Critical role for CXC chemokine ligand 16 (SR-PSOX) in Th1 response mediated by NKT cells

The transmembrane chemokine CXCL 16 (CXCL16), which is the same molecule as the scavenger receptor that binds phosphatidylserine and oxidized lipoprotein (SR-PSOX), has been shown to mediate chemotaxis and adhesion of CXC chemokine receptor 6-expressing cells such as NKT and activated Th1 cells. We generated SR-PSOX/CXCL16-deficient mice and examined the role of this chemokine in vivo. The mutant mice showed a reduced number of liver NKT cells, and decreased production of IFN-gamma and IL-4 by administration of alpha-galactosylceramide (alphaGalCer). Of note, the alphaGalCer-induced production of IFN-gamma was more severely impaired than the production of IL-4 in SR-PSOX-deficient mice. In this context, SR-PSOX-deficient mice showed impaired sensitivity to alphaGalCer-induced anti-tumor effect mediated by IFN-gamma from NKT cells. NKT cells from wild-type mice showed impaired production of IFN-gamma, but not IL-4, after their culture with alphaGalCer and APCs from mutant mice. Moreover, Propionibacterium acnes-induced in vivo Th1 responses were severely impaired in SR-PSOX-deficient as well as NKT KO mice. Taken together, SR-PSOX/CXCL16 plays an important role in not only the production of IFN-gamma by NKT cells, but also promotion of Th1-inclined immune responses mediated by NKT cells.     

Glucosylation of phenols and phenylalkyl alcohols using cultured plant cells

Cultured plant cells of Eucalyptus perriniana can convert phenol and phenylalkyl alcohols [C(6)H(5)(CH(2))(n)OH, n=0-3] into the corresponding beta-D-glucopyranosides in a good yield. The cells preferentially glucosylated phenylmethanol (n=1, 59% yield) rather than phenol (n=0, 49%), 2-phenylethanol (n=2, 38%), and 3-phenylpropan-1-ol (n=3, 20%). On the other hand, 2-, 3-, and 4-hydroxyphenylmethanols were also glucosylated to (hydroxymethyl)phenyl beta-D-glucopyranosides and (hydroxyphenyl)methyl beta-D-glucopyranosides by cultured E. perriniana cells.     

Isolation and characterization of microsatellite loci in <Emphasis Type="Italic">Melicope quadrilocularis</Emphasis> (Rutaceae), an endemic plant species of the Bonin Islands,Japan, and cross-species amplification in closely related taxa

Twelve microsatellite loci were isolated and characterized for Melicope quadrilocularis, an insular endemic tree species of the Bonin Islands. The observed number of alleles at each locus ranged from 1 to 18. The range of expected heterozygosity was 0.0000–0.9445. The inter-specific applicability of these loci was evaluated by analyzing two other endemic species and one endemic variety of Melicope that are also distributed on the Bonin Islands. All primer pairs for the 12 loci tested successfully amplified the loci in all taxa, except for primers for four loci in M. nishimurae.     

Multiple losses of photosynthesis in Nitzschia (Bacillariophyceae)

In order to obtain insights into the evolution of colorless (apochlorotic) diatoms, we investigated newly established apochlorotic strains of Nitzschia spp. using light and electron microscopy and molecular phylogenetic analyses. Fluorescence microscopic observations demonstrated that the apochlorotic diatoms lack chlorophylls. Transmission electron microscopy of two apochlorotic strains also demonstrated that their plastids lacked thylakoids; instead, having four‐membrane‐bound organelles without thylakoids, similar to nonphotosynthetic plastid remnants. From the apochlorotic strains, we also found plastid small subunit rRNA genes that were unusually long branched in phylogenetic analyses, as observed in other nonphotosynthetic plastids. Molecular phylogenetic analysis of the nucleus‐encoded large subunit rRNA genes showed eight distinct lineages for apochlorotic diatoms. The eight apochlorotic lineages were not monophyletic, suggesting that the loss of photosynthesis took place multiple times independently within Nitzschia. Several diatoms, including Nitzschia spp., are mixotrophic, which is an expected mode of nutrition that would help explain the evolutionary switch from a photosynthetic lifestyle to a heterotrophic lifestyle.     

High butyric acid amounts induce oxidative stress,alter calcium homeostasis,and cause neurite retraction in nerve growth factor-treated PC12 cells

Butyric acid (BA) is a common secondary metabolite by-product produced by oral pathogenic bacteria and is detected in high amounts in the gingival tissue of patients with periodontal disease. Previous works have demonstrated that BA can cause oxidative stress in various cell types; however, this was never explored using neuronal cells. Here, we exposed nerve growth factor (NGF)-treated PC12 cells to varying BA concentrations (0.5, 1.0, 5.0 mM). We measured total heme, H₂O₂, catalase, and calcium levels through biochemical assays and visualized the neurite outgrowth after BA treatment. Similarly, we determined the effects of other common periodontal short-chain fatty acids (SCFAs) on neurite outgrowth for comparison. We found that high (1.0 and 5.0 mM) BA concentrations induced oxidative stress and altered calcium homeostasis, whereas low (0.5 mM) BA concentration had no significant effect. Moreover, compared to other SCFAs, we established that only BA was able to induce neurite retraction.     

Diabetes-Related Ankyrin Repeat Protein (DARP/Ankrd23) Modifies Glucose Homeostasis by Modulating AMPK Activity in Skeletal Muscle

Skeletal muscle is the major site for glucose disposal, the impairment of which closely associates with the glucose intolerance in diabetic patients. Diabetes-related ankyrin repeat protein (DARP/Ankrd23) is a member of muscle ankyrin repeat proteins, whose expression is enhanced in the skeletal muscle under diabetic conditions; however, its role in energy metabolism remains poorly understood. Here we report a novel role of DARP in the regulation of glucose homeostasis through modulating AMP-activated protein kinase (AMPK) activity. DARP is highly preferentially expressed in skeletal muscle, and its expression was substantially upregulated during myotube differentiation of C2C12 myoblasts. Interestingly, DARP-/- mice demonstrated better glucose tolerance despite similar body weight, while their insulin sensitivity did not differ from that in wildtype mice. We found that phosphorylation of AMPK, which mediates insulin-independent glucose uptake, in skeletal muscle was significantly enhanced in DARP-/- mice compared to that in wildtype mice. Gene silencing of DARP in C2C12 myotubes enhanced AMPK phosphorylation, whereas overexpression of DARP in C2C12 myoblasts reduced it. Moreover, DARP-silencing increased glucose uptake and oxidation in myotubes, which was abrogated by the treatment with AICAR, an AMPK activator. Of note, improved glucose tolerance in DARP-/- mice was abolished when mice were treated with AICAR. Mechanistically, gene silencing of DARP enhanced protein expression of LKB1 that is a major upstream kinase for AMPK in myotubes in vitro and the skeletal muscle in vivo. Together with the altered expression under diabetic conditions, our data strongly suggest that DARP plays an important role in the regulation of glucose homeostasis under physiological and pathological conditions, and thus DARP is a new therapeutic target for the treatment of diabetes mellitus.     

Effects of Eliminating Pyruvate Node Pathways and of Coexpression of Heterogeneous Carboxylation Enzymes on Succinate Production by Enterobacter aerogenes

Lowering the pH in bacterium-based succinate fermentation is considered a feasible approach to reduce total production costs. Newly isolated Enterobacter aerogenes strain AJ110637, a rapid carbon source assimilator under weakly acidic (pH 5.0) conditions, was selected as a platform for succinate production. Our previous work showed that the ΔadhE/PCK strain, developed from AJ110637 with inactivated ethanol dehydrogenase and introduced Actinobacillus succinogenes phosphoenolpyruvate carboxykinase (PCK), generated succinate as a major product of anaerobic mixed-acid fermentation from glucose under weakly acidic conditions (pH <6.2). To further improve the production of succinate by the ΔadhE/PCK strain, metabolically engineered strains were designed based on the elimination of pathways that produced undesirable products and the introduction of two carboxylation pathways from phosphoenolpyruvate and pyruvate to oxaloacetate. The highest production of succinate was observed with strain ES04/PCK+PYC, which had inactivated ethanol, lactate, acetate, and 2,3-butanediol pathways and coexpressed PCK and Corynebacterium glutamicum pyruvate carboxylase (PYC). This strain produced succinate from glucose with over 70% yield (gram per gram) without any measurable formation of ethanol, lactate, or 2,3-butanediol under weakly acidic conditions. The impact of lowering the pH from 7.0 to 5.5 on succinate production in this strain was evaluated under pH-controlled batch culture conditions and showed that the lower pH decreased the succinate titer but increased its yield. These findings can be applied to identify additional engineering targets to increase succinate production.     

Disruption of Heat Shock Protein 90 (Hsp90)-Protein Kinase Cδ (PKCδ) Interaction by (?)-Maackiain Suppresses Histamine H1 Receptor Gene Transcription in HeLa Cells

The histamine H₁ receptor (H1R) gene is an allergic disease sensitive gene, and its expression level is strongly correlated with the severity of allergic symptoms. (−)-Maackiain was identified as a Kujin-derived anti-allergic compound that suppresses the up-regulation of the H1R gene. However, the underlying mechanism of H1R gene suppression remains unknown. Here, we sought to identify a target protein of (−)-maackiain and investigate its mechanism of action. A fluorescence quenching assay and immunoblot analysis identified heat shock protein 90 (Hsp90) as a target protein of (−)-maackiain. A pull-down assay revealed that (−)-maackiain disrupted the interaction of Hsp90 with PKCδ, resulting in the suppression of phorbol 12-myristate 13-acetate (PMA)-induced up-regulation of H1R gene expression in HeLa cells. Additional Hsp90 inhibitors, including 17-(allylamino)-17-demethoxygeldanamycin, celastrol, and novobiocin also suppressed PMA-induced H1R gene up-regulation. 17-(Allylamino)-17-demethoxygeldanamycin inhibited PKCδ translocation to the Golgi and phosphorylation of Tyr³¹¹ on PKCδ. These data suggest that (−)-maackiain is a novel Hsp90 pathway inhibitor. The underlying mechanism of the suppression of PMA-induced up-regulation of H1R gene expression by (−)-maackiain and Hsp90 inhibitors is the inhibition of PKCδ activation through the disruption of Hsp90-PKCδ interaction. Involvement of Hsp90 in H1R gene up-regulation suggests that suppression of the Hsp90 pathway could be a novel therapeutic strategy for allergic rhinitis.