Genetic Engineering of Zymobacter palmae for Production of Ethanol from Xylose

Its metabolic characteristics suggest that Zymobacter palmae gen. nov., sp. nov. could serve as a useful new ethanol-fermenting bacterium, but its biotechnological exploitation will require certain genetic modifications. We therefore engineered Z. palmae so as to broaden the range of its fermentable sugar substrates to include the pentose sugar xylose. The Escherichia coli genes encoding the xylose catabolic enzymes xylose isomerase, xylulokinase, transaldolase, and transketolase were introduced into Z. palmae, where their expression was driven by the Zymomonas mobilis glyceraldehyde-3-phosphate dehydrogenase promoter. When cultured with 40 g/liter xylose, the recombinant Z. palmae strain was able to ferment 16.4 g/liter xylose within 5 days, producing 91% of the theoretical yield of ethanol with no accumulation of organic acids as metabolic by-products. Notably, xylose acclimation enhanced both the expression of xylose catabolic enzymes and the rate of xylose uptake into recombinant Z. palmae, which enabled the acclimated organism to completely and simultaneously ferment a mixture of 40 g/liter glucose and 40 g/liter xylose within 8 h, producing 95% of the theoretical yield of ethanol. Thus, efficient fermentation of a mixture of glucose and xylose to ethanol can be accomplished by using Z. palmae expressing E. coli xylose catabolic enzymes.     

Cutting edge: negative regulation of immune synapse formation by anchoring lipid raft to cytoskeleton through Cbp-EBP50-ERM assembly.

Ag recognition by T lymphocytes induces immune synapse formation and recruitment of signaling molecules into a lipid raft. Cbp/PAG is a Csk-associated membrane adapter protein exclusively localized in a lipid raft. We identified NHERF/EBP50 as a Cbp-binding molecule, which connects the membrane raft and cytoskeleton by binding to both Cbp through its PDZ domain and ezrin-radixin-moesin through the C terminus. Overexpression of Cbp reduced the mobility of the raft on the cell surface of unstimulated T cells and prevented synapse formation and subsequent T cell activation, whereas a mutant incapable of EBP50 binding restored both synapse formation and activation. These results suggest that anchoring of lipid raft to the cytoskeleton through Cbp-EBP50-ezrin-radixin-moesin assembly regulates membrane dynamism for synapse formation and T cell activation.     

Insulin modulates thyrotropin-induced follicle reconstruction and iodine metabolism in hog thyroid cells cultured in a chemically defined medium

Effects of insulin on cultured hog thyroid cells were examined in a chemically defined medium supplemented with thyrotropin. Thyrotropin-induced follicle reconstruction and recovery of functional activities were stimulated by the addition of insulin to the culture medium. The iodide uptake and organification at the maximum stimulation exceeded 3 times those of the control values. The minimum effective dose of insulin was 60 microU/ml. The insulin stimulation occurred after a latency period of 24 h. Since no growth promoting effect of insulin was observed during the cultivation period in the present study, insulin appears to act in concert with thyrotropin in regenerating the characteristic properties of the differentiated cells during cultivation.     

A new brain-penetrant glucosylceramide synthase inhibitor as potential Therapeutics for Gaucher disease

Gaucher disease (GD), the most common lysosomal storage disorders, is caused by GBA gene mutations resulting in glycosphingolipids accumulations in various tissues, such as the brain. While suppressing glycosphingolipid accumulation is the central strategy for treating peripheral symptoms of GD, there is no effective treatment for the central nervous system symptoms. As glycosphingolipid biosynthesis starts from ceramide glycosylation by glucosylceramide synthase (GCS), inhibiting GCS in the brain is a promising strategy for neurological GD. Herein, we discovered T-036, a potent and brain-penetrant GCS inhibitor with a unique chemical structure and binding property. T-036 does not harbor an aliphatic amine moiety and has a noncompetitive inhibition mode to the substrates, unlike other known inhibitors. T-036 exhibited sufficient exposure and a significant reduction of glucosylsphingolipids in the plasma and brain of the GD mouse model. Therefore, T-036 could be a promising lead molecule for treating central nervous system symptoms of GD.

     

Activated Macrophage Survival Is Coordinated by TAK1 Binding Proteins

Macrophages play diverse roles in tissue homeostasis and immunity, and canonically activated macrophages are critically associated with acute inflammatory responses. It is known that activated macrophages undergo cell death after transient activation in some settings, and the viability of macrophages impacts on inflammatory status. Here we report that TGFβ- activated kinase (TAK1) activators, TAK1-binding protein 1 (TAB1) and TAK1-binding protein 2 (TAB2), are critical molecules in the regulation of activated macrophage survival. While deletion of Tak1 induced cell death in bone marrow derived macrophages even without activation, Tab1 or Tab2 deletion alone did not profoundly affect survival of naïve macrophages. However, in lipopolysaccharide (LPS)-activated macrophages, even single deletion of Tab1 or Tab2 resulted in macrophage death with both necrotic and apoptotic features. We show that TAB1 and TAB2 were redundantly involved in LPS-induced TAK1 activation in macrophages. These results demonstrate that TAK1 activity is the key to activated macrophage survival. Finally, in an in vivo setting, Tab1 deficiency impaired increase of peritoneal macrophages upon LPS challenge, suggesting that TAK1 complex regulation of macrophages may participate in in vivo macrophage homeostasis. Our results demonstrate that TAB1 and TAB2 are required for activated macrophages, making TAB1 and TAB2 effective targets to control inflammation by modulating macrophage survival.     

Redescription of Scymnodon ichiharai Yano and Tanaka 1984 (Squaliformes: Somniosidae) from the western North Pacific,with comments on the definition of somniosid genera

Ichthyological Research - The somniosid species Scymnodon ichiharai Yano and Tanaka 1984 is redescribed based on the type specimens and additional material from Taiwan and Japan. The range of this...     

Mediation of ecdysone synthesis in Manduca sexta by a hemolymph enzyme

The prothoracic glands of Manduca sexta synthesize dehydroecdysone, which is rapidly converted to ecdysone through the mediation of a hemolymph enzyme, a 3 β-forming-3-ketosteroid reductase. The hemolymph protein fraction (HPF) containing this enzyme was obtained from diapausing and non-diapausing pupae, isolated abdomens, surgically manipulated pupae, etc., and in all cases had the capacity to affect the conversion of dehydroecdysone to ecdysone. The enzyme is heat labile, is inactivated by trypsin, and has a molecular weight of between 20,000 and 30,000. The data indicate that the conversion of dehydroecdysone to ecdysone exhibits linear kinetics and may be dependent on both the enzyme concentration and the concentration of NADPH at the beginning of the reaction but may be limited by the absolute amount of reducing equivalents after 10 min, under the experimental conditions utilized. The capacity of the enzyme to reduce dehydroecdysone was titered in the hemolymph during the last larval instar and during prepupal and pupal life with maximum capacity exhibited at the beginning of the instar, on day 8 of larval life and at day 1 of pupal life. Even at its lowest point at day 5, 1 ml of hemolymph was able to convert 77 pmol (～35 ng) dehydroecdysone to ecdysone in 1 min. These results require a new interpretation of the control of molting in the Lepidoptera.     

Impaired Air Conditioning within the Nasal Cavity in Flat-Faced Homo

We are flat-faced hominins with an external nose that protrudes from the face. This feature was derived in the genus Homo, along with facial flattening and reorientation to form a high nasal cavity. The nasal passage conditions the inhaled air in terms of temperature and humidity to match the conditions required in the lung, and its anatomical variation is believed to be evolutionarily sensitive to the ambient atmospheric conditions of a given habitat. In this study, we used computational fluid dynamics (CFD) with three-dimensional topology models of the nasal passage under the same simulation conditions, to investigate air-conditioning performance in humans, chimpanzees, and macaques. The CFD simulation showed a horizontal straight flow of inhaled air in chimpanzees and macaques, contrasting with the upward and curved flow in humans. The inhaled air is conditioned poorly in humans compared with nonhuman primates. Virtual modifications to the human external nose topology, in which the nasal vestibule and valve are modified to resemble those of chimpanzees, change the airflow to be horizontal, but have little influence on the air-conditioning performance in humans. These findings suggest that morphological variation of the nasal passage topology was only weakly sensitive to the ambient atmosphere conditions; rather, the high nasal cavity in humans was formed simply by evolutionary facial reorganization in the divergence of Homo from the other hominin lineages, impairing the air-conditioning performance. Even though the inhaled air is not adjusted well within the nasal cavity in humans, it can be fully conditioned subsequently in the pharyngeal cavity, which is lengthened in the flat-faced Homo. Thus, the air-conditioning faculty in the nasal passages was probably impaired in early Homo members, although they have survived successfully under the fluctuating climate of the Plio-Pleistocene, and then they moved “Out of Africa” to explore the more severe climates of Eurasia.     

Compartmentation and localization of bioactive metabolites in different organs of Allium roylei

To investigate the involvement of Allium roylei metabolites in the plant’s defenses, a comprehensive analysis of the content of cysteine sulfoxides, flavonols, polyphenols, ascorbic acid, and saponins was carried out in the various organs of this species. Metabolomics high performance liquid chromatography (HPLC), spectral-based analysis, and histochemcial studies have given important insight to the validity of saponins as a key component involved in plant protection. The root-basal stem, bulb, and leaf extracts exhibited 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging activity with inhibition concentration (IC₅₀) ranging from 0.649 to 0.757?mg/mL. The antimicrobial properties of the saponin and flavonoid crude extracts were evaluated. The saponin extracts demonstrated significant antifungal activity depending on the applied concentration, and the growth inhibition rate of the tested fungal pathogens ranged from 1.07 to 47.76%. No appreciable antibacterial activity was recorded in the same sample.