Purification and characterization of aldoxime dehydratase of the head blight fungus, Fusarium graminearum

Fungal aldoxime dehydratase (Oxd) of Fusarium graminearum MAFF305135 was purified and characterized for the first time from its overexpressing Escherichia coli transformant. The enzyme showed about 20% identity with known Oxds, and had similar enzymatic properties with nitrilase-linked Oxd from the Bacillus strain. It belongs to a group of phenylacetaldoxime dehydratases (EC 4.99.1.7), based on its substrate specificity and kinetic analysis.     

Microtubule-cyclodextrin conjugate: functionalization of motile filament with molecular inclusion ability

A microtubule-beta-cyclodextrin conjugate was prepared on a kinesin-adsorbed glass surface by chemical and biochemical means. Fluorescence microscope observation and a motility assay indicated that the conjugate simultaneously expressed an inherent motor function and an inclusion property.     

Cell-to-cell movement of the CAPRICE protein in Arabidopsis root epidermal cell differentiation

CAPRICE (CPC), a small, R3-type Myb-like protein, is a positive regulator of root hair development in Arabidopsis. Cell-to-cell movement of CPC is important for the differentiation of epidermal cells into trichoblasts (root hair cells). CPC is transported from atrichoblasts (hairless cells), where it is expressed, to trichoblasts, and generally accumulates in their nuclei. Using truncated versions of CPC fused to GFP, we identified a signal domain that is necessary and sufficient for CPC cell-to-cell movement. This domain includes the N-terminal region and a part of the Myb domain. Amino acid substitution experiments indicated that W76 and M78 in the Myb domain are critical for targeted transport, and that W76 is crucial for the nuclear accumulation of CPC:GFP. To evaluate the tissue-specificity of CPC movement, CPC:GFP was expressed in the stele using the SHR promoter and in trichoblasts using the EGL3 promoter. CPC:GFP was able to move from trichoblasts to atrichoblasts but could not exit from the stele, suggesting the involvement of tissue-specific regulatory factors in the intercellular movement of CPC. Analyses with a secretion inhibitor, Brefeldin A, and with an rhd3 mutant defective in the secretion process in root epidermis suggested that intercellular CPC movement is mediated through plasmodesmata. Furthermore, the fusion of CPC to tandem-GFPs defined the capability of CPC to increase the size exclusion limit of plasmodesmata.     

General synthesis and biological evaluation of alpha-1-C-substituted derivatives of fagomine (2-deoxynojirimycin-alpha-C-glycosides)

A general synthesis of alpha-1-C-substituted derivatives of fagomine (2-deoxynojirimycin-alpha-C-glycosides) by ring-opening reactions of an aziridine with various heteroatomic nucleophiles, including thiol, amine, alcohol, carboxylate and phosphate, is described. The nine-step reaction sequence proceeded in an overall yield of 14-28% from tri-O-benzyl-D-glucal. Biological evaluation of alpha-1-C-substituted derivatives of fagomine, of the 2-deoxy analog of alpha-homonojirimycin 19 and its 1,N-anhydro derivative 22 as glycosidase inhibitors is reported. The glycosyl phosphate mimetic 15k was found to display no inhibitory activity towards glycogen phosphorylase b and phosphoglucomutase.     

Functional polymorphisms of HSPA5: possible association with bipolar disorder

Altered endoplasmic reticulum stress (ER) response signaling is suggested in bipolar disorder. Previously, we preliminarily reported the genetic association of HSPA5 (GRP78/BiP) with bipolar disorder. Here, we extended our analysis by increasing the number of Japanese case-control samples and NIMH Genetics Initiative bipolar trio samples (NIMH trios), and also analyzed schizophrenia samples. In Japanese, nominally significant association of one haplotype was observed in extended samples of bipolar disorder but not in schizophrenia. In NIMH trios, no association was found in total samples. However, an exploratory analysis suggested that the other haplotype was significantly over-transmitted to probands only from the paternal side. The associated haplotype in Japanese or NIMH pedigrees shared three common polymorphisms in the promotor, which was found to alter promotor activity. These findings suggested promotor polymorphisms of HSPA5 may affect the interindividual variability of ER stress response and may confer a genetic risk factor for bipolar disorder.     

Beyond vasodilation: the antioxidant effect of adrenomedullin in Dahl salt-sensitive rat aorta

We have investigated the antioxidant effect of adrenomedullin (AM) on endothelial function in the Dahl salt-sensitive (DS) rat hypertension model. Dahl salt-resistant (DR) and DS rats were fed an 8% NaCl diet. In addition, the DS rats were subcutaneously infused with either saline or recombinant human AM for 4 weeks. Although systolic blood pressures measured weekly in AM- and saline-infused rats did not significantly differ, aortic O2*- levels were significantly (P<0.01) higher in the latter. Likewise, both endothelial nitric oxide synthase (eNOS) mRNA and protein were significantly higher in saline-infused DS rats. Infusion of AM reduced both O2*- and eNOS expression to levels comparable to those seen in DR rats. AM infusion also upregulated the gene expression of guanosine-5'-triphosphate cyclohydrolase I and downregulated the expression of p22(phox), suggesting that AM increased the NOS coupling and bioavailability of NO. AM possesses significant antioxidant properties that improve endothelial function.     

Role of microtubules and centrosomes in the eccentric relocation of the germinal vesicle upon meiosis reinitiation in sea-cucumber oocytes

In the oocytes of many animals, the germinal vesicle (GV) relocates from the center to the periphery of the oocyte upon meiosis reinitiation, which is a prerequisite to the formation of meiotic spindles beneath the cell surface in order for meiosis to succeed. In the present study, we have investigated nuclear positioning using sea-cucumber oocytes. Upon meiosis reinitiation, the GV relocates to the cell periphery beneath a surface protuberance. After GV breakdown, polar bodies were extruded from the top of the protuberance, which we therefore called the animal pole process. The GV relocation was inhibited by nocodazole but not by cytochalasin. Immunofluorescent staining and electron microscopy of microtubular arrays revealed that: (i) in immature oocytes, two centrosomes were situated beneath the animal pole process far apart from the GV, anchoring to the cortex via astral microtubules; (ii) upon meiosis reinitiation, microtubular bundles were newly formed between the centrosomes and the GV; and (iii) the microtubular bundles became short as GV migration proceeded. These observations suggest that microtubules and centrosomes participate in GV relocation. A very large mass of annulate lamellae, having a 20-microm diameter, was found in the vegetal pole of the oocytes.