Carbon metabolism in Bradyrhizobium japonicum bacteroids

Abstract Carbon metabolism in Bradyrhizobium japonicum bacteroids is reviewed. Additionally, the bacteroid tricarboxylic acid (TCA) cycle and its regulation under oxygen-limited conditions is considered, with emphasis on possible sites of TCA cycle rate-limiting reactions. Furthermore, we consider other adaptive pathways that may be employed by these organisms while in symbiosis. These pathways include: (1) a poly-β-hydroxy-butyrate shunt, (2) a malate-aspartate shuttle, (3) an α-ketoglutarate-glutamate shunt, (4) a modified dicarboxylic acid cycle, and (5) fermentation pathways leading to lactate, acetaldehyde and ethanol. The effects of oxygen limitation on host carbon metabolism are also considered briefly.     

The use of amphipathic maleimides to study membrane-associated proteins

A series of amphiphilic polymethylenecarboxymaleimides has been synthesized for use as sulfhydryl reagents applicable to membrane proteins. Physical properties of the compounds which are relevant to their proposed mode of action have been determined. By comparing rates of reaction in aqueous and aprotic solvents, the compounds have been shown to react exclusively with the thiolate ion. The effects of the reagents on three membrane-associated proteins are reported, and in two cases a comparative study has been made of the effects on the proteins in the absence of membranes. A mechanism is proposed whereby the reagents are anchored at the lipid/water interface by the negatively charged carboxyl group, thus siting the reactive maleimide in a plane whose depth is defined by the length of the reagent. Supporting evidence for this model is provided by the inability of the reagents to traverse membranes, and variation of their inhibitory potency with chain length when the proteins are embedded in the membrane, but not when extracted into solution. As examples of general use of the reagents to probe sulfhydryl groups in membrane proteins, the reagents have been used to (a) determine the depths in the membrane at which two populations of sulfhydryl groups occur in the mitochondrial phosphate transporter; (b) locate a single sulfhydryl associated with the active site ofD--hydroxybutyrate dehydrogenase in the inner mitochondrial membrane; (c) examine sulfhydryl groups in theD-3-glyceraldehyde phosphate dehydrogenase associated with the human red blood cell membrane.     

Two-step purification of full-length nerve growth factor receptor and maintenance of receptor-specific binding activity

A method for the purification of full-length nerve growth factor receptor (NGFRc) using membranes from three different cell lines was developed. We emphasized recovery of NGFRc that retained specific binding activity. Lipids were required to preserve binding activity during solubilization and throughout the purification procedure. Phosphatidylcholine was used for this purpose. Lectin affinity chromatography followed by high-resolution anion-exchange chromatography was used, and a 3000-fold increase in specific binding activity was obtained for NGFRc from human melanoma A875 membranes. Seven percent of the original binding activity was recovered as pure NGFRc. NGFRc binding activity eluted at 0.35 M NaCl in anion-exchange chromatography of solubilized A875, rat pheochromocytoma PC12, and human neuroblastoma MC-IXC membranes. Eight and three percent of the original binding activity were recovered as highly enriched NGFRc from membranes prepared from PC12 and MC-IXC cells, respectively. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of highly enriched, 125I-labeled NGFRc revealed several protein species. After chromatography, identification of proteins as NGFRc was verified both by immunoprecipitation using receptor-specific monoclonal antibodies and by covalent cross-linking to 125I-NGF using N-hydroxysuccinimidyl-4-azidobenzoate. Predominantly, NGFRc was recovered as a mixture of species of 80 and 160-180 kDa. Small amounts of larger species as well as smaller species were observed, consistent with minor amounts of receptor aggregation and proteolysis occurring during purification.     

An endochitinase gene expressed at high levels in the stylar transmitting tissue of tomatoes

A gene (pMON9617; Chi2;1) identified by screening a tomato pistil cDNA library has been found to encode a protein similar in sequence to class II chitinases. Using a baculovirus expression system we show that the Chi2;1 protein is an active endochitinase. The Chi2;1 protein is most similar in sequence to a previously described stylar chitinase (SK2) isolated from potato. Both proteins lack the diagnostic N-terminal cysteine-rich regions and the C-terminal vacuolar targeting signals of class I chitinases and appear to define a novel type of class II endochitinases associated with flowers. Chi2;1 is expressed predominantly in floral organs and its expression within these organs is temporally regulated. The highest level of expression is found in the transmitting tissue of the style where Chi2;1 mRNA begins to accumulate just prior to anthesis. In vegetative tissue, low levels of Chi2;1 mRNA were detected, and these levels did not increase in response to wounding or treatment with ethephon. mRNA from Chi2;1 orthologs was not detected in most other angiosperms tested, even including some members of the Solanaceae, and it is therefore unlikely that Chi2;1 is essential for stylar function. A fragment containing 1.4 kilobase pairs of 5-flanking DNA from the Chi2;1 gene was shown to drive high-level expression of an attached reporter gene in the styles of transgenic tomatoes. This fragment has utility for engineering expression of other coding regions in styles.     

Floristic diversity and co-occurrences in a subtropical broad-leaved forest and two contrasting regrowth stands in central-west Yunnan Province,China

Many of the natural forested ecosystems that still remain in mainland China are being cleared with potentially detrimental effects on woody plant species diversity on both local and regional scales. The most extensive stand of subtropical broad-leaved forest remaining in China is located in Yunnan Province. In an effort to document the influence of human-induced disturbance on Yunnan's woody flora, floristic inventories were conducted in a stand of primary forest and in regrowth stands located in its interior and along its outer margin in the Xujiaba Nature Sanctuary in the Ailao Mountain Range. Of particular interest was the location of the disturbance relative to the primary forest source area. A total of 134 woody plant species representing 74 genera and 43 families were recorded. The floristics of the two regrowth stands were significantly different from each other, with < 10% of their respective floras comprised of co-occurring species. The interior regrowth stand had a higher number of co-occurring species with the primary forest; however, > 40% were still non-co-occurring.The principal families represented in the primary forest and the interior regrowth stand were Aquifoliaceae, Berberidaceae, Fagaceae, Lauraceae, Rosaceae, Smilacaceae, Symplocaceae, Theaceae, and Vacciniaceae. The three dominant species with relative importance values ranging from > 5% to 18% in both the primary forest and the interior regrowth stand were Castanopsis wattii, Lithocarpus jingdongensis, and Symplocos sumuntia. The edge regrowth stands had the lowest species diversity and were dominated by the native pine Pinus yunnanensis, with a relative importance of 24%. The principal families represented in the edge regrowth stand were Betulaceae, Ericaceae, Fagaceae, Myricaceae, Pinaceae, and Theaceae. Only the Fagaceae and Theaceae were well-represented in all three stands. The results of the study document the low species diversity in post-cutting regrowth on the margins of the primary forest as compared with post-cutting regrowth in the forest interior.     

What's new: To boldly glow…. Applications of laser scanning confocal microscopy in developmental biology

The laser scanning confocal microscope (LSCM)

1 LSCM: laser scanning confocal microscope; FISH: fluorescence in situ hybridisation; DiO₆: 3,3′-dihexyloxacarbocyanine iodide; NBD-ceramide: 6-((N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)amino)-caproyl)sphingosine; DiO: 3,3′-dioctadecyloxacarbocyanine perchlorate; DiI: 1,1′-dioctadecyl-3,3,3′,3′-tetramethyl-indocarbocyanine perchlorate; CCD: charge-coupled device; DIC: differential interference contrast; FURA2: (-(2-(5-carboxyoxazol-2-yl)-6-aminobenzofuran-5-oxy)-2-)2′-amino-5′-methylphenoxy)-ethane-N,N,N′,N′-tetraacetic acid, sodium salt);BCECF: 2′,7′-bis-(carboxyethyl)-5-(and-6-)-carboxyfluorescein;fluo-3: 1-(2-amino-5-(2,7-dichloro-6-hydroxy-3-oxo-3H-xanthen-9-yl)-2-(2′amino-5′-methylphenoxy)-ethane-N,N,N′,N′,-tetraacetic acid, ammonium salt; DAPI: 4′,6-diamidino-2-phenylindole, dihydrochloride; PET: positron emission tomogrophy; CT: computer-assisted tomogrophy; CiD: cubitus interruptus dominus; MRC: Medical Research Council; TOTO-1: benzothiazolium-4-quinolinium dimer; YOYO-1: benzoxazolium-4-quinolinium dimer; ex.: excitation wavelength; em.: emission wavelength.

is now established as an invaluable tool in developmental biology for improved light microscope imaging of fluorescently labelled eggs, embryos and developing tissues. The universal application of the LSCM in biomedical research has stimulated improvements to the microscopes themselves and the synthesis of novel probes for imaging biological structures and physiological processes. Moreover the ability of the LSCM to produce an optical series in perfect register has made computer 3-D reconstruction and analysis of light microscope images a practical option.     

Reconciling international human rights and cultural relativism: the case of female circumcision

How can we reconcile, in a non-ethnocentric fashion, the enforcement of international, universal human rights standards with the protection of cultural diversity? Examining this question, taking the controversy over female circumcision as a case study, this article will try to bridge the gap between the traditional anthropological view that human rights are non-existent -- or completely relativised to particular cultures -- and the view of Western naturalistic philosophers (including Lockeian philosophers in the natural rights tradition, and Aquinas and neo-Thomists in the natural law tradition) that they are universal -- simply derived from a basic human nature we all share. After briefly defending a universalist conception of human rights, the article will provide a critique of female circumcision as a human rights violation by three principal means: by an internal critique of the practice using the condoning cultures' own functionalist criteria; by identifying supra-national norms the cultures subscribe to which conflict with the practice; and by the identification of traditional and novel values in the cultures, conducive to those norms. Through this analysis, it will be seen that cultural survival, diversity and flourishing need not be incompatible with upholding international, universal human rights standards.     

IL-1α and TNFα act synergistically to stimulate production of myeloid colony-stimulating factors by cultured human bone marrow stromal cells and cloned stromal cell strains

Human bone marrow stromal cells repond to stimulation by the monokines IL-1 and TNF by producing colony-stimulating factors such as GM-CSF and G-CSF. In this study we show that IL-1α and TNFα act synergistically to stimulate GM-CSF and G-CSF production by cultured marrow stromal cells. We further show that IL-1α and TNFα synergistically stimulate production of GM-CSF and G-CSF by a clonal stroma-derived cell strain. Although IL-1 and TNF share many of the same biological activities, we show that IL-1α and TNFα have an unequal ability to induce myeloid-CSF production by both cultures, with IL-1α being the more potent inducer. We found that induction by IL-1α and TNFα was independent of cell proliferation. The effect of IL-1α and TNFα on production of the two myeloid-CSFs by the clonal cells was significantly greater than the unfractionated passaged stromal cultures, having the greater effect on G-CSF production. The clonally derived stromal cells constitutively produced colony-stimulating activity, in particular GM-CSF, at levels easily detected by ELISA. These findings show that, in addition to the overlapping and additive activities of IL-1α and TNFα, they can interact synergistically. Our findings further suggest that a small subpopulation of stroma cells may be the major producer of G-CSF in the marrow microenvironment during immune response. © 1994 wiley-Liss, Inc.