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Maintenance of genomic stability during eukaryotic cell division relies on the Spindle Assembly Checkpoint (SAC), which has evolved as a surveillance mechanism that monitors kinetochore-microtubule attachment and prevents APC/C-mediated mitotic exit until all chromosomes are properly attached to the mitotic spindle. Reversible protein phosphorylation has long been accredited as a regulatory mechanism of the SAC. Nevertheless, knowledge of how several mitotic kinases act in concert within the signaling pathway to orchestrate SAC function is still emerging. In a recent study, we undertook a comprehensive dissection of the hierarchical framework controlling SAC function in Drosophila cells. We found that Polo lies at the top of the SAC pathway promoting the efficient recruitment of Mps1 to unattached kinetochores. This renders Mps1 fully active to control BubR1 phosphorylation that generates the 3F3/2 phosphoepitope at tensionless kinetochores. We have proposed that Polo is required for SAC function and that the molecular outcome of Mps1-dependent 3F3/2 formation is to promote the association of Cdc20 with BubR1 allowing proper kinetochore recruitment of Cdc20 and efficient assembly of the Mitotic Checkpoint Complex (MCC) required for a sustained SAC response.     

A calcium‐mediated actin redistribution at egg activation in Drosophila

Egg activation is the essential process in which mature oocytes gain the competency to proceed into embryonic development. Many events of egg activation are conserved, including an initial rise of intracellular calcium. In some species, such as echinoderms and mammals, changes in the actin cytoskeleton occur around the time of fertilization and egg activation. However, the interplay between calcium and actin during egg activation remains unclear. Here, we use imaging, genetics, pharmacological treatment, and physical manipulation to elucidate the relationship between calcium and actin in living Drosophila eggs. We show that, before egg activation, actin is smoothly distributed between ridges in the cortex of the dehydrated mature oocytes. At the onset of egg activation, we observe actin spreading out as the egg swells though the intake of fluid. We show that a relaxed actin cytoskeleton is required for the intracellular rise of calcium to initiate and propagate. Once the swelling is complete and the calcium wave is traversing the egg, it leads to a reorganization of actin in a wavelike manner. After the calcium wave, the actin cytoskeleton has an even distribution of foci at the cortex. Together, our data show that calcium resets the actin cytoskeleton at egg activation, a model that we propose to be likely conserved in other species.     

New insights into the functional roles of reactive oxygen species during embryo sac development and fertilization in Arabidopsis thaliana

Previously considered as toxic by-products of aerobic metabolism, reactive oxygen species (ROS) are emerging as essential signaling molecules in eukaryotes. Recent evidence showed that maintenance of ROS homeostasis during female gametophyte development is crucial for embryo sac patterning and fertilization. Although ROS are exclusively detected in the central cell of mature embryo sacs, the study of mutants deficient in ROS homeostasis suggests that controlled oxidative bursts might take place earlier during gametophyte development. Also, a ROS burst that depends on pollination takes place inside the embryo sac. This oxidative response might be required for pollen tube growth arrest and for sperm cell release. In this mini-review, we will focus on new insights into the role of ROS during female gametophyte development and fertilization. Special focus will be made on the mitochondrial Mn-Superoxide dismutase (MSD1), which has been recently reported to be essential for maintaining ROS homeostasis during embryo sac formation.     

Antimicrobial and antiviral activity-guided fractionation from Scutia buxifolia Reissek extracts

Antimicrobial and antiviral activities of the fractions from Scutia buxifolia stem bark and leaves were evaluated. Best antimicrobial results occurred with the ethyl acetate (EA) and n-butanolic (NB) fractions from the leaves against Micrococcus sp. (minimal inhibitory concentration—MIC = 62.5 μg/ml), and NB fraction from stem bark and leaves against Klebsiella pneumoniae and Enterococcus faecalis (MIC = 62.5 μg/ml). The most active fractions were selected and fractioned into silica column to perform an in vitro antibiofilm assay, which evidenced subfractions EA2 and EA3 as the more active against Candida albicans (biofilm inhibitory concentration—BIC = 582 ± 0.01 μg/ml) and Staphylococcus aureus (BIC = 360 ± 0.007 μg/ml), respectively. The NB (selectivity index—SI = 25.78) and the EA (SI = 15.97) fractions from the stem bark, and the EA (SI = 14.13) fraction from the leaves exhibited a potential antiviral activity towards Herpes Simplex Virus type 1 whereas EA2 and EA3 subfractions from leaves (SI = 12.59 and 10.06, respectively), and NB2 subfraction from stem bark (SI = 12.34) maintained this good activity. Phenolic acids and flavonoids (gallic acid, chlorogenic acid, caffeic acid, rutin, isoquercitrin, quercitrin and quercetin) were identified by HPLC and may be partially responsible for the antimicrobial and antiherpes activities observed. The results obtained in this study showed that Scutia buxifolia has antibiofilm and anti-herpetic activities and that these properties are reported for the first time for this species.     

Gamma-Decanolactone Alters the Expression of GluN2B,A1 Receptors,and COX-2 and Reduces DNA Damage in the PTZ-Induced Seizure Model After Subchronic Treatment in Mice

Neurochemical Research - Gamma-decanolactone (GD) has been shown to reduce epileptic behavior in different models, inflammatory decreasing, oxidative stress, and genotoxic parameters. This study...     

Scedosporium Cell Wall: From Carbohydrate-Containing Structures to Host–Pathogen Interactions

Scedosporium species are filamentous fungi usually found in sewage and soil from human-impacted areas. They cause a wide range of diseases in humans, from superficial infections, such as mycetoma, to invasive and disseminated cases, especially associated in immunocompromised patients. Scedosporium species are also related to lung colonization in individuals presenting cystic fibrosis and are considered one of the most frequent fungal pathogens associated to this pathology. Scedosporium cell wall contains glycosylated molecules involved in important biological events related to virulence and pathogenicity and represents a significant source of antigens. Polysaccharides, peptidopolysaccharides, O-linked oligosaccharides and glycosphingolipids have been identified on the Scedosporium surface. Their primary structures were determined based on a combination of techniques including gas chromatography, ESI-MS, and ¹H and ¹³C nuclear magnetic resonance. Peptidorhamnnomannans are common cell wall components among Scedosporium species. Comparing different species, minor structural differences in the carbohydrate portions were detected which could be useful to understand variations in virulence observed among the species. N- and O-linked peptidorhamnomannans are major pathogen-associated molecular patterns and, along with α-glucans, play important roles in triggering host innate immunity. Glycosphingolipids, such as glucosylceramides, have highly conserved structures in Scedosporium species and are crucial for fungal growth and virulence. The present review presents current knowledge on structural and functional aspects of Scedosporium glycoconjugates that are relevant for understanding pathogenicity mechanisms and could contribute to the design of new agents capable of inhibiting growth and differentiation of Scedosporium species. Other cell components such as melanin and ectophosphatases will be also included.

     

Glycoprofiling Bifidobacterial Consumption of Galacto-Oligosaccharides by Mass Spectrometry Reveals Strain-Specific,Preferential Consumption of Glycans

Galacto-oligosaccharides (GOS) are versatile food ingredients that possess prebiotic properties. However, at present there is a lack of precise analytical methods to demonstrate specific GOS consumption by bifidobacteria. To better understand the role of GOS as prebiotics, purified GOS (pGOS) without disaccharides and monosaccharides was prepared and used in bacterial fermentation experiments. Growth curves showed that all bifidobacteria assayed utilized and grew on pGOS preparations. We used a novel mass spectrometry approach involving matrix-assisted laser desorption ionization-Fourier transform ion cyclotron resonance (MALDI-FTICR) to determine the composition of oligosaccharides in GOS syrup preparations. MALDI-FTICR analysis of spent fermentation media demonstrated that there was preferential consumption of selected pGOS species by different bifidobacteria. The approach described here demonstrates that MALDI-FTICR is a rapid-throughput tool for comprehensive profiling of oligosaccharides in GOS mixtures. In addition, the selective consumption of certain GOS species by different bifidobacteria suggests a means for targeting prebiotics to enrich select bifidobacterial species.Galacto-oligosaccharides (GOS) are nondigestible carbohydrates and versatile food ingredients that possess prebiotic properties (1). In addition, other health benefits have been reported to result from consumption of these oligosaccharides, such as stimulation of intestinal mobility and mineral absorption, elimination of ammonium, and colon cancer prevention, as well as protection against certain pathogenic bacterial infections (6, 11, 19).The physicochemical characteristics of GOS have enabled them to be incorporated in food as prebiotic ingredients. GOS have been of interest in acidic beverages and fermented milk formulations since they exhibit increased thermal stability in acidic environments compared to fructo-oligosaccharides (16, 21). Thus, in the past decade, the applications of GOS in human food products have included dairy products, sugar replacements, diet supplements, and infant formula (11).Commercial GOS preparations are produced by enzymatic treatment of lactose with β-galactosidases from different sources, such as fungi, yeast, or bacteria, which results in a mixture of oligomers with various chain lengths (1). Thus, the basic structure of GOS includes a lactose core at the reducing end, which is typically elongated with up to six galactose residues. Structural diversity in GOS preparations is dependent on the enzyme used in the transgalactosylation reaction and the experimental conditions used, such as pH and temperature (5).Considerable effort has been made to understand the effects of GOS in vivo, and most studies have described the impact of GOS on intestinal bacterial population shifts and production of short-chain fatty acids attributed to bacterial fermentation. While there have a been a variety of in vitro studies characterizing the growth of different gut microbes on GOS, the majority of these studies used commercially available preparations of GOS. These commercial preparations contain high concentrations of monosaccharides (i.e., galactose and glucose) and the disaccharide lactose, both of which remain in the product after the transgalactosylation reaction. However, monosaccharides are the preferred substrates for most microorganisms when they are available in a mixed-carbon source (2). Thus, to evaluate growth on GOS, removal of monosaccharides and lactose is helpful (15).An analytical method currently used to measure GOS in food and feed products is high-pH anion-exchange chromatography (HPAEC) coupled to analysis with a pulse amperometric detector (PAD) (4). Van Laere and coworkers have used this method to monitor GOS fermentation in Bifidobacterium adolescentis cultures (20). However, HPAEC-PAD analysis is time-consuming and thus a low-throughput method. More importantly, due to the detector, in HPAEC-PAD analysis there is a differential response to oligosaccharides with higher degrees of polymerization (DP). Thus, new analytical approaches are needed to specifically characterize the consumption of GOS and other prebiotics by probiotic bacteria.We have previously developed analytical methods employing high-mass-accuracy and high-resolution Fourier transform ion cyclotron resonance (FTICR) mass spectrometry (MS) to characterize bacterial consumption of human milk oligosaccharides and fructo-oligosaccharides (9, 10, 14, 17). The matrix-assisted laser desorption ionization (MALDI)-FTICR method is a sensitive and robust analytical method with high-performance capability, and it allows rapid and unambiguous assignment of oligosaccharide signals.The aims of the present study were to investigate the oligosaccharide composition of GOS syrup preparations using MALDI-FTICR MS, to test lactose-free purified GOS (pGOS) as a sole carbon source in bifidobacterial fermentation experiments, and to determine the pGOS consumption profile by MALDI-FTICR MS. Four major bifidobacterial phylotypes, B. adolescentis, Bifidobacterium breve, Bifidobacterium longum subsp. infantis, and Bifidobacterium longum subsp. longum, were used, and our results demonstrate that there is differential consumption of individual GOS species by various bifidobacteria, which provides a conceptual basis for targeted enrichment of specific bifidobacterial strains using specific GOS fractions.