Characterization of a cDNA encoding cysteine proteinase inhibitor from Chinese cabbage (Brassica campestris L. ssp. pekinensis) flower buds

A cDNA encoding a new phytocystatin isotype named BCPI-1 was isolated from a cDNA library of Chinese cabbage flower buds. The BCPI-1 clone encodes 199 amino acids resulting in a protein much larger than other known phytocystatins. BCPI-1 has an unusually long C-terminus. A BCPI-1 fusion protein expressed in Escherichia coli strongly inhibits the enzymatic activity of papain, a cysteine proteinase. Genomic Southern blot analysis revealed that the BCPI gene is a member of a small multi-gene family in Chinese cabbage. Northern blot analysis showed that it is differentially expressed in the flower bud, leaf and root.     

Biodegradation of the mixtures of 4-chlorophenol and phenol by Comamonas testosteroni CPW301

A 4-chlorophenol (4-CP)-degrading bacterium, strain CPW301, was isolated from soil and identified as Comamonas testosteroni. This strain dechlorinated and degraded 4-CP via a meta-cleavage pathway. CPW301 could also utilize phenol as a carbon and energy source without the accumulation of any metabolites via the same meta-cleavage pathway. When phenol was added as a additional substrate, CPW301 could degrade 4-CP and phenol simultaneously. The addition of phenol greatly accelerated the degradation of 4-CP due to the increased cell mass. The simultaneous degradation of the 4-CP and phenol is useful not only for enhanced cell growth but also for the bioremediation of both compounds, which are normally present in hazardous waste sites as a mixture.     

Human xeroderma pigmentosum group G gene encodes a DNA endonuclease.

Because of defective nucleotide excision repair of ultraviolet damaged DNA, xeroderma pigmentosum (XP) patients suffer from a high incidence of skin cancers. Cell fusion studies have identified seven XP complementation groups, A to G. Previous studies have implicated the products of these seven XP genes in the recognition of ultraviolet-induced DNA damage and in incision of the damage-containing DNA strand. Here, we express the XPG-encoded protein in Sf9 insect cells and purify it to homogeneity. We demonstrate that XPG is a single-strand specific DNA endonuclease, thus identifying the catalytic role of the protein in nucleotide excision repair. We suggest that XPG nuclease acts on the single-stranded region created as a result of the combined action of the XPB helicase and XPD helicase at the DNA damage site.     

MHC antigen expression by melanomas recovered from mice treated with allogeneic mouse fibroblasts genetically modified for interleukin-2 secretion and the expression of melanoma-associated antigens

Recent approaches toward the immunotherapy of neoplastic disease involve the introduction of expression-competent genes for interleukin-2 (IL-2) into autologous malignant cells. Treatment of tumor-bearing experimental animals with the IL-2-secreting cells successfully induces partial and at times complete remissions. In most instances, however, although delayed, progressive tumor growth continues. Here, certain of the characteristic of B16 melanomas (H-2^b) persisting in C57BL/6 mice (H-2^b) treated with an IL-2-secreting, melanoma-antigen-positive cellular immunogen (RLBA-IL-2 cells) are described. Unlike the melanoma cells first injected, B16 cells recovered from mice treated with RLBA-IL-2 cells were deficient in the experssion of MHC class I, but not class II determinants. Deficient MHC class I expression correlated with the cells' resistance to cytotoxic T lymphocytes (CTL) from the spleens of mice immunized with RLBA-IL-2 cells. Melanomas persisting in mice treated with non-IL-2-secreting, melanoma-antigen-positive cell constructs (RLBA-ZipNeo cells) were also deficient in the expression of MHC class I determinants, and the melanoma cells were resistant to CTL from mice immunized with RLBA-ZipNeo cells. Thus, the expression of melanoma-associated antigens rather than IL-2-secretion correlated with deficient MHC class I expression by the persistent melanomas. This point was substantiated by the expression of MHC class I antigens by melanomas persisting in mice treated with IL-2-secreting, melanoma-antigen-negative LM cells (LM-IL-2); it was equivalent to that of melanomas in untreated mice. The involvement of MHC class I antigens in the immune resistance of persistent melanoma cells from mice treated with the melanoma-autigen-positive immunogens was indicated by the effect of interferon (IFN) orN-methyl-N-nitro-N-nitrosoguanidine (MNNG) on the susceptibility of the cells to anti-melanoma CTL. Treatment of the resistant melanomas with IFN or MNNG stimulated MHC class I antigen expression and restored the cells' sensitivity to CTL from mice immunized with IL-2-secreting or nonsecreting, melanoma-antigen-positive cellular immunogens. Prior treatment of the treated cells with antibodies to MHC class I determinants inhibited the cells' susceptibility to CTL from mice immunized with RLBA-IL-2 cells.     

Larval development of Crangon hakodatei Rathbun (Decapoda: Crangonidae) reared in the laboratory

Complete larval development of Crangon hakodatei Rathbun isdescribed, based on material hatched in the laboratory fromovigerous females. The species has six zoeal stages and onepostlarval stage. The morphological characters of the larvaland postlarval stages are described with illustrative figuresand compared with those of two congeneric species. The zoealstages of C. hakodatei can be distinguished from those of otherCrangon species in the number of segments of the antennule peduncle,the number of setae on the antennal scale and basis of the maxillipeds,and the stages of appearance of pereiopods. The first zoealstage in the seven species of Crangon are compared and an annotatedkey for distinguishing them is also provided.     

Effect of Root Zone Temperature on the Mineral Composition of Xylem Sap and Plasma Membrane K+-Mg++-ATPase Activity of Grafted-Cucumber and -Figleaf Gourd Root Systems

Cucumber (Cucumis sativus L.) seedlings were grafted onto cucumber-(CG) or figleaf gourd- (FG, Cucurbita ficifolia Bouché)seedlings in order to determine the effect of solution temperature(12, 22, and 32°C) on the mineral composition of xylem sapand the plasma membrane K⁺-Mg⁺⁺-ATPase activities of the roots.Low solution temperature (12°C) lowered the concentrationof NO^–₃ and H₂PO^–₄ in xylem sap of CG plants butnot of FG plants. Concentrations of K⁺, Ca⁺⁺ and Mg⁺⁺ in xylemsap were less affected than anions by solution temperature.The plasma membrane of FG plants grown in 12°C solutiontemperature showed the highest K⁺- Mg⁺⁺-ATPase activity at allATP concentrations up to 3 mM and at low reaction temperatureup to 12°C, indicating resistance of figleaf gourd to lowroot temperature. (Received December 27, 1994; Accepted March 10, 1995)     

Identification of three genetic loci controlling leaf senescence in Arabidopsis thaliana

Four mutants that show the delayed leaf senescence phenotype were isolated from Arabidopsis thaliana . Genetic analyses revealed that they are all monogenic recessive mutations and fall into three complementation groups, identifying three genetic loci controlling leaf senescence in Arabidopsis . Mutations in these loci cause delay in all senescence parameters examined, including chlorophyll content, photochemical efficiency of photosystem II, relative amount of the large subunit of Rubisco, and RNase and peroxidase activity. Delay of the senescence symptoms was observed during both age-dependent in planta senescence and dark-induced artificial senescence in all of the mutant plants. The results indicate that the three genes defined by the mutations are key genetic elements controlling functional leaf senescence and provide decisive genetic evidence that leaf senescence is a genetically programmed phenomenon controlled by several monogenic loci in Arabidopsis . The results further suggest that the three genes function at a common step of age-dependent and dark-induced senescence processes. It is further shown that one of the mutations is allelic to ein2-1 , an ethylene-insensitive mutation, confirming the role of ethylene signal transduction pathway in leaf senescence of Arabidopsis .     

The role of EMF1 in regulating the vegetative and reproductive transition in Arabidopsis thaliana (Brassicaceae)

To understand the genetic regulation of vegetative to reproductive transition in higher plants, further characterization of the Arabidopsis mutant embryonic flower1, emf1, was conducted. Using three flowering symptoms, we showed that emf1 mutants could only grow reproductive and not rosette shoots under five different growth conditions. The mutant embryos did not produce the typical tunica–corpus shoot apical structures at the heart-, torpedo-, and mature stages. The divergent shoot apical development during mutant and wild-type embryogenesis indicated that the wild-type EMF1 gene was expressed in early embryogenesis. Mutations in the EMF1 gene affected the embryonic shoot apical development and caused the germinating embryo and regenerating callus to grow inflorescence, instead of rosette, shoots. Our results support the hypothesis that the EMF1 gene regulates the switch between vegetative and reproductive growth in Arabidopsis.     

Ginkgolide B production in cultured cells derived from Ginkgo biloba L. leaves

Callus cultures and cell suspension cultures derived from Ginkgo biloba L. leaves produced ginkgolidc B. In cell suspension cultures, the production reached a maximum by the 13th day of subculture and followed by a sharp decrease. The medium of Murashige and Skoog induced the highest ginkgolide B content in cultures while the medium of Schenk and Hildebrandt promoted cell growth. For the maximal production of ginkgolide B, cells were cultured in Murashige and Skoog medium modified to contain 1.0 mg/l of -naphthaleneacetic acid, 0.1 mg/1 of kinetin, 30 g/1 sucrose and 1.25 mM potassium phosphate with a molar ratio of ammonium to nitrate ions of 1 3.Abbreviations B5 Gamborg et al (1968) medium - GKB Ginkgolide B - MS Murashige and Skoog (1962) medium - NAA -naphthaleneacetic aicd - SH Schenk and Hildebrandt (1972) medium     

Seasonal ectomycorrhizal fungal biomass development on loblolly pine (Pinus taeda L.) seedlings

Ergosterol, a membrane sterol found in fungi but not in plants, was used to estimate live mycelial biomass in ectomycorrhizae. Loblolly pine (Pinus taeda L.) seeds were sown in April 1993 and grown with standard nursery culture practices. Correlations between total seedling ergosterol and visual assessment of mycorrhizal colonization were high during July and August but low as ectomycorrhizal development continued into the growing season. Percentages of mycelial dry weight over lateral roots decreased from 9% in July to 2.5% in November because seedling lateral root dry weight accumulated faster than mycelial dry weight. Total ergosterol per seedling increased from July through February. As lateral root dry weight ceased to increase during winter months, ectomycorrhizal mycelia became the major carbohydrate sink of pine seedlings. No distinctive seasonal pattern of soil ergosterol content was observed. The impact of ectomycorrhizal fungi on plant carbohydrate source-sink dynamics can be quantitatively estimated with ergosterol analysis but not with conventional visual determination.