CONTRIBUTION OF POLLEN MORPHOLOGY TO SYSTEMATICS OF COLLOMIA (POLEMONIACEAE)

Pollen morphology of 14 species of Collomia (Polemoniaceae) was examined by light microscopy, and by both scanning and transmission electron microscopy. Four distinct pollen types were observed which are based principally upon 1) shape, number and distribution of apertures, and 2) surface sculpturing: Type 1—zonocolporate with striate ridges; Type 2—zonocolporate with striato-reticulate ridges; Type 3—pantoporate with radiate ridges; Type 4—pantoporate with irregularly reticulate ridges. Evaluation of pollen morphology reveals considerable discrepancy with respect to presently accepted sectional classification. Collomia grandiflora of sect. Collomia has a pollen type similar to that of members of sect. Collomiastrum and is now interpreted as representing an independent evolutionary line derived from the latter section. Collomia diversifolia of sect. Courtoisia has a pollen morphology similar to that of sect. Collomia. whereas C. heterophylla of the same section possesses pollen unique within the genus. This last pollen type shows close similarity to the pollen of members of Polemonium, Gilia, Leptodactylon, and Ipomopsis. Pollen of C. tinctoria and C. tracyi of sect. Collomia are anomalous within Polemoniaceae. No significant difference in exine stratification was discernible among the four pollen types.     

Microbial Metabolism of a Parathion-Xylene Pesticide Formulation

A mixed bacterial culture was adapted to growth on a mixed carbon substrate consisting of the pesticide parathion and its xylene-based formulation. The environmental growth parameters of temperature, pH, and dissolved oxygen concentration were optimized to obtain complete metabolism of parathion from this mixed carbon substrate. This adapted culture grew rapidly (mu = 0.7 per h) on the pesticide formulation at high parathion suspensions (3,000 mg/liter). Carbon utilization from this mixed substrate was strongly dependent on pH. At slightly acidic pH, xylene was preferentially metabolized, whereas at slightly alkaline pH, parathion was preferentially metabolized. Diethylthiophosphoric acid, a metabolite from parathion, and toluic acid, a metabolite from xylene, also influenced the selection of the primary carbon source.     

Effects of Luteolin on Human Breast Cancer Using Gene Expression Array: Inferring Novel Genes

Taraxacum officinale (dandelion) is often used in traditional Chinese medicine for the treatment of cancer; however, the downstream regulatory genes and signaling pathways mediating its effects on breast cancer remain unclear. The present study aimed to explore the effects of luteolin, the main biologically active compound of T. officinale, on gene expression profiles in MDA-MB-231 and MCF-7 breast cancer cells. The results revealed that luteolin effectively inhibited the proliferation and motility of the MDA-MB-231 and MCF-7 cells. The mRNA expression profiles were determined using gene expression array analysis and analyzed using a bioinformatics approach. A total of 41 differentially expressed genes (DEGs) were found in the luteolin-treated MDA-MB-231 and MCF-7 cells. A Gene Ontology analysis revealed that the DEGs, including AP2B1, APP, GPNMB and DLST, mainly functioned as oncogenes. The human protein atlas database also found that AP2B1, APP, GPNMB and DLST were highly expressed in breast cancer and that AP2B1 (cut-off value, 75%) was significantly associated with survival rate (p = 0.044). In addition, a Kyoto Encyclopedia of Genes and Genomes pathway analysis revealed that the DEGs were involved in T-cell leukemia virus 1 infection and differentiation. On the whole, the findings of the present study provide a scientific basis that may be used to evaluate the potential benefits of luteolin in human breast cancer. Further studies are required, however, to fully elucidate the role of the related molecular pathways.     

Functional roles of ATP-binding residues in the catalytic site of human mitochondrial NAD(P)+-dependent malic enzyme

Human mitochondrial NAD(P)+-dependent malic enzyme is inhibited by ATP. The X-ray crystal structures have revealed that two ATP molecules occupy both the active and exo site of the enzyme, suggesting that ATP might act as an allosteric inhibitor of the enzyme. However, mutagenesis studies and kinetic evidences indicated that the catalytic activity of the enzyme is inhibited by ATP through a competitive inhibition mechanism in the active site and not in the exo site. Three amino acid residues, Arg165, Asn259, and Glu314, which are hydrogen-bonded with NAD+ or ATP, are chosen to characterize their possible roles on the inhibitory effect of ATP for the enzyme. Our kinetic data clearly demonstrate that Arg165 is essential for catalysis. The R165A enzyme had very low enzyme activity, and it was only slightly inhibited by ATP and not activated by fumarate. The values of K(m,NAD) and K(i,ATP) to both NAD+ and malate were elevated. Elimination of the guanidino side chain of R165 made the enzyme defective on the binding of NAD+ and ATP, and it caused the charge imbalance in the active site. These effects possibly caused the enzyme to malfunction on its catalytic power. The N259A enzyme was less inhibited by ATP but could be fully activated by fumarate at a similar extent compared with the wild-type enzyme. For the N259A enzyme, the value of K(i,ATP) to NAD+ but not to malate was elevated, indicating that the hydrogen bonding between ATP and the amide side chain of this residue is important for the binding stability of ATP. Removal of this side chain did not cause any harmful effect on the fumarate-induced activation of the enzyme. The E314A enzyme, however, was severely inhibited by ATP and only slightly activated by fumarate. The values of K(m,malate), K(m,NAD), and K(i,ATP) to both NAD+ and malate for E314A were reduced to about 2-7-folds compared with those of the wild-type enzyme. It can be concluded that mutation of Glu314 to Ala eliminated the repulsive effects between Glu314 and malate, NAD+, or ATP, and thus the binding affinities of malate, NAD+, and ATP in the active site of the enzyme were enhanced.     

Heterology expression of the Arabidopsis C-repeat/dehydration response element binding factor 1 gene confers elevated tolerance to chilling and oxidative stresses in transgenic tomato

In an attempt to improve stress tolerance of tomato (Lycopersicon esculentum) plants, an expression vector containing an Arabidopsis C-repeat/dehydration responsive element binding factor 1 (CBF1) cDNA driven by a cauliflower mosaic virus 35S promoter was transferred into tomato plants. Transgenic expression of CBF1 was proved by northern- and western-blot analyses. The degree of chilling tolerance of transgenic T(1) and T(2) plants was found to be significantly greater than that of wild-type tomato plants as measured by survival rate, chlorophyll fluorescence value, and radical elongation. The transgenic tomato plants exhibited patterns of growth retardation; however, they resumed normal growth after GA(3) (gibberellic acid) treatment. More importantly, GA(3)-treated transgenic plants still exhibited a greater degree of chilling tolerance compared with wild-type plants. Subtractive hybridization was performed to isolate the responsive genes of heterologous Arabidopsis CBF1 in transgenic tomato plants. CATALASE1 (CAT1) was obtained and showed activation in transgenic tomato plants. The CAT1 gene and catalase activity were also highly induced in the transgenic tomato plants. The level of H(2)O(2) in the transgenic plants was lower than that in the wild-type plants under either normal or cold conditions. The transgenic plants also exhibited considerable tolerance against oxidative damage induced by methyl viologen. Results from the current study suggest that heterologous CBF1 expression in transgenic tomato plants may induce several oxidative-stress responsive genes to protect from chilling stress.     

Winter spatial distribution of fish larvae assemblages relative to the hydrography of the waters surrounding Taiwan

The aim of this study was to investigate the species composition and distribution of fish larvae in relation to hydrographic conditions in the waters surrounding Taiwan Island (TI) in February 2003. In total, 242 kinds of fish larvae belonging to 127 genera and 75 families were recognized. Among these, 109 taxa were identified to the family or genus level, others to the species level. The 12 predominant types, which constituted 71% of the total fish larvae, were Engraulis japonica, Scomber sp., Diaphus spp., Benthosema pterotum, Carangoides ferdau, Embolichthys mitsukurii, Maurolicus sp., unidentified Myctophidae, Gonostoma gracile, Trichiurus lepturus, unidentified Gobiidae, and Myctophum asperum. The distribution of fish larvae showed a clear association with water masses around TI, with higher abundances and lower species richness northwest of TI where the China Coastal Current prevails, and lower abundances and higher species diversity east of TI where the Kuroshio Current dominates. Cluster analysis distinguished three station groups and four species groups, and the distribution patterns of fish larvae also corresponded to hydrographic conditions. The total abundances of fish larvae and eight of the 12 predominant taxa showed significant and positive correlations with zooplankton abundance, which suggests that food source might be a key factor determining the abundance and distribution of fish larvae during the winter.     

Condensin-Dependent Chromatin Compaction Represses Transcription Globally during Quiescence

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Ex vivo immunological evaluation of stable mixed chimeric patients after matched related donor allogeneic transplantation in sickle cell disease

Background aimsAllogeneic hematopoietic stem cell transplantation is curative for sickle cell disease, and the use of matched related donors, non-myeloablative conditioning and sirolimus immunosuppression results in stable mixed chimerism without graft-versus-host disease (GVHD). However, the time to terminate sirolimus while maintaining mixed chimerism is unclear.MethodsIn this study, we developed a two-way mixed lymphocyte reaction (MLR) to evaluate ex vivo immunoreaction in mixed chimeric patients.ResultsIn co-culture of peripheral blood mononuclear cells (PBMCs) from two healthy controls (without irradiation), we detected proliferation at various ratios of PBMC mixtures (1:9 to 9:1) as well as various concentrations of sirolimus, suggesting that two-way MLR is applicable to patients (having >10% chimerism) undergoing sirolimus treatment. In two-way MLR using PBMCs (including donor and recipient cells) from mixed chimeric patients (n = 28), greater ex vivo proliferation was observed <6 months compared with >6 months post-transplant and healthy control PBMC monoculture. Robust ex vivo proliferation was observed in a patient with acute GVHD, and persistent ex vivo proliferation (until 2 years) was observed in a patient with decreasing donor chimerism.ConclusionsIn summary, we demonstrated that in two-way MLR, ex vivo immunoreaction decreases to low levels ~6 months post-transplant. These findings suggest a rationale to continue immunosuppression for 6 months.     

Sequence dependence of Drosophila topoisomerase II in plasmid relaxation and DNA binding

The sequence dependence of Drosophila topoisomerase II supercoil relaxation and binding activities has been examined. The DNA substrates used in binding experiments were two fragments from Drosophila heat shock locus 87A7. One of these DNA fragments includes the coding region for the heat shock protein hsp70, and the other includes the intergenic non-coding region that separates two divergently transcribed copies of the hsp70 gene at the locus. The intergenic region was previously shown to have a much higher density of topoisomerase cleavage sites than the hsp70 coding region. Competition nitrocellulose filter binding assays demonstrate a preferential binding of the intergene fragment, and that binding specificity increases with increasing ionic strength. Dissociation kinetics indicate a greater kinetic stability of topoisomerase II complexes with the intergene DNA fragment. To study topoisomerase II relaxation activity, we used supercoiled plasmids that contained the same fragments from locus 87A7 cloned as inserts. The relative relaxation rates of the two plasmids were determined under several conditions of ionic strength, and when the plasmid substrates were included in separate reactions or when they were mixed in a single reaction. The relaxation properties of these two plasmids can be explained by a coincidence of high-affinity binding sites, strong cleavage sites, and sites used during the catalysis of strand passage events by topoisomerase II. Sequence dependence of topoisomerase II catalytic activity may therefore parallel the sequence dependence of DNA cleavage by this enzyme.     

Improving the remaining activity of lignocellulolytic enzymes by membrane entrapment

The production of bioethanol by the conversion of lignocellulosic waste has attracted much interest in recent years because of its low cost and great potential availability. However, the high cost of the enzyme required for this conversion is often considered to be the major bottleneck in the commercial lignocellulosic ethanol industry. In this work, the hydrolysis of rice straw by free and entrapped lignocellulolytic enzymes (cellulase, xylanase and laccase) was carried out at pH 5.5 and 37 °C. The hydrolysis of rice straw by enzymes entrapped in a membrane produced a higher monosaccharide content: 601.05 mg/g rice straw for entrapped enzymes vs. 465.46 mg/g rice straw for free enzymes. This study has shown that enzyme entrapment is an important technique for the efficient use and reuse of enzymes in industrial applications and also for the rapid separation of saccharide products from the reaction medium, thus improving the remaining enzymatic activities.