Liver Fatty Acid-binding Protein Binds Monoacylglycerol in Vitro and in Mouse Liver Cytosol

Liver fatty acid-binding protein (LFABP; FABP1) is expressed both in liver and intestinal mucosa. Mice null for LFABP were recently shown to have altered metabolism of not only fatty acids but also monoacylglycerol, the two major products of dietary triacylglycerol hydrolysis (Lagakos, W. S., Gajda, A. M., Agellon, L., Binas, B., Choi, V., Mandap, B., Russnak, T., Zhou, Y. X., and Storch, J. (2011) Am. J. Physiol. Gastrointest. Liver Physiol. 300, G803–G814). Nevertheless, the binding and transport of monoacylglycerol (MG) by LFABP are uncertain, with conflicting reports in the literature as to whether this single chain amphiphile is in fact bound by LFABP. In the present studies, gel filtration chromatography of liver cytosol from LFABP^−/− mice shows the absence of the low molecular weight peak of radiolabeled monoolein present in the fractions that contain LFABP in cytosol from wild type mice, indicating that LFABP binds sn-2 MG in vivo. Furthermore, solution-state NMR spectroscopy demonstrates two molecules of sn-2 monoolein bound in the LFABP binding pocket in positions similar to those found for oleate binding. Equilibrium binding affinities are ∼2-fold lower for MG compared with fatty acid. Finally, kinetic studies examining the transfer of a fluorescent MG analog show that the rate of transfer of MG is 7-fold faster from LFABP to phospholipid membranes than from membranes to membranes and occurs by an aqueous diffusion mechanism. These results provide strong support for monoacylglycerol as a physiological ligand for LFABP and further suggest that LFABP functions in the efficient intracellular transport of MG.     

Mammalian Target of Rapamycin (mTor) Mediates Tau Protein Dyshomeostasis: IMPLICATION FOR ALZHEIMER DISEASE*

Previous evidence from post-mortem Alzheimer disease (AD) brains and drug (especially rapamycin)-oriented in vitro and in vivo models implicated an aberrant accumulation of the mammalian target of rapamycin (mTor) in tangle-bearing neurons in AD brains and its role in the formation of abnormally hyperphosphorylated tau. Compelling evidence indicated that the sequential molecular events such as the synthesis and phosphorylation of tau can be regulated through p70 S6 kinase, the well characterized immediate downstream target of mTor. In the present study, we further identified that the active form of mTor per se accumulates in tangle-bearing neurons, particularly those at early stages in AD brains. By using mass spectrometry and Western blotting, we identified three phosphoepitopes of tau directly phosphorylated by mTor. We have developed a variety of stable cell lines with genetic modification of mTor activity using SH-SY5Y neuroblastoma cells as background. In these cellular systems, we not only confirmed the tau phosphorylation sites found in vitro but also found that mTor mediates the synthesis and aggregation of tau, resulting in compromised microtubule stability. Changes of mTor activity cause fluctuation of the level of a battery of tau kinases such as protein kinase A, v-Akt murine thymoma viral oncogene homolog-1, glycogen synthase kinase 3β, cyclin-dependent kinase 5, and tau protein phosphatase 2A. These results implicate mTor in promoting an imbalance of tau homeostasis, a condition required for neurons to maintain physiological function.     

Differing Epidemiological Dynamics of Influenza B Virus Lineages in Guangzhou,Southern China, 2009-2010

The epidemiological and evolutionary dynamics of the two cocirculating lineages of influenza B virus, Victoria and Yamagata, are poorly understood, especially in tropical or subtropical areas of Southeast Asia. We performed a phylogenetic analysis of the hemagglutinin (HA) and neuraminidase (NA) sequences of influenza B viruses isolated in Guangzhou, a southern Chinese city, during 2009 to 2010 and compared the demographic and clinical features of infected patients. We identified multiple viral introductions of Victoria strains from both Chinese and international sources, which formed two phylogenetically and antigenically distinct clades (Victoria 1 and 2), some of which persisted between seasons. We identified one dominant Yamagata introduction from outside China during 2009. Our phylogenetic analysis reveals the occurrence of reassortment events among the Victoria and Yamagata lineages and also within the Victoria lineage. We found no significant difference in clinical severity by influenza B lineage, with the exceptions that (i) the Yamagata lineage infected older people than either Victoria lineage and (ii) fewer upper respiratory tract infections were caused by the Victoria 2 than the Victoria 1 clade. Overall, our study reveals the complex epidemiological dynamics of different influenza B lineages within a single geographic locality and has implications for vaccination policy in southern China.     

Characterization and expression of gamma-interferon-inducible lysosomal thiol reductase (GILT) gene in rainbow trout (Oncorhynchus mykiss) with implications for GILT in innate immune response

The interferon-γ-inducible lysosomal thiol reductase (GILT) has been demonstrated to play an important role in the processing and presentation of MHC class II-restricted antigen (Ag) by catalyzing disulfide bond reduction. In this study, a rainbow trout cDNA (designated as rGILT) was cloned and identified from Oncorhynchus mykiss. The open reading frame of rGILT consists of 759 bases encoding a protein of 253 amino acids with an estimated molecular mass of 28.23 kDa and a theoretical isoelectric point of 4.94. The rGILT exhibited a characteristic GILT signature sequence CQHGX₂ECX₂NX₄C and CXXC motif. Phylogenetic analysis suggested that rGILT had been derived from a common ancestor with other GILT proteins. RT-PCR results showed that rGILT and rIFN-γ (rainbow trout IFN-γ) mRNA was expressed in a tissue-specific manner and obviously up-regulated in splenocytes and the cells from head kidney after induction with LPS. Recombinant rGILT fused with His6 tag was efficiently expressed in Escherichia coli BL21 (DE3) and purified by Ni-NTA affinity chromatography. Further study revealed that rGILT was capable of catalyzing the reduction of the interchain disulfide bonds from intact IgG. This study shows that rGILT may be involved in the immune response to bacteria challenge and maintain first line of innate immune defense at basal level in O. mykiss. It also provides the basis for investigating on the role of GILT using O. mykiss as an animal model for related studies.     

Cadmium removal and 2,4-dichlorophenol degradation by immobilized Phanerochaete chrysosporium loaded with nitrogen-doped TiO2 nanoparticles

Phanerochaete chrysosporium has been identified as an effective bioremediation agent for its biosorption and degradation ability. However, the applications of P. chrysosporium are limited owing to its long degradation time and low resistance to pollutants. In this research, nitrogen-doped TiO₂ nanoparticles were loaded on P. chrysosporium to improve the remediation capacity for pollutants. The removal efficiencies were maintained at a high level: 84.2 % for Cd(II) and 78.9 % for 2,4-dichlorophenol (2,4-DCP) in the wide pH range of 4.0 to 7.0 in 60 h. The removal capacity of immobilized P. chrysosporium loaded with nitrogen-doped TiO₂ nanoparticles (PTNs) was strongly affected by the initial Cd(II) and 2,4-DCP concentrations. The hyphae of PTNs became tight, and a large amount of crystals adhered to them after the reaction. Fourier transform infrared spectroscopy showed that carboxyl, amino, and hydroxyl groups on the surface of PTNs were responsible for the biosorption. In the degradation process, 2,4-DCP was broken down into o-chlorotoluene and 4-hexene-1-ol. These results showed that PTNs is promising for simultaneous removal of Cd(II) and 2,4-DCP from wastewater.     

A novel locus essential for spreading of Cytophaga hutchinsonii colonies on agar

Cytophaga hutchinsonii is an aerobic cellulolytic gliding bacterium. The mechanism of its cell motility over surfaces without flagella and type IV pili is not known. In this study, mariner-based transposon mutagenesis was used to identify a new locus CHU_1797 essential for colony spreading on both hard and soft agar surfaces through gliding. CHU_1797 encodes a putative outer membrane protein of 348 amino acids with unknown function, and proteins which have high sequence similarity to CHU_1797 were widespread in the members of the phylum Bacteroidetes. The disruption of CHU_1797 suppressed spreading toward glucose on an agar surface, but had no significant effect on cellulose degradation for cells already in contact with cellulose. SEM observation showed that the mutant cells also regularly arranged on the surface of cellulose fiber similar with that of the wild type strain. These results indicated that the colony spreading ability on agar surfaces was not required for cellulose degradation by C. hutchinsonii. This was the first study focused on the relationship between cell motility and cellulose degradation of C. hutchinsonii.     

Additional data for a new Theileria sp. from China based on the sequences of ribosomal RNA internal transcribed spacers

Theileria sinensis was recently isolated and named as an independent Theileria species that infects cattle in China. To date, this parasite has been described based on its morphology, transmission and molecular studies, indicating that it should be classified as a distinct species. To test the validity of this taxon, the two internal transcribed spacers (ITS1 and ITS2) and the 5.8S rRNA gene were cloned and sequenced from three T. sinensis isolates. The complete ITS sequences were compared with those of other Theileria sp. available in GenBank. Phylogenetic analyses based on sequence data for the complete ITS sequences indicate that T. sinensis lies in a distinct clade that is separate from that of T. buffeli/orientalis and T. annulata. Sequence comparisons indicate that different T. sinensis isolates possess unique sizes of ITS1 and ITS2 as well as species-specific nucleotide sequences. This analysis provides new molecular data to support the classification of T. sinensis as a distinct species from other known Theileria spp. based on ITS sequences.     

Actinopolyspora dayingensis sp. nov., a novel halophilic actinomycete isolated from a hypersaline lake

A novel halophilic, filamentous actinomycete, designated TRM 4064^T, was isolated from a hypersaline habitat in Sichuan Province, China. Phylogenetic analysis based on an almost-complete 16S rRNA gene sequence of strain TRM 4064^T showed that it was most closely related to Actinopolyspora mortivallis (99.1 % sequence similarity). The sequence similarities between strain TRM 4064^T and other Actinopolyspora species with validly-published names were <97.0 %. However, it had relatively low mean values for DNA–DNA relatedness with the A. mortivallis DSM 44261^T (23.2 %). Optimal growth occurred at 37 °C, pH 7.0 and in the presence of 13 % (w/v) NaCl. The whole-cell sugar pattern consists of xylose, glucose, ribose and arabinose. The predominant menaquinones are MK-10(H₄) (38.2 %), MK-9(H₄) (25.1 %), MK-9(H₂) (28.6 %) and MK-8(H₄) (7.3 %). The major fatty acids are anteiso-C_17:0 (36.9 %) and iso-C_17:0 (19.3 %). The diagnostic phospholipids detected were diphosphatidylglycerol (DPG), phosphatidylglycerol (PG), phosphatidylcholine (PC), phosphatidylinositol (PI) and two unknown phospholipids. The G+C content of the genomic DNA of the type strain is 66.3 mol%. Strain TRM 4064^T therefore represents a novel species of the genus Actinopolyspora, for which the name Actinopolyspora dayingensis sp. nov. is proposed. The type strain is TRM 4064^T (= KCTC 19979^T = CCTCC AA 2010010^T).