies the water-holding capacity of proteins, their solubility and the enzymatic activity. Salty taste, bitterness and off-flavor are the most affected characteristics. However, these effects could be compensated by using longer post-salting periods and by employing masking agents. It is possible to reduce the sodium content in fish and meat products using magnesium as one of the ingredients, allowing to obtain new products with similar physicochemical characteristics and safety conditions.     

Akt activity protects rheumatoid synovial fibroblasts from Fas-induced apoptosis by inhibition of Bid cleavage

Introduction  

Synovial hyperplasia is a main feature of rheumatoid arthritis pathology that leads to cartilage and bone damage in the inflamed joints. Impaired apoptosis of resident synoviocytes is pivotal in this process. Apoptosis resistance seems to involve defects in the extrinsic and intrinsic apoptotic pathways. The aim of this study was to investigate the association of PI3Kinase/Akt and the mitochondrial apoptotic pathway in the resistance of rheumatoid arthritis (RA) fibroblast like synovial cells (FLS) to Fas-mediated apoptosis.     

Proteomic Profiling of the TRAF3 Interactome Network Reveals a New Role for the ER-to-Golgi Transport Compartments in Innate Immunity

Tumor Necrosis Factor receptor-associated factor-3 (TRAF3) is a central mediator important for inducing type I interferon (IFN) production in response to intracellular double-stranded RNA (dsRNA). Here, we report the identification of Sec16A and p115, two proteins of the ER-to-Golgi vesicular transport system, as novel components of the TRAF3 interactome network. Notably, in non-infected cells, TRAF3 was found associated with markers of the ER-Exit-Sites (ERES), ER-to-Golgi intermediate compartment (ERGIC) and the cis-Golgi apparatus. Upon dsRNA and dsDNA sensing however, the Golgi apparatus fragmented into cytoplasmic punctated structures containing TRAF3 allowing its colocalization and interaction with Mitochondrial AntiViral Signaling (MAVS), the essential mitochondria-bound RIG-I-like Helicase (RLH) adaptor. In contrast, retention of TRAF3 at the ER-to-Golgi vesicular transport system blunted the ability of TRAF3 to interact with MAVS upon viral infection and consequently decreased type I IFN response. Moreover, depletion of Sec16A and p115 led to a drastic disorganization of the Golgi paralleled by the relocalization of TRAF3, which under these conditions was unable to associate with MAVS. Consequently, upon dsRNA and dsDNA sensing, ablation of Sec16A and p115 was found to inhibit IRF3 activation and anti-viral gene expression. Reciprocally, mild overexpression of Sec16A or p115 in Hec1B cells increased the activation of IFNβ, ISG56 and NF-κB -dependent promoters following viral infection and ectopic expression of MAVS and Tank-binding kinase-1 (TBK1). In line with these results, TRAF3 was found enriched in immunocomplexes composed of p115, Sec16A and TBK1 upon infection. Hence, we propose a model where dsDNA and dsRNA sensing induces the formation of membrane-bound compartments originating from the Golgi, which mediate the dynamic association of TRAF3 with MAVS leading to an optimal induction of innate immune responses.     

Functional regulation of the structure-specific endonuclease FEN1 by the human cytomegalovirus protein IE1 suggests a role for the re-initiation of stalled viral replication forks

Flap endonuclease 1 (FEN1) is a member of the family of structure-specific endonucleases implicated in regulation of DNA damage response and DNA replication. So far, knowledge on the role of FEN1 during viral infections is limited. Previous publications indicated that poxviruses encode a conserved protein that acts in a manner similar to FEN1 to stimulate homologous recombination, double-strand break (DSB) repair and full-size genome formation. Only recently, cellular FEN1 has been identified as a key component for hepatitis B virus cccDNA formation. Here, we report on a novel functional interaction between Flap endonuclease 1 (FEN1) and the human cytomegalovirus (HCMV) immediate early protein 1 (IE1). Our results provide evidence that IE1 manipulates FEN1 in an unprecedented manner: we observed that direct IE1 binding does not only enhance FEN1 protein stability but also phosphorylation at serine 187. This correlates with nucleolar exclusion of FEN1 stimulating its DSB-generating gap endonuclease activity. Depletion of FEN1 and inhibition of its enzymatic activity during HCMV infection significantly reduced nascent viral DNA synthesis demonstrating a supportive role for efficient HCMV DNA replication. Furthermore, our results indicate that FEN1 is required for the formation of DSBs during HCMV infection suggesting that IE1 acts as viral activator of FEN1 in order to re-initiate stalled replication forks. In summary, we propose a novel mechanism of viral FEN1 activation to overcome replication fork barriers at difficult-to-replicate sites in viral genomes.