Lysis of Bacillus subtilis Cells by Glycerol and Sucrose Esters of Fatty Acids

The lytic action of glycerol and sucrose esters of fatty acids with different carbon chain lengths on the exponentially growing cells of Bacillus subtilis 168 was investigated. Of each series of esters, glycerol dodecanoate and sucrose hexadecanoate were the most active. Lysis at 1 h after the addition of 0.1 mM glycerol dodecanoate or 20 μg of sucrose hexadecanoate per ml was 81 or 79%, respectively, as evaluated by the reduction in optical density. During this treatment a great loss of viability occurred that preceded lysis. The results that were obtained suggest that autolysis is induced by these esters. The esters caused morphological changes in the cells, but a seeming adaptation of the cells to esters was seen.     

Thresholds of retinal and extraretinal photoreceptors measured by photobehavioral response in catfish,Silurus asotus

Summary Light thresholds of retinal and extraretinal photoreceptors in catfish were examined by the photobehavioral response using a method in which reflex body movements are recorded. Thresholds were determined in six groups, (A) intact, (B) ophthalmectomized, (C) pinealectomized, (D) ophthalmectomized + pinealectomized, (E) ophthalmectomized, pinealectomized and skinless over the brain (skinless fish) and (F) ophthalmectomized, pinealectomized and dorsally covered with aluminum foil over the brain (covered fish). All these fishes displayed short term activity to white light stimulation after being dark adapted for more than 5 h. The lowest threshold was obtained in the intact group (2.0×10^–4 W/cm²). The thresholds of lateral eyes and the pineal organ were 3.4×10^–3 and 1.5×10^–2 W/cm², respectively. Without lateral eyes and pineal organ, catfish still responded to light, indicating the possible existence of extraretinal nonpineal photoreceptors (ENPs). The threshold of ENPs was 3.3 W/cm². The localization of ENPs was assumed to be in the brain from the experiment with the combination of skinless and covered fish.Abbreviation ENP extraretinal nonpineal photoreceptor     

Quantitative analysis of major axis development inViburnum dilatatum andV. wrightii (Caprifoliaceae)

The architectural development ofViburnum dilatatum andV. wrightii was investigated quantitatively. In both species, the major axis is developed from terminal buds of vegetative shoots and from axillary buds on the most distal nodes of reproductive shoots. The architecture of the two species is formed mainly by four branching patterns: a monopodial pattern (M), a sympodial pattern producing a pair of opposite daughter shoots (SP), a sympodial pattern producing a single daughter shoot (SS), and a pattern terminated with a dormant or dead bud (D). In the process of the architecture formation, four successive stages are recognized: 1) height growth, where the M pattern is dominant; 2) crown formation, in addition to the M pattern, the SP pattern occurs frequently; 3) crown expansion, the M and SP patterns are also frequent; 4) over mature, the M pattern becomes dominant again. These four stages are common to the two species, butViburnum wrightii proceeds with the crown formation stage more rapidly and stays in the crown expansion stage for a longer time thanV. dilatatum. The crown ofViburnum wrightii is thus more branched than that ofV. dilatatum.     

Maternal Effects of mto1 Mutation, That Causes Overaccumulation of Soluble Methionine, on the Expression of a Soybean {beta}Conglycinin Gene Promoter-GUS Fusion in Transgenic Arabidopsis thaliana

ß-Conglycinin, the 7S seed storage protein of soybean(Glycine max [L.] Merr.), is comprised mainly of three subunits,designated , ' and ß. Expression of the gene encodingthe ß subunit is unique because its expression hasbeen shown to be down-regulated by exogenously applied L-methioninein immature soybean cotyledon cultures in vitro. Arabidopsisthaliana strain carrying a mto1-1 mutation overaccumulates solublemethionine. By using this mutant, we analyzed the effects ofmethionine on expression of the ß subunit gene invivo. Reciprocal crosses were made between the mto1-1 mutantand a transgenic A. thaliana strain, designated SNTß3,which carries a ß-glucuronidase (GUS) reporter geneunder the control of the promoter region of the ßsubunit gene. Analysis of GUS activity in F₁ seeds indicatedthat the GUS activity was dramatically repressed when the mto1-1mutant plants were used as female parents. We constructed astrain which carries both the transgene and mto1-1 mutationin the homozygous state. Analyses of the GUS activity in seedsof this double homozygous strain indicated that the GUS activitywas repressed to 2.5% of control by introduction of the mto1-1mutation. These results indicate that the ß subunitgene promoter activity in seeds is down-regulated by maternalgenotype and suggest that soluble methionine, or its mobilemetabolite, is translocated from mother plants to repress ßsubunit gene expression in seeds. ⁵Present address: Division of Biological Sciences, GraduateSchool of Science, Hokkaido University, Kita-ku, Sapporo, 060Japan ⁶Present address: Department of Biotechnology, Faculty of Agriculture,The University of Tokyo, Bunkyo-ku, Tokyo, 113 Japan     

Influence of Molecular Sizes of Cryptococcus neoformans Capsular Polysaccharide on Phagocytosis

The role of capsular polysaccharides (CPS) of Cryptococcus neoformans in phagocytosis by murine alveolar macrophages was investigated in four strains of C. neoformans serotype A, YC-11, YC-5, YC-27 and YC-13. Phagocytosis rates increased markedly after adding 10% mouse serum, compared to fetal calf serum. The reverse relation between capsular thickness of C. neoformans and phagocytosis by alveolar macrophages was observed except in YC-27, which had thin capsules and high virulence. The phagocytosis rate in mice serum was 17.3% in YC-11 (capsule thickness 2.8-3.5 μm), 39.8% in YC-5 (capsule size 0.8-1.5 μm), 20.3% in YC-27 (capsule size 0.6-1.1 μm), and 62.8% in YC-13 (capsule not detected microscopically). The CPS of YC-11, YC-5, and YC-27 analyzed by gel-filtration using CL-2B showed high molecular fractions near the void volume. However, the CPS of YC-13 showed only low molecular fractions. The widely eluted CPS of YC-11 was separated into 3 fractions and each fraction was added in the phagocytosis assay of YC-13. Phagocytosis was markedly suppressed particularly by the addition of a higher molecular fraction. These results suggest that phagocytosis of C. neoformans by alveolar macrophages is influenced by the molecular sizes of the CPS.     

A human gene that restores the DNA-repair defect in SCID mice is located on 8p11.1→q11.1

In order to map the gene that is responsible for the DNA-repair defect in severe combined immune deficient (SCID) mice, a mixture of microcells independently isolated from mouse A9 cells containing pSV2neo-tagged human chromosomes 5, 7, 8, 9, 11, 15, 18 or 20 were fused with SCID fibroblast cell lines SCVA2 and SCVA4, which were originally established from lung tissue of the C.B.17-scid/scid mouse by SV40 virus transfection. After irradiation with ⁶⁰Co -rays and selection with antibiotic G418, 12 independent clones were obtained, of which 4 contained an intact chromosome 8, 3 clones contained a deleted chromosome 8 [del(8)q22qter or del(8)q23 qter] and remaining 5 had no detectable or specific human chromosome. We further independently transferred a single human chromosome 8 or 11 into the SCVA cells via microcell fusion, and examined the radiation sensitivity of the microcell hybrids. Complementation of the radiation sensitivity was correlated with the presence of human chromosome 8 in microcell hybirds, whereas no correlation was observed in clones following the transfer of human chromosome 11. Thus, the results indicate that human chromosome 8 restored high sensitivity to ionizing radiation. A number of subclones that were radiation resistant or sensitive were isolated from the microcell hybrids. The concordance of the radiation sensitivity with the presence or absence of specific DNA fragments on chromosome 8 indicates that the human gene is located on the centromeric region of chromosome 8, i.e., 8p11.1 q11.1.     

Assay of nicotinamide deamidase activity using high-performance liquid chromatography

A rapid, simple and reproducible method has been developed for the determination of nicotinamide deamidase activity using high-performance liquid chromatography (HPLC). Nicotinic acid (NA) liberated from nicotinamide (NAA) after a 15-min enzyme reaction was determined directly by HPLC without further separation steps. Both NA, the product, and NAA, the substrate were separated by reversed-phase ion-pair isocratic chromatography and detected at 261 nm. The present method could be applied to the measurement of deamidase activity in crude cell extracts prepared from several bacterial strains. The Michaelis constant of nicotinamide deamidase in Enterobacter agglomerans was 36 μM for NAA. This method is useful for the measurement of nicotinamide deamidase from various sources.     

Rice phloem thioredoxin h has the capacity to mediate its own cell-to-cell transport through plasmodesmata

Rice (Oryza sativa L.) phloem sieve tubes contain RPP13-1, a thioredoxin h protein that moves around the plant via the translocation stream. Such phloem-mobile proteins are thought to be synthesized in the companion cells prior to being transferred, through plasmodesmata, to the enucleate sieve-tube members. In this study, in-situ hybridization experiments confirmed that expression of RPP13-1 is restricted to companion cells within the mature phloem. To test the hypothesis that RPP13-1 enters the sieve tube, via plasmodesmata, recombinant RPP13-1 was expressed in Escherichia coli, extracted, purified and fluorescently labeled with fluorescein isothiocyanate (FITC) for use in microinjection experiments into tobacco (Nicotiana tabacum L.) mesophyll cells. The FITC-RPP13-1 moved from the injected cell into surrounding cells, whereas the E. coli thioredoxin, an evolutionary homolog of RPP13-1, when similarly labeled and injected, failed to move in this same experimental system. In addition, co-injection of RPP13-1 and FITC-dextrans established that RPP13-1 can induce an increase in plasmodesmal size exclusion limit to a value greater than 9.4 but less than 20 kDa. Nine mutant forms of RPP13-1 were constructed and tested for their capacity to move from cell to cell; two such mutants were found to be incapable of movement. Crystal-structure prediction studies were performed on wild-type and mutant RPP13-1 to identify the location of structural motifs required for protein trafficking through plasmodesmata. These studies are discussed with respect to plasmodesmal-mediated transport of macromolecules within the companion cell-sieve tube complex. Received: 6 June 1997 / Accepted 25 June 1997     

Clothing microclimate temperatures during thermal comfort in boys,young and older men

To examine the effects of age-related differences in thermoregulatory function on the clothing microclimate temperature (T _m) andT _m fluctuations while maintaining thermal comfort in daily life, 5 boys (group B, 10–11 years), 5 young men (group Y, 20–21 years) and 5 older men (group O, 60–65 years) volunteered to take part in this study. The subjects were asked to maintain thermal comfort as closely as possible in their daily lives.T _m (temperatures between the skin surface and the innermost garment) at four sites (chest, back, upper arm, and thigh), skin temperature on the chest (T _chest) and ambient temperature (T _a) were measured over a period of 8–12 h from morning to evening on one day in each of the seasons, spring, summer, autumn, and winter. Records of ability to maintain thermal comfort and of adjustment of their clothes were kept by each subject.T _a during periods of thermal comfort did not differ among the groups in any of the seasons. In group Y,T _m was significantly lower at the thigh than at the other sites in spring, autumn, and winter (P<0.05) and fluctuations (CV) ofT _m were significantly larger at the thigh than at other sites in autumn and winter (P<0.05). Similar tendencies were observed forT _m and CV ofT _m in group B. However,T _m and CV ofT _m in group O did not differ by site except for the autumnT _m. Group O had a smaller CV at the thigh in winter (P<0.05), compared to groups B and Y, suggesting a smaller regional difference inT _m fluctuation in group O. Group O adjusted their clothes even on the lower limbs (together with upper body) in order to maintain thermal comfort in accordance with changes inT _a, while groups B and Y did so only on their upper bodies. These results sugest that compared to boys and young men, lower thermoregulatory function in older men may affectT _m and CV ofT _m as a result of clothing on lower limbs being adjusted differently in order to maintain thermal comfort.     

cDNA Cloning of a Wounding-Inducible Gene Encoding a Plastid {omega}-3 Fatty Acid Desaturase from Tobacco

A cDNA encoding the plastid