Glial cells contribute more to iron and aluminum accumulation but are more resistant to oxidative stress than neuronal cells

Iron (Fe) and aluminum (Al) have been implicated in the pathogenesis of Alzheimer's disease (AD). In this study, we examined neuronal and glial cells to clarify which contributes most to metal accumulation after internalization through the transferrin-independent iron uptake (Tf-IU) systems in primary neuronal and glial predominant (NP and GP) cells from rat cerebral cortex, which affect the accumulation of transition metals in a variety of cultured cells. Al more significantly upregulated the Tf-IU activity in GP cells than in NP cells. GP cells were more resistant to Fe and Al exposure than NP cells. However, a chemiluminescence analysis specific for reactive oxygen species (ROS) showed that ROS levels in Fe- or Al-loaded NP cells were twice as high as in Fe- or Al-loaded GP cells. Northern blot analysis and gel retardation assay showed that the Al and Fe exposure taken up by the cells suppress Tf receptor mRNA expression to a greater extent in GP than NP cells, indicating that Al and Fe more markedly accumulate in glial than in neuronal cells. These results suggest that glial cells rather than neuronal cells contribute to the metal accumulation and are more resistant to oxidative stress caused by metals than neuronal cells. The present study may help to explain the pathogenesis of neurodegeneration in AD disorders caused by metal-generated oxidative stress.     

Co-localization of receptor and transducer proteins in the glycosphingolipid-enriched, low density, detergent-insoluble membrane fraction of sea urchin sperm

The low density, detergent-insoluble membrane fraction (LD-DIM), where gangliosides are likely to be highly enriched, was prepared from sperm of two sea urchin species, Hemicentrotus pulcherrimus and Strongylocentrotus purpuratus. Immunoblotting showed the presence in the LD-DIM of two receptors for egg ligands, a glycosylphosphatidylinositol (GPI)-anchored protein, and four proteins which may be involved in signal transduction. Co-immunoprecipitation revealed that at least three proteins, the speract receptor, the 63[emsp4 ]kDa GPI-anchored protein and the subunit of a heterotrimeric Gs protein, are localized in the LD-DIM. This suggests that the LD-DIM fraction may be a membrane microdomain for speract–speract receptor interaction, as well as the subsequent signal transduction pathway involved in induction of sperm respiration, motility and possibly the acrosome reaction.     

Flavonol glycosides from Asplenium foreziense and its five related taxa and A. incisum

The flavonoids of Asplenium foreziense, A. fontanum subsp. fontanum and subsp. pseudofontanum, A. obovatum subsp. obovatum var. obovatum and var. protobillotii, A. obovatum subsp. lanceolatum, and A. incisum were isolated and identified for chemotaxonomic survey. A major constituent of all taxa was kaempferol 3-O-gentiobioside. As minor compounds, kaempferol 3,7-O-glycoside and/or kaempferol 3-O-glycoside were found in A. fontanum, A. obovatum and A. foreziense, and kaempferol 3-O-gentiobioside-4'-O-glucoside, kaempferol 3-O-glucoside and quercetin 3-O-diglucoside in A. incisum. It was suggested that A. foreziense, A. fontanum including subsp. pseudofontanum and A. obovatum including subsp. lanceolatum are not only morphologically but also chemotaxonomically related. The East Asian A. incisum was chemically and geographically different from these taxa.     

Molecular cloning and expression of the mouse N-acetylneuraminic acid 9-phosphate synthase which does not have deaminoneuraminic acid (KDN) 9-phosphate synthase activity

A cDNA of the mouse homologue of Escherichia coli N-acetylneuraminic acid (Neu5Ac) synthase (neuB gene product) was cloned by the PCR-based method. The mouse homologue consists of 359 amino acids, and the cDNA sequence displays 33% identity to that of the E. coli Neu5Ac synthase. The recombinant mouse homologue which is transiently expressed in HeLa cells does not exhibit the Neu5Ac synthase activity, which catalyzes condensation of phosphoenolpyruvate (PEP) and N-acetylmannosamine (ManNAc) to synthesize Neu5Ac, but the Neu5Ac 9-phosphate (Neu5Ac-9-P) synthase activity, which catalyzes condensation of PEP and ManNAc 6-phosphate (ManNAc-6-P) to synthesize Neu5Ac-9-P. Thus, the mouse homologue of E. coli Neu5Ac synthase is the Neu5Ac-9-P synthase. The Neu5Ac-9-P synthase is a cytosolic enzyme and ubiquitously distributed in mouse various tissues. Notably, the Neu5Ac-9-P synthase can not catalyze the synthesis of deaminoneuraminic acid (KDN) or KDN-9-P from PEP and Man or ManNAc-6-P, thus suggesting that the enzyme is not involved in the synthesis of KDN. This is consistent with the previous observation that only a very low activity to synthesize KDN is found in mouse B16 cells [Angata, T., et al. (1999) Biochem. Biophys. Res. Commun. 261, 326-331].     

Mouse gene trap approach: identification of novel genes and characterization of their biological functions.

The mouse gene trap strategy is an insertional mutagenesis involving an exogenous DNA, termed the trap vector, as a mutagen that produces a mutation in the mouse genome and a sequence tag to facilitate the isolation of the mutated genes. The trap vector consists of a reporter gene whose expression mimics that of the endogenous genes mutated and a selection marker that sorts cells bearing the inserted vector. Gene trap is a powerful method for identifying genes important in biological phenomena. Moreover, the method produces mutant organisms whose phenotypes provide invaluable information about the biological functions of the genes responsible for these phenotypes. Indeed, a number of genes essential for mouse embryogenesis have been identified by the gene trap method. Here, we describe the principle, results, and perspectives for applications of gene trap approach to the study of cell differentiation and lineage commitment.     

Synthesis of neoglycoconjugates containing deaminated neuraminic acid (KDN) using rat liver alpha2,6-sialyltransferase

2-Keto-3-deoxy-D- glycero -D- galacto -nononic acid (KDN) was introduced into asialotransferrin and N -acetyllactosamine (LacNAc) from CMP-KDN by using rat liver Galbeta1-->4GlcNAc alpha2, 6- sialyltransferase to form KDN-transferrin and KDN-LacNAc. These structures contain terminal KDNalpha2-->6Gal-residues, a glycotope that has not yet been described in natural glycoconjugates. KDN was transferred to all four Gal residues in asialotransferrin by this enzyme. The incorporation efficiency of KDN from CMP-KDN into asialotransferrin was about half that of Neu5Ac from CMP-Neu5Ac, based on the V max/ K m values for these donor substrates, 0.0527 min-1and 0.119 min-1, respectively. The KDNalpha2-->6Gal linkage was resistant to exosialidase treatment, in contrast to the sensitivity of the Neu5Acalpha2-->6Gal linkage. Interestingly, Sambucus sieboldiana agglutinin (SSA) was shown to prefer KDN-transferrin to the corresponding Neu5Ac-transferrin, as estimated by slot-blot analysis. The use of an alpha2,6-sialyltransferase to synthesize neoglycoproteins containing KDN has not been previously reported. Their facile synthesis using CMP-KDN and sialyltransferases with different specificities offers new possibilities to study the function of neo-KDN- glycoconjugates, and to explore their use in glycotechnology.      

Flexible Ion‐Conducting Composite Membranes for Lithium Batteries

The use of metallic lithium anodes enables higher energy density and higher specific capacity Li‐based batteries. However, it is essential to suppress lithium dendrite growth during electrodeposition. Li‐ion‐conducting ceramics (LICC) can mechanically suppress dendritic growth but are too fragile and also have low Li‐ion conductivity. Here, a simple, versatile, and scalable procedure for fabricating flexible Li‐ion‐conducting composite membranes composed of a single layer of LICC particles firmly embedded in a polymer matrix with their top and bottom surfaces exposed to allow for ionic transport is described. The membranes are thin (<100 μm) and possess high Li‐ion conductance at thicknesses where LICC disks are mechanically unstable. It is demonstrated that these membranes suppress Li dendrite growth even when the shear modulus of the matrix is lower than that of lithium. It is anticipated that these membranes enable the use of metallic lithium anodes in conventional and solid‐state Li‐ion batteries as well as in future Li? S and Li? O₂ batteries.     

Quantitative proteomics of Arabidopsis shoot microsomal proteins reveals a cross‐talk between excess zinc and iron deficiency

Iron (Fe) deficiency significantly effects plant growth and development. Plant symptoms under excess zinc (Zn) resemble symptoms of Fe‐deficient plants. To understand cross‐talk between excess Zn and Fe deficiency, we investigated physiological parameters of Arabidopsis plants and applied iTRAQ‐OFFGEL quantitative proteomic approach to examine protein expression changes in microsomal fraction from Arabidopsis shoots under those physiological conditions. Arabidopsis plants manifested shoot inhibition and chlorosis symptoms when grown on Fe‐deficient media compared to basal MGRL solid medium. iTRAQ‐OFFGEL approach identified 909 differentially expressed proteins common to all three biological replicates; the majority were transporters or proteins involved in photosynthesis, and ribosomal proteins. Interestingly, protein expression changes between excess Zn and Fe deficiency showed similar pattern. Further, the changes due to excess Zn were dramatically restored by the addition of Fe. To obtain biological insight into Zn and Fe cross‐talk, we focused on transporters, where STP4 and STP13 sugar transporters were predominantly expressed and responsive to Fe‐deficient conditions. Plants grown on Fe‐deficient conditions showed significantly increased level of sugars. These results suggest that Fe deficiency might lead to the disruption of sugar synthesis and utilization.