Human Parainfluenza Virus Type 2 V Protein Inhibits TRAF6-Mediated Ubiquitination of IRF7 To Prevent TLR7- and TLR9-Dependent Interferon Induction

Paramyxovirus V proteins block Toll-like receptor 7 (TLR7)- and TLR9-dependent signaling leading to alpha interferon production. Our recent study has provided evidence that interaction of the V proteins with IRF7 is important for the blockade. However, the detailed mechanisms still remain unclear. Here we reexamined the interaction of the human parainfluenza virus type 2 (HPIV2) V protein with signaling molecules involved in TLR7/9-dependent signaling. Immunoprecipitation experiments in HEK293T cells transfected with V protein and one of the signaling molecules revealed that the V protein interacted with not only IRF7 but also TRAF6, IKKα, and MyD88. Whereas overexpression of TRAF6 markedly enhanced the level of V protein associating with IRF7, IKKα, and MyD88 in HEK293T cells, the level of V protein associating with TRAF6 was little affected by overexpression of IRF7, IKKα, and MyD88. Moreover, knockdown or knockout of endogenous TRAF6 in HEK293T or mouse embryonic fibroblast cells resulted in dissociation of the V protein from IRF7, IKKα, and MyD88. These results demonstrate that binding of the V protein to IRF7, IKKα, and MyD88 is largely indirect and mediated by endogenous TRAF6. It was found that the V protein inhibited TRAF6-mediated lysine 63 (K63)-linked polyubiquitination of IRF7, which is prerequisite for IRF7 activation. Disruption of the tryptophan-rich motif of the V protein significantly affected its TRAF6-binding efficiency, which correlated well with the magnitude of inhibition of K63-linked polyubiquitination and the resultant activation of IRF7. Taken together, these results suggest that the HPIV2 V protein prevents TLR7/9-dependent interferon induction by inhibiting TRAF6-mediated K63-linked polyubiquitination of IRF7.     

Blend miscibility of cellulose propionate with poly(N-vinyl pyrrolidone-co-methyl methacrylate)

The blend miscibility of cellulose propionate (CP) with poly(N-vinyl pyrrolidone-co-methyl methacrylate) (P(VP-co-MMA)) was investigated. The degree of substitution (DS) of CP used ranged from 1.6 to >2.9, and samples for the vinyl polymer component were prepared in a full range of VP:MMA compositions. Through DSC analysis and solid-state ¹³C NMR and FT-IR measurements, we revealed that CPs of DS < 2.7 were miscible with P(VP-co-MMA)s of VP ≥ ∼10 mol% on a scale within a few nanometers, in virtue of hydrogen-bonding interactions between CP-hydroxyls and VP-carbonyls. When the DS of CP exceeded 2.7, the miscibility was restricted to the polymer pairs using P(VP-co-MMA)s of VP = ca. 10–40 mol%; the scale of mixing in the blends concerned was somewhat larger (ca. 5–20 nm), however. The appearance of such a “miscibility window” was interpretable as an effect of intramolecular repulsion in the copolymer component. Results of DMA and birefringence measurements indicated that the miscible blending of CP with the vinyl polymer invited synergistic improvements in thermomechanical and optical properties of the respective constituent polymers. Additionally, it was found that the VP:MMA composition range corresponding to the miscibility window was expanded by modification of the CP component into cellulose acetate propionate.     

Novel contribution of cell surface and intracellular M1‐muscarinic acetylcholine receptors to synaptic plasticity in hippocampus

Muscarinic acetylcholine receptors (mAChRs) are well known to transmit extracellular cholinergic signals into the cytoplasm from their position on the cell surface. However, we show here that M1‐mAChRs are also highly expressed on intracellular membranes in neurons of the telencephalon and activate signaling cascades distinct from those of cell surface receptors, contributing uniquely to synaptic plasticity. Radioligand‐binding experiments with cell‐permeable and ‐impermeable ligands and immunohistochemical observations revealed intracellular and surface distributions of M1‐mAChRs in the hippocampus and cortex of rats, mice, and humans, in contrast to the selective occurrence on the cell surface in other tissues. All intracellular muscarinic‐binding sites were abolished in M1‐mAChR‐gene‐knockout mice. Activation of cell surface M1‐mAChRs in rat hippocampal neurons evoked phosphatidylinositol hydrolysis and network oscillations at theta rhythm, and transiently enhanced long‐term potentiation. On the other hand, activation of intracellular M1‐mAChRs phosphorylated extracellular‐regulated kinase 1/2 and gradually enhanced long‐term potentiation. Our data thus demonstrate that M1‐mAChRs function at both surface and intracellular sites in telencephalon neurons including the hippocampus, suggesting a new mode of cholinergic transmission in the central nervous system.     

Omega-3 polyunsaturated fatty acid has an anti-oxidant effect via the Nrf-2/HO-1 pathway in 3T3-L1 adipocytes

Oxidative stress is produced in adipose tissue of obese subjects and has been associated with obesity-related disorders. Recent studies have shown that omega-3 polyunsaturated fatty acid (ω3-PUFA) has beneficial effects in preventing atherosclerotic diseases and insulin resistance in adipose tissue. However, the role of ω3-PUFA on adipocytes has not been elucidated. In this study, 3T3-L1 adipocytes were treated with ω3-PUFA and its metabolites, eicosapentaenoic acid (EPA), docosahexaenoic acid (DHA), or 4-hydroxy hexenal (4-HHE). ω3-PUFA and its metabolites dose-dependently increased mRNA and protein levels of the anti-oxidative enzyme, heme oxygenase-1 (HO-1); whereas no changes in the well-known anti-oxidant molecules, superoxide dismutase, catalase, and glutathione peroxidase, were observed. Knockdown of nuclear factor erythroid 2-related factor 2 (Nrf-2) significantly reduced EPA, DHA or 4-HHE-induced HO-1 mRNA and protein expression. Also, pretreatment with ω3-PUFA prevented H₂O₂-induced cytotoxicity in a HO-1 dependent manner. In conclusion, treatment with EPA and DHA induced HO-1 through the activation of Nrf-2 and prevented oxidative stress in 3T3-L1 adipocytes. This anti-oxidant defense may be of high therapeutic value for clinical conditions associated with systemic oxidative stress.     

Ester Synthesis in Various Organic Solvents by Three Kinds of Lipase Preparations Derived from Pseudomonas fragi 22.39 B

DNA-synthesis stimulatory activity was found in some tryptic fragments of human /¡-casein by using BALB/c3T3 cells, and two of them were identified as the /¡-casein fragments of Arg[l] to Lys[18] (/¡-CN(f 1 -18)) and of Gln[105] to Lys[l 17] (/¡-CN(fl05 -117)).     

Preparation of Optically Pure cis-2,4-Dimethyl-l-cyclohexanones,the Key Intermediates in Cycloheximide Synthesis,Using Microbial Resolution

Asymmetric hydrolysis of acetate (10) of (±)-t-2,t-4-dimethyl-r-l-cyclohexanol with Bacillus subtilis var. niger gave (?)-(lS,2S,4S)-2,4-dimethyl-l-cyclohexanol (6a) and (+)-(1R,2R,4R)-acetate (10b) with high optical purities. Optically pure (?) and (+)-alcohols (6a and 6b) were prepared via corresponding 3,5-dinitrobenzoates. Oxidation of alcohols (6a and 6b) with chromic acid gave optically pure (?)-(2S,4S) and (+)-(2R,4R)-2,4-dimethyl-l-cyclohexanones (2a and 2b), respectively.     

NADPH Production from NADP+ by a Formate-utilizing Methanogenic Bacterium

Resting cells of the methanogen strain HU, a formate-utilizing methanogenic bacterium, was able to utilize formate or hydrogen as electron donor for the production of NADPH from NADP⁺ under suitable conditions. In the presence of 0.2% Triton X-100 and 0.3 m potassium phosphate, pH 9.0 at 30°C, the resting cells could convert ca. 60% of the exogenous NADP⁺ into NADPH yielding ca. 6 g NADPH/liter. Phosphate ions greatly enhanced the NADPH production.     

Synthesis of Streptovitacin-C2 Isomers

(2R*,4S*,6S*,αS*)- and (2R,4R,6R,αS)-Streptovitacin-C₂ (STV-C₂) (1a and 1b) were synthesized by an aldol condensation of (2R*,4S*)- or (2R,4R)-2,4-dimethyl-2-trimethylsiloxy-1-cyclohexanone (15a or 15b) with 4-(2-oxoethyl)-2,6-piperidinedione (16), which was followed by desilylation of the products. The stereochemistry of the synthesized STV-C₂ isomers (1a and 1b) was elucidated by NMR. STV-C₂ isomers (1a and 1b) did not show strong antimicrobial activity against Saccharomyces cerevisiae and Pyricularia oryzae.     

The Preparation and Geometrical Isomerism of Substituted α-Methylthio-Cinnamic Acids

α-Methylthio-cinnamic acid and its substituted analogues (III) were synthesized from their respective β-aryl-α-thiopyruvic acids (II). In connection with the study on the tautomeric ene-thiol structure of β-aryl-α-thiopyruvic acids (II), 4-arylidenerl,3-oxathiolan-5-one (IV) were prepared from compounds II.     

Solid State 13C-NMR Analysis of Cell Wall Components of Fusarium oxysporum

An abdominal fat accumulation complicated by high blood triglycerides is regarded as a risk factor of metabolic syndrome. Feeding powdered nacre, mother of pearl, from Pinctada maxima, resulted in reduced body weight, visceral fat amount, and blood triglyceride level without influencing the food intake, body length, or amount of muscular tissue, suggesting that nacre powder specifically could decrease visceral fat.