Induction of endoreduplication in Chinese hamsters V79 cells by cytosine arabinoside

Endoreduplication (ER) could be induced very effectively in Chinese hamster V79 cells exposed to cytosine arabinoside (1-β-D-arabinofuranosylcytosine; Ara-C). Cells were cultured for 48 hours in Ara-C containing medium. ER frequency increases rapidly after Ara-C release. About 60% of metaphase cells were endoreduplicated at 8–10 hours after release from Ara-C (5 μg/ml). Induction of ER also depends on Ara-C concentrations.     

The Effect of Electrotonus on the Olfactory Epithelium

The effect of electrotonus on the slow potential of the olfactory epithelium of the frog was studied. The "on"-slow potential induced by a general odor like amyl acetate increased its magnitude in accordance with increase of anodal current, while it decreased its magnitude with increase of cathodal current. Similar relations were also found in the case of the vapors of organic solvents like ethyl ether of low concentrations. Conversely, the on-slow potential induced by the vapors of organic solvents of high concentration decreased its magnitude in accordance with the increase of anodal current, while it increased its magnitude with the increase of cathodal current. The "off"-slow potential induced by the vapors of organic solvents of high concentration showed a potential change under the action of electrotonic currents which is similar to the change of the on-slow potential induced by general odors. It was concluded that there are two receptive processes in the olfactory cell. One is an ordinary excitatory process which produces an electronegative slow potential in response to general odors. The other is a process of a different kind which is activated only by the vapor of an organic solvent of high concentration and which shows an entirely opposite reaction from that generally found in excitable tissues when an electrotonic current is applied.     

Role of ω-3 fatty acid desaturases in the regulation of the level of trienoic fatty acids during leaf cell maturation

Trienoic fatty acids, namely -linolenic acid and hexadecatrienoic acid, present in leaf lipids are produced by -3 fatty acid desaturases located in the endoplasmic reticulum and plastid membranes. The changes in the level of trienoic fatty acids during leaf maturation were investigated in wild-type plants of Arabidopsis thaliana (L.) Heynh. and in the fad7 mutant deficient in the activity of a plastid -3 desaturase. The levels of trienoic fatty acids increased in 26 °C- and 15 °C-grown wild-type plants with maturation of leaves. The increase in trienoic fatty acids was mainly due to galactolipids enriched in plastid membranes. In addition, the relative levels of trienoic fatty acids in major glycerolipids, including phospholipids enriched in the endoplasmic reticulum membranes, also increased with leaf maturation. By contrast, when the fad7 mutant was grown at 26 °C, the relative levels of trienoic fatty acids in individual lipids decreased with leaf maturation. The decreases in the levels of trienoic fatty acids, however, were alleviated when the fad7 mutant was grown at 15 °C. These results suggest that the plastid -3 desaturase plays a major role in increasing the levels of trienoic fatty acids with leaf maturation.Abbreviations 163 hexadecatrienoic acid - 183 -linolenic acid - DGD digalactosyldiacylglycerol - MGD monogalactosyldiacylglycerol - PC phosphatidylcholine - PE phosphatidylethanolamine - TA trienoic fatty acid - WT wild type - -3 refers to the position of the double bond from the methyl end of a fatty acid This research was supported in part by Grants-in-Aid for Scientific research (#07251214 and #06804050 to K.I.) from the Ministry of Education, Science and Culture, Japan, and by the research grant from Shorai Foundation.     

Establishing a new genus,Chiharadinium gen. nov. (Peridiniales,Dinophyceae) for a tidal pool dinoflagellate formerly known as Scrippsiella hexapraecingula

To determine its accurate taxonomic position, a tidal pool bloom-forming dinoflagellate, Scrippsiella hexapraecingula was re-investigated using light, scanning and transmission electron microscopy together with a phylogenetic analysis based on concatenated ribosomal DNA sequences. The culture strains used in this study were established from intertidal rock pool samples taken from Jogashima, Kanagawa prefecture and Heisaura, Chiba prefecture, Japan and were identified as S. hexapraecingula originally described by Horiguchi and Chihara from a tidal pool in Hachijo Island, Tokyo, Japan in 1983. The thecal plate arrangement was determined as Po, X, 4′, 3a, 6″, 6c, 5s, 5″′, 2″″. The internal structure was investigated for the first time. The organism has typical dinoflagellate cellular organelles such as a dinokaryotic nucleus, mitochondria with tubular cristae, trichocysts and pusule. The chloroplast was single and connected to the central pyrenoid (stalked type). The eyespot found in the sulcus is of the B type with two rows of superficial intraplastidal lipid globules directly overlain by an extraplastidal single layer of crystalline bricks enveloped by a common membrane. The apical pore is plugged by a double-layered stub-like structure. Stalk building material for attachment covered the apical pore. Phylogenetic analysis indicated that S. hexapraecingula was most closely related to a freshwater dinoflagellate, Peridiniopsis borgei, the type species of the genus Peridiniopsis. However, clear differences exist between these two organisms, including their thecal plate arrangement, habitat and habit. As a result, a new genus, Chiharadinium Dawut & T. Horiguchi gen. nov. has been proposed rather than attempting to accommodate S. hexapraecingula in the genus Peridiniopsis. The new combination, Chiharadinium hexapraecingulum (T. Horiguchi & Chihara) Dawut & T. Horiguchi comb. nov. has been proposed.     

Spectral and electrochemical properties of vanadyl hexadecafluorophthalocyanine

A vanadyl complex with perfluorinate phthalocyanine, VOPcF₁₆, was prepared. The monomer-dimer solvent dependence was confirmed based on the solvent effect for the Q-band position-that is, VOPcF₁₆ exists as a monomer in a nonpolar solvent such as benzene, but dimerizes in a polar solvent such as acetone. Electron spin resonance data also supported the solvent dependence found. In addition, the substituent effect of fluorine atoms on the redox properties was investigated by measuring the cyclic voltammograms in dichloromethane. On the reduction side, three redox couples were observed, the first two of which were assigned as being due to the reduction of the phthalocyanine ring (to LUMO), whose potentials are 0.4–0.5 V higher than those of the tetra-t-butyl and octabutoxy derivatives, VOPc(t-Bu)₄ and VOPc(O-n-Bu)₈.     

Initiation of Cyclin B Degradation by the 26S Proteasome upon Egg Activation

Immediately before the transition from metaphase to anaphase, the protein kinase activity of maturation or M-phase promoting factor (MPF) is inactivated by a mechanism that involves the degradation of its regulatory subunit, cyclin B. The availability of biologically active goldfish cyclin B produced in Escherichia coli and purified goldfish proteasomes (a nonlysosomal large protease) has allowed the role of proteasomes in the regulation of cyclin degradation to be examined for the first time. The 26S, but not the 20S proteasome, digested recombinant 49-kD cyclin B at lysine 57 (K57), producing a 42-kD truncated form. The 42-kD cyclin was also produced by the digestion of native cyclin B forming a complex with cdc2, a catalytic subunit of MPF, and a fragment transiently appeared during cyclin degradation when eggs were released from metaphase II arrest by egg activation. Mutant cyclin at K57 was resistant to both digestion by the 26S proteasome and degradation at metaphase/anaphase transition in Xenopus egg extracts. The results of this study indicate that the destruction of cyclin B is initiated by the ATP-dependent and ubiquitin-independent proteolytic activity of 26S proteasome through the first cutting in the NH₂ terminus of cyclin (at K57 in the case of goldfish cyclin B). We also surmise that this cut allows the cyclin to be ubiquitinated for further destruction by ubiquitin-dependent activity of the 26S proteasome that leads to MPF inactivation.     

CD25 T-lymphocytes induce CD11b on eosinophils in allergic nasal mucosa

In the allergic mucosa, there is a significant increase in numbers of CD25(+) cells and activated eosinophils. To determine whether a link exists between the activated T-lymphocytes and tissue eosinophils in nasal allergy, we studied CD25(+) cells in the nasal mucosa and compared the levels of soluble IL-2 receptor (sIL-2R) both in the serum and the nasal secretions, and further investigated expression of CD11b on eosinophils in the nasal lavage fluids and peripheral blood of patients with nasal allergy. We also examined the effects of the culture supernatant of Con A- and IL-2-activated T-lymphocytes on CD11b expression on eosinophils in the present study. The concentration of sIL-2R in the nasal secretions from patients with Japanese cedar pollinosis (JCP) was significantly higher than that from normal subjects (p < 0.01). The sIL-2R level was significantly higher in the nasal secretions than in the sera in patients (p < 0.01), and CD11b expression on eosinophils from nasal hvage fluid was significandy higher than that of eosinophils from peripheral blood of the same individuals (p < 0.01). The activated T-lymphocytes promoted eosinophil activation with upregulation of CD11b in vitro, and eosinophils in the nasal secretions from patients significantly expressed more CD11b in vivo. These results indicate that activation of T-lymphocytes is linked to eosinophil activation in nasal allergy.     

Identification of cells by backscattered electron imaging of silver stained bulk tissues in scanning electron microscopy

Anuran tadpole tail muscle was stained en bloc by a modified light microscope silver stain for light microscopy and freeze-fractured in liquid nitrogen after partial dehydration with ethanol. The fractured specimens were observed in both secondary electron and backscattered electron modes in a scanning electron microscope. Since the cell nuclei specifically stained with silver provided high contrast against the unstained background due to atomic number contrast of backscattered electron image, various cells were easily identified by a comparison of secondary electron images and compositional images of backscattered electron signals.     

Mast cells contain spleen-type prostaglandin D synthetase

Prostaglandin D synthetase activity in the cytosol (100,000 x g, 1-h supernatant) fraction of peritoneal mast cells of adult rats (105.0 nmol/min/mg protein) was the highest among such activities in various rat tissues and cells. As judged by the absolute requirement for glutathione for the reaction (Km = 300 microM), the Km value for prostaglandin H2 (200 microM), and insensitivity of the activity to 1 mM 1-chloro-2,4-dinitrobenzene, the enzyme in mast cells was similar to rat spleen prostaglandin D synthetase and differed from rat brain prostaglandin D synthetase or glutathione S-transferase, all of which catalyze the isomerase reaction from prostaglandin H2 to prostaglandin D2. In immunotitration analyses, the activity in mast cells showed a titration curve exactly identical with that of the purified spleen-type enzyme and almost completely absorbed by an excess amount of antibody against this enzyme, but it remained unchanged after incubation with antibodies against the brain-type enzyme and glutathione S-transferase isozymes thus far purified. In Western blot after two-dimensional electrophoresis of crude extracts of mast cells, a single immunoreactive spot was observed with antibody against the spleen-type enzyme at the same position as that of the purified enzyme (Mr = 26,000, pI = 5.2). Furthermore, the immunoreactive protein obtained from mast cells showed the same peptide fingerprints as those of the purified spleen-type enzyme, after partial digestion with Staphylococcus aureus V8 protease or trypsin. In immunoperoxidase staining, the immunoreactivity of the spleen-type enzyme was found in the cytosol of tissue mast cells in various organs such as thymus, intestine, stomach, and skin of adult rats. These findings indicate that prostaglandin D2 is produced by the spleen-type synthetase in mast cells of various tissues.