Fission yeast Skp1 is required for spindle morphology and nuclear membrane segregation at anaphase

Skp1 is a core component of the Skp1-Cullin-1-F-box ubiquitin ligase. Here, we show a novel role for fission yeast Skp1 in mitotic progression. Temperature-sensitive skp1-A7 mutants enter mitosis, but fail to execute anaphase. Time-lapse imaging shows that spindles in this mutant form intranuclear arch-like structures, which eventually collapse abruptly. The two spindle poles are also seen to move backward to the cell centre rather than towards the cell ends. These abnormal phenotypes appear to stem from defects in nuclear membrane segregation. Our results show that Skp1 is required for coordinated structural alterations of mitotic spindles and nuclear membranes.     

Reconstruction of microtubules; entry into interphase

Microtubules display dramatic morphological alterations from mitotic spindles to fibrous interphase structures upon exit from mitosis. In this issue of Developmental Cell, Zimmerman et al. shed a novel light on the molecular mechanism of microtubule structure reorganization during cytokinesis.     

Inhibitory effects of ellagi- and gallotannins on rat intestinal alpha-glucosidase complexes

The clove ellagitannins and their related polygalloyl-glucoses inhibited maltase activity of rat intestinal alpha-glucosidases. The structure-activity relationship study of those galloylglucoses, varying the extent of galloylation on the glucose core, with the ellagitannins, indicated that an increasing number of galloyl units in the molecule lead to an increase in the inhibitory activity. Penta-O-galloyl-beta-D-glucose, with five galloyl groups showed the highest inhibitory activity. On the other hand, hexahydroxydiphenoyl units contained in the ellagitannins had little effect on the activity. After separation of maltase-glucoamylase and sucrase-isomaltase complexes from the crude mixture of the rat alpha-glucosidases, the inhibitory activities of the galloylglucose derivatives against each complex were examined. The inhibitory influence on the maltase-glucoamylase complex was more potent than on the sucrase-isomaltase complex.     

Inhibitory effects of licoisoflavones A and B and sophoraisoflavone A of Sophra mooracroftiana Beth ex Baker on copper-ion-induced protein oxidative modification of mice brain homogenate, in vitro

We present the results of an in vitro investigation of the inhibitory effects of licoisoflavones A and B and sophoraisoflavone A isolated from Sophra mooracroftiana BETH ex BAKER on copper-induced protein oxidative modification of mice brain homogenate in vitro. Although inhibitory effect of sophoraisoflavone A was stronger than those of licoisoflavones A and B, genistein as a related isoflavone, and mannitol as a hydroxy radical scavenger, inhibitory effects of licoisoflavones A and B were weaker than those of genistein and mannitol. These results demonstrated that the difference of inhibitory effects are dependent on the relation between chemical structures of these isoflavones, such as hydroxy group or benzopyran, and oxidative stress.     

Comparison of antioxidative and chelating effects of daidzein and daidzin on protein oxidative modification by copper in vitro

Daidzein and its glycoside daidzin are isoflavones. Their antioxidative effects were compared in vitro. Although both compounds inhibited protein oxidative modification by copper, the inhibitory effect of daidzein was stronger than that of daidzin. Because daidzein showed a greater affinity for Cu²⁺, the antioxidant effect of these isoflavones may be dependent on their respective copper-chelating abilities.     

Input-output budget of nitrogen and the effect of experimentally changed deposition in the forest ecosystems in central Japan

To evaluate the current nitrogen (N) status in Japanese forests, field measurements of rainfall, throughfall, litter layer percolation, and soil solution percolation were conducted in a red pine stand (Kannondai) and a deciduous stand (Yasato) located in central Japan. N input via throughfall was 31 and 14 kg ha(-1) year(-1) and output below rooting zone was 9.6 and 5.5 kg ha1 year(-1) in Kannondai and in Yasato, respectively. Two thirds of input N were retained in plant-soil systems. Manipulation of N input was carried out. Ionic constituents were removed from throughfall with ion exchange resin at removal sites and ammonium nitrate containing twice the N of the throughfall was applied at N addition sites periodically. SO4(2-) output below 20-cm soil layer changed depending on the input, while NO3- output was regulated mainly by the internal cycle and effect of manipulation was undetected. These Japanese stands were generally considered to have a larger capacity to assimilate N than NITREX sites in Europe. However, N output fluxes had large spatial variability and some sites in Kannondai showed high N leaching below rooting zone almost balanced with the input via throughfall.     

Air exposure promotes fibroblast growth with increased expression of mitogen-activated protein kinase cascade

Subepithelial tissue cell types in vivo are separated from air by the surface-covering epithelial layer of various organs, e.g., the skin, cornea, and respiratory and upper alimentary tracts. The epithelial defect caused by inflammatory, traumatic or surgical injury would be expected to expose the subepithelial tissue-localized fibroblasts to influx air. However, it is unclear what effects air stimulation elicits in fibroblast growth, which is critical for wound healing. To address this question, we examined the proliferation of 3T3 fibroblasts with bromodeoxyuridine (BrdU) uptake, using fibroblast-embedded collagen gel culture with or without air exposure. The BrdU intake of air-exposed fibroblasts was about 6 times that of air-nonexposed cells. To further characterize this fibroblast growth, we examined the expression of mitogen-activated protein kinase (MAPK) cascade, which plays a key role in the growth-signaling pathway of various cell types. Immunohistochemistry and Western blotting showed that air exposure increased MAPK cascade expression of the cells more strongly than air nonexposure. The data indicate that air exposure promotes MAPK cascade-associated fibroblast growth, suggesting in turn that in wound repair air stimulation itself may be involved in the basic mechanisms of subepithelial fibroblast proliferation and that it may be related to the pathogenesis of excessive fibroplasia through fibroblast overgrowth.     

Synthesis of a molt-inhibiting hormone of the American crayfish, Procambarus clarkii, and determination of the location of its disulfide linkages

A molt-inhibiting hormone (Prc-MIH) of the American crayfish, Procambarus clarkii, a member of the type II CHH family, was chemically synthesized and the location of its three disulfide linkages was determined. Prc-MIH consists of 75 amino acid residues and was synthesized by a thioester method. Two peptide segments, Boc-[Cys(Acm)(7,24,27), Lys(Boc)(19)]-Prc-MIH(1-39)-SCH(2)CH(2)CO-Nle-NH(2) and H-[Cys(Acm)(40,44,53), Lys(Boc)(42,51,67)]-Prc-MIH(40-75)-NH(2), were prepared using peptides obtained via the Boc solid-phase method. Condensation of the building blocks in the presence of silver chloride, 3,4-dihydro-3-hydroxy-4-oxo-1,2,3-benzotriazine, and N, N-diisopropylethylamine, followed by removal of the protecting groups, gave the reduced form of Prc-MIH(1-75)-NH(2). This product was converted to the native form of Prc-MIH (synthetic Prc-MIH) in a buffer which contained cysteine and cystine. The synthetic Prc-MIH showed the same behavior by RP-HPLC and biological activity assays as the natural Prc-MIH. The disulfide bond between Cys7 and Cys44 was determined by isolation of a fragment from an enzymatic digest of the synthetic Prc-MIH by RP-HPLC, followed by mass analysis. The disulfide bonds between Cys24 and Cys40 and between Cys27 and Cys53 were determined by comparing the elution position of an enzymatic digest of the synthetic Prc-MIH with authentic chemically synthesized samples, which contained three types of possible disulfide linkages.     

Molecular Cloning and Characterization of a Microsatellite Locus Found in an RAPD Marker of a Spider Mite, Panonychus Citri (Acari: Tetranychidae)

Genetic markers were searched using PCR with 40 kinds of decanucleotide primers to investigate DNA polymorphism in Panonychuscitri. A region consisting of a variable number of CT tandem repeats (microsatellite) was found in a fragment amplified with the OPB10 primer. The microsatellite differed in size by ca. 100bp among several P. citri populations screened and was derived from at least seven alleles. This region was characteristic of P. mori and P. osmanthi, but was lacking in P. ulmi. The flanking regions were highly conserved among these species.     

Interspecific Diversity of Mitochondrial COI Sequences in Japanese Panonychus Species (Acari: Tetranychidae)

Sequences of a part of the mitochondrial cytochrome oxidase subunit I (COI) gene were analyzed in four Japanese Panonychus species to determine their phylogenetic relationships. Neighbor-Joining and maximum likelihood analysis resulted in a high bootstrap support of the relationships within the genus Panonychus. In contrast with a previous study based on ribosomal DNA data, the COI phylogeny suggested that P. mori was more distantly related to P. citri than to P. ulmi. This study shows for the first time that P. osmanthi is closely related to P. citri. Intraspecific variation analysis shows that the genetic distance between two local populations of P. mori is higher than between P. citri and P. osmanthi.