Systematic analysis of SNARE localization in the filamentous fungus Aspergillus oryzae

In spite of their great importance for both applied and basic biology, studies on vesicular trafficking in filamentous fungi have been so far very limited. Here, we identified 21 genes, which might be a total set, encoding putative SNARE proteins that are key factors for vesicular trafficking, taking advantage of available whole genome sequence in the filamentous fungus Aspergillus oryzae. The subsequent systematic analysis to determine the localization of putative SNAREs using EGFP-fused chimeras revealed that most putative SNAREs show similar subcellular distribution to their counterparts in the budding yeast. However, there existed some characteristic features of SNAREs in A. oryzae, such as SNARE localization at/near the septum and the presence of apparently non-redundant plasma membrane Qa-SNAREs. Overall, this analysis allowed us to provide an overview of vesicular trafficking and organelle distribution in A. oryzae.     

Antiviral Effect of Theaflavins on Tobacco Mosaic Virus

Theaflavins, the oxidized products formed from tea leaf catechins during black tea fermentation, showed an antiviral activity on TMV. From the survey of the interactions of theaflavins with RNA and its related substances, it was presumed that theaflavins disturbed the replication cycle of TMV through binding to TMV-RNA.     

Isolation and characterization of some novel genes of the apolipoprotein A-I family in Japanese eel, <Emphasis Type="Italic">Anguilla japonica</Emphasis>

Apolipoproteins such as apolipoprotein (apo) A-I, apoA-IV, and apoE are lipid binding proteins synthesized mainly in the liver and the intestine and play an important role in the transfer of exogenous or endogenous lipids through the circulatory system. To investigate the mechanism of lipid transport in fish, we have isolated some novel genes of the apoA-I family, apoIA-I (apoA-I isoform) 1–11, from Japanese eel by PCR amplification. Some of the isolated genes of apoIA-I corresponded to 28kDa-1 cDNAs which had already been deposited into the database and encoded an apolipoprotein with molecular weight of 28 kDa in the LDL, whereas others seemed to be novel genes. The structural organization of all apoIA-Is consisted of four exons separated by three introns. ApoIA-I10 had a total length of 3232 bp, whereas other genes except for apoIA-I9 ranged from 1280 to 1441 bp. The sequences of apoIA-Is at the exon-intron junctions were mostly consistent with the consensus sequence (GT/AG) at exon-intron boundaries, whereas the sequences of 3′ splice acceptor in intron 1 of apoIA-I1-7 were (AC) but not (AG). The deduced amino acid sequences of all apoIA-Is contained a putative signal peptide and a propeptide of 17 and 5 amino acid residues, respectively. The mature proteins of apoIA-I1-3, 7, and 8 consisted of 237 amino acids, whereas those of apoIA-I4-6 consisted of 239 amino acids. The mature apoIA-I10 sequence showed 65% identity to amino acid sequence of apoIA-I11 which was associated with an apolipoprotein with molecular weight of 23 kDa in the VLDL. All these mature apoIA-I sequences satisfied the common structural features depicted for the exchangeable apolipoproteins such as apoA-I, apoA-IV, and apoE but apoIA-I11 lacked internal repeats 7, 8, and 9 when compared with other members of apoA-I family. Phylogenetic analysis showed that these novel apoIA-Is isolated from Japanese eel were much closer to apoA-I than apoA-IV and apoE, suggesting new members of the apoA-I family.     

A genetically explicit model of speciation by sensory drive within a continuous population in aquatic environments

Background  

The sensory drive hypothesis predicts that divergent sensory adaptation in different habitats may lead to premating isolation upon secondary contact of populations. Speciation by sensory drive has traditionally been treated as a special case of speciation as a byproduct of adaptation to divergent environments in geographically isolated populations. However, if habitats are heterogeneous, local adaptation in the sensory systems may cause the emergence of reproductively isolated species from a single unstructured population. In polychromatic fishes, visual sensitivity might become adapted to local ambient light regimes and the sensitivity might influence female preferences for male nuptial color. In this paper, we investigate the possibility of speciation by sensory drive as a byproduct of divergent visual adaptation within a single initially unstructured population. We use models based on explicit genetic mechanisms for color vision and nuptial coloration.     

Barotropic and thermotropic bilayer phase behavior of positional isomers of unsaturated mixed-chain phosphatidylcholines

The bilayer phase transitions of six kinds of mixed-chain phosphatidylcholines (PCs) with an unsaturated acyl chain in the sn-1 or sn-2 position, 1-oleoyl-2-stearoyl- (OSPC), 1-stearoyl-2-oleoyl- (SOPC), 1-oleoyl-2-palmitoyl- (OPPC), 1-palmitoyl-2-oleoyl- (POPC), 1-oleoyl-2-myristoyl- (OMPC) and 1-myristoyl-2-oleoyl-sn-glycero-3-phosphocholine (MOPC), were observed by means of differential scanning calorimetry (DSC) and high-pressure light transmittance measurements. Bilayer membranes of SOPC, POPC and MOPC with an unsaturated acyl chain in the sn-2 position exhibited only one phase transition, which was identified as the main transition between the lamellar gel (L_β) and liquid crystalline (L_α) phases. On the other hand, the bilayer membranes of OSPC, OPPC and OMPC with an unsaturated acyl chain in the sn-1 position exhibited not only the main transition but also a transition from the lamellar crystal (L_c) to the L_β (or L_α) phase. The stability of their gel phases was markedly affected by pressure and chain length of the saturated acyl chain in the sn-2 position. Considering the effective chain lengths of unsaturated mixed-chain PCs, the difference in the effective chain length between the sn-1 and sn-2 acyl chains was proven to be closely related to the temperature difference of the main transition. That is, a mismatch of the effective chain length promotes a temperature difference of the main transition between the positional isomers. Anomalously small volume changes of the L_c/L_α transition for the OPPC and OMPC bilayers were found despite their large enthalpy changes. This behavior is attributable to the existence of a cis double bond and to significant inequivalence between the sn-1 and sn-2 acyl chains, which brings about a small volume change for chain melting due to loose chain packing, corresponding to a large partial molar volume, even in the L_c phase. Further, the bilayer behavior of unsaturated mixed-chain PCs containing an unsaturated acyl chain in the sn-1 or sn-2 position was well explained by the chemical-potential diagram of a lipid in each phase.     

A G protein-coupled receptor responsive to bile acids

So far some nuclear receptors for bile acids have been identified. However, no cell surface receptor for bile acids has yet been reported. We found that a novel G protein-coupled receptor, TGR5, is responsive to bile acids as a cell-surface receptor. Bile acids specifically induced receptor internalization, the activation of extracellular signal-regulated kinase mitogen-activated protein kinase, the increase of guanosine 5'-O-3-thio-triphosphate binding in membrane fractions, and intracellular cAMP production in Chinese hamster ovary cells expressing TGR5. Our quantitative analyses for TGR5 mRNA showed that it was abundantly expressed in monocytes/macrophages in human and rabbit. Treatment with bile acids was found to suppress the functions of rabbit alveolar macrophages including phagocytosis and lipopolysaccharide-stimulated cytokine productions. We prepared a monocytic cell line expressing TGR5 by transfecting a TGR5 cDNA into THP-1 cells that did not express TGR5 originally. Treatment with bile acids suppressed the cytokine productions in the THP-1 cells expressing TGR5, whereas it did not influence those in the original THP-1 cells, suggesting that TGR5 is implicated in the suppression of macrophage functions by bile acids.     

Distinct DNA methylation activity of Dnmt3a and Dnmt3b towards naked and nucleosomal DNA

In mammals, the resetting of DNA methylation patterns in early embryos and germ cells is crucial for development. De novo type DNA methyltransferases Dnmt3a and Dnmt3b are responsible for creating DNA methylation patterns during embryogenesis and in germ cells. Although their in vitro DNA methylation properties are similar, Dnmt3a and Dnmt3b methylate different genomic DNA regions in vivo. In the present study, we have examined the DNA methylation activity of Dnmt3a and Dnmt3b towards nucleosomes reconstituted from recombinant histones and DNAs, and compared it to that of the corresponding naked DNAs. Dnmt3a showed higher DNA methylation activity than Dnmt3b towards naked DNA and the naked part of nucleosomal DNA. On the other hand, Dnmt3a scarcely methylated the DNA within the nucleosome core region, while Dnmt3b significantly did, although the activity was low. We propose that the preferential DNA methylation activity of Dnmt3a towards the naked part of nucleosomal DNA and the significant methylation activity of Dnmt3b towards the nucleosome core region contribute to their distinct methylation of genomic DNA in vivo.     

Pannexin1 channels act downstream of P2X7 receptors in ATP-induced murine T-cell death

Death of murine T cells induced by extracellular ATP is mainly triggered by activation of purinergic P2X₇ receptors (P2X₇Rs). However, a link between P2X₇Rs and pannexin1 (Panx1) channels, which are non-selective, has been recently demonstrated in other cell types. In this work, we characterized the expression and cellular distribution of pannexin family members (Panxs 1, 2 and 3) in isolated T cells. Panx1 was the main pannexin family member clearly detected in both helper (CD4⁺) and cytotoxic (CD8⁺) T cells, whereas low levels of Panx2 were found in both T-cell subsets. Using pharmacological and genetic approaches, Panx1 channels were found to mediate most ATP-induced ethidium uptake since this was drastically reduced by Panx1 channel blockers (¹⁰Panx1, Probenecid and low carbenoxolone concentration) and absent in T cells derived from Panx1^−/− mice. Moreover, electrophysiological measurements in wild-type CD4⁺ cells treated with ATP unitary current events and pharmacological sensitivity compatible with Panx1 channels were found. In addition, ATP release from T cells treated with 4Br-{"type":"entrez-nucleotide","attrs":{"text":"A23187","term_id":"833253","term_text":"A23187"}}A23187, a calcium ionophore, was completely blocked with inhibitors of both connexin hemichannels and Panx1 channels. Panx1 channel blockers drastically reduced the ATP-induced T-cell mortality, indicating that Panx1 channels mediate the ATP-induced T-cell death. However, mortality was not reduced in T cells of Panx1^−/− mice, in which levels of P2X₇Rs and ATP-induced intracellular free Ca²⁺ responses were enhanced suggesting that P2X₇Rs take over Panx1 channels lose-function in mediating the onset of cell death induced by extracellular ATP.