A Novel Role for hGas7b in Microtubular Maintenance: POSSIBLE IMPLICATION IN TAU-ASSOCIATED PATHOLOGY IN ALZHEIMER DISEASE*

Here, we report a novel role for hGas7b (human growth arrest specific protein 7b) in the regulation of microtubules. Using a bioinformatic approach, we studied the actin-binding protein hGas7b with a structural similarity to the WW domain of a peptidyl prolyl cis/trans isomerase, Pin1, that facilitates microtubule assembly. Thus, we have demonstrated that hGas7b binds Tau at the WW motif and that the hGas7b/Tau protein complex interacts with the microtubules, promoting tubulin polymerization. Tau, in turn, contributes to protein stability of hGas7b. Furthermore, we observed decreased levels of hGas7b in the brains from patients with Alzheimer disease. These results suggest an important role for hGas7b in microtubular maintenance and possible implication in Alzheimer disease.     

The distribution and intracellular location of Fas and Fas Ligand following gastric carcinogenesis: Fas Ligand expressing gastric carcinoma cells can inhibit local immune response

Previous reports indicated that Fas Ligand (FasL) in gastric carcinoma might support tumour cells to evade host immune attack. However, the mechanism induced by the Fas/FasL system has not yet been described on the basis of comparison of normal and malignant tissues in terms of the features of regional location of Fas and FasL. By using immunostaining methods, we studied the distribution and regional location of Fas and FasL in gastric epithelial cells (GECs), gastric carcinoma cells (GCCs), normal gastric stroma-infiltrating lymphoid cells (NGILs) and tumour-infiltrating lymphoid cells (TILs) in 59 tissue specimens of human gastric carcinoma. The expression of Fas within the entire GECs was higher than that in all GCCs (P < 0.0001); however, the expression of Fas in NGILs was lower than that in TILs (P < 0.0001). The expression of FasL showed no significant difference between GECs and GCCs, or between NGILs and TILs. When we analyzed the Fas/FasL expression on cytomembrane (CM) in GECs and GCCs, Fas-in-CM was detected in 79.4% and 33.33% (P < 0.05), compared with 3.03% and 56.67%, respectively, for FasL-in-CM (P < 0.001). Our results suggest that there is indeed a possible mechanism to assist cancer cells to evade host immune attack, and this mechanism depends on the dynamic state of Fas/FasL expression, that is, Fas showed a tendency to be expressed within the cells, whereas FasL showed a tendency to be expressed on the cell membrane following carcinogenesis.     

Assessing temporal dynamics of predation and effectiveness of caterpillar visual defense using sawfly larval color and resting posture as a model

Caterpillars (Lepidoptera and Symphyta larvae) employ diverse visual defensive tactics, and effectiveness of such tactics may be highly dynamic across time due to seasonal changes in the predator assemblages and their preferences. However, this has rarely been studied especially in tropical regions. Here we assessed temporal changes in the defensive value of caterpillar color and shape, using six types of plasticine dummy caterpillars: three colors (green, black, and white) × two shapes (curled and straight). These dummy caterpillars were deployed five times over different seasons in tropical forests of Xishuangbanna (China) and, as a comparison, twice in a temperate forest of Hirosaki (Japan). The colors and shapes of dummy caterpillars simulate visual traits of black sawfly larvae which take the curled resting posture in tropical rainforests of Xishuangbanna, apparently masquerading excrements commonly found on plants, while in Hirosaki there is no black-curled sawfly larvae and few excrements on plants. We found no significant effects of caterpillar colors or shapes on predation in Hirosaki. In contrast, black and curled caterpillars received significantly lower predation by birds in Xishuangbanna constantly across time. However, we were unable to provide evidence that the black-curled sawfly larvae are masquerading as excrements. Shapes of the dummy caterpillars also affected the predation by ants and parasitoid wasps at certain times. This is the first report on ecological function of the curled posture of sawfly larvae, and we demonstrated the importance to assess the temporal dynamics of predation and effectiveness of defensive tactics in tropical forests.     

Separation and analysis of free ceramides containing 2-hydroxy fatty acids in Sphingobacterium species

Abstract The occurrence of free ceramides was shown in the chloroform-methanol extractable lipids of 16 strains of Sphingobacterium including three species: S. versatilis, S. multivorum and S. mizutae . The predominant long-chain base was identified as a branched-chain, saturated dihydroxy base with a carbon chain consisting of 17 carbon atoms, while the most abundant fatty acid was 2-hydroxy-13-methyltetradecanoic acid. The major molecular species of the intact ceramides were identified as LCB- d - iso -17 : 0-2-OH iso -15 : 0FA, LCB- d - iso -17 : 0- iso -15 : 0FA and LCB- d -n16 : 0- iso -15 : 0FA.     

Identification of a key amino acid residue of Streptomyces phospholipase D for thermostability by in vivo DNA shuffling

To isolate thermostability-related amino acid residues of Streptomyces phospholipase D (PLD), we constructed a chimeral genes library between two highly homologous plds, which exhibited different thermostabilities, by an in vivo DNA shuffling method using Escherichia coli that has a mutation of a single-stranded DNA-binding protein gene. To confirm the location of the recombination site, we carried out the restriction mapping of 68 chimeral pld genes. The recombination sites were widely dispersed over the entire pld sequence. Moreover, we examined six chimeral PLDs by comparing their thermostabilities with those of parental PLDs. To identify a thermostability-related amino acid residue, we investigated the thermostability of chimera C that was the most thermolabile among the six chimeras. We identified the thermostability-related factor Gly-188, which is located in the alpha-7 helix of PLD from Streptomyces septatus TH-2 (TH-2PLD). TH-2PLD mutants, in which Gly-188 was substituted with Phe, Val or Trp, exhibited higher thermostabilities than that of the parental PLD. Gly-188 substituted with the Phe mutant, which was the most stable among the mutants, showed an enzyme activity almost the same as that of TH-2PLD as determine by kinetic analysis.     

Changes in Crassulacean Acid Metabolism of Kalanchoe blossfeldiana by Different Nitrogen Sources

Kalanchoe blossfeldiana Poelln. cv. Hikan (a Crassulacean acidmetabolism (CAM) plant) was grown in pots containing soil for6 months and then cultured in nutrient solution containing 10mM nitrate or ammonium as a sole nitrogen source for 2 or 3months, under a long-day (16 h) condition. Plant growth was better in the nitrate medium. Leaves of thenitrate-grown plants showed greater diurnal fluctuations intitratable acidity and malate content than those of the ammonium-grownplants. The diurnal patterns in CO₂ exchange of nitrate-grownplants were basically similar for both groups, but the amountof net CO₂ uptake at night was twice as large in the nitrate-grownplants. The leaves of the nitrate-grown plants had 1.3 to 2.5times higher activities of phosphoenolpyruvate carboxylase (PEPC),phosphofructokinase (PFK) and NAD glycelaldehyde-3-phosphatedehydrogenase (G3PDH). These results indicate that K. blossfeldianagrown in nitrate medium showed more CAM activity than thosein ammonium medium. (Received August 13, 1987; Accepted February 22, 1988)     

A novel vaccine strategy to induce mycobacterial antigen-specific Th1 responses by utilizing the C-terminal domain of heat shock protein 70

Heat shock protein 70 (HSP70) is a member of a highly conserved superfamily of intracellular chaperones called stress proteins that can activate innate and adaptive immune responses. We evaluated the effect of a fusion DNA vaccine that encoded mycobacterial HSP70 and MPT51, a major secreted protein of Mycobacterium tuberculosis. Spleen cells from mice immunized with fusion DNA of full-length HSP70 and MPT51 produced a higher amount of interferon-γ (IFN-γ) in response to the CD4+, but not the CD8+ T-cell epitope peptide on MPT51 than those from mice immunized with MPT51 DNA. Furthermore, because HSP70 comprises the N-terminal ATPase domain and the C-terminal peptide-binding domain, we attempted to identify the domain responsible for its enhancing effect. The fusion DNA vaccine that encoded the C-terminal domain of HSP70 and MPT51 induced a higher MPT51-specific IFN-γ production by CD4+ T cells than the vaccine that encoded MPT51 alone, whereas that with the N-terminal domain did not. Similar results were obtained by immunization with the fusion proteins. These results suggest that the DNA vaccine that encodes a chimeric antigen molecule fused with mycobacterial HSP70, especially with its C-terminal domain, can induce a stronger antigen-specific T-helper cell type 1 response than antigen DNA alone.     

Molecular Dynamics Calculations of Wild Type vs. Mutant Protein C: Relationship Between Binding Affinity to Endothelial Cell Protein C Receptor and Hereditary Disease

Abstract

Molecular dynamics simulations of the protein C γ-carboxyglutamic acid (Gla) domain and endothelial cell protein C receptor (EPCR) complex were performed to determine the effect of a hereditary disease, which results in a mutation (Gla 25 → Lys) in the protein C Gla domain. Our results suggest that the Gla 25 → Lys mutation causes a significant reduction in the binding force between protein C Gla domain and EPCR due to destabilization of the helix structure of EPCR and displacement of a Ca²⁺ ion.