The power of bioenergy-related standards to protect biodiversity.

The sustainable production of bioenergy is vital to avoiding negative impacts on environmental goods such as climate, soil, water, and especially biodiversity. We propose three key issues that should be addressed in any biodiversity risk-mitigation strategy: conservation of areas of significant biodiversity value; mitigation of negative effects related to indirect land-use change; and promotion of agricultural practices with few negative impacts on biodiversity. Focusing on biodiversity concerns, we compared principles and criteria set to address biodiversity and other environmental and social issues in seven standards (defined here as commodity-based standards or roundtables, or relevant European legislation): five voluntary initiatives related to bioenergy feedstocks, the Renewable Transport Fuel Obligation (United Kingdom), and the European Renewable Energy Source Directive. Conservation of areas of significant biodiversity value was fairly well covered by these standards. Nevertheless, mitigation of negative impacts related to indirect land-use change was underrepresented. Although the EU directive, with its bonus system for the use of degraded land and a subquota system for noncrop biofuels, offered the most robust standards to mitigate potential negative effects, all of the standards fell short in promoting agricultural practices with low negative impacts on biodiversity. We strongly recommend that each standard be benchmarked against related standards, as we have done here, and that efforts should be made to strengthen the elements that are weak or missing. This would be a significant step toward achieving a bioenergy industry that safeguards Earth's living heritage.

Glucose-induced fibronectin expression in endothelial cells is mediated by protein kinase C.

One of the crucial pathophysiological changes in diabetic angiopathy might be the glucose-induced synthesis of components of the extracellular matrix-like fibronectin. This effect has been described for endothelial and mesangial cells. In order to gain further insight into the mechanisms how glucose-induced fibronectin expression is regulated, confluent monolayers of human umbilical vein endothelial cells (HUVEC) were incubated with varying concentrations of D-glucose (5-30 mM) in a time-dependent manner. Using Westernblotting techniques with a monoclonal antibody, a 3 +/- 0.2-fold increase of the 220 kDa-signal corresponding to human fibronectin was visible after an incubation time of 6 h indicating de-novo synthesis. Using incubation times of 6 or 10 days, glucose-mediated fibronectin overexpression was still visible, reaching a maximum at 15 mM D-glucose. The corresponding maximum values were 2.4 +/- 0.3 (6 d) and 2.3 +/- 0.2 (10 d). There was a concomitant glucose-dependent onset of expression of Protein Kinase C (PKC)-isoforms PKC-delta (2.5 +/- 0.3-fold), PKC-epsilon (2 +/- 0.2-fold) and PKC-zeta (1.8 +/- 0.2-fold), which reached a maximum after 12 h and was still visible during long-term culture. Induction of fibronectin expression was also obtained using the PKC-activating phorbolester Phorbol-12-myristat-13-acetat (PMA) which mimicks glucose-induced PKC activation. Glucose-induced fibronectin expression was decreased by the PKC-inhibitor H-7 (1- [5-isoquinolinylsulfonyl]-2-methylpiperazine). These experiments suggest that: 1. Short-term induction of fibronectin expression mediated by glucose is probably PKC-triggered since concomitant induction of PKC-isoforms PKC-delta, PKC-epsilon and PKC-zeta was observed. 2. Long-term incubation with D-glucose leads to an ongoing concentration-dependent coexpression of fibronection and various PKC-isoforms. If glucose-induced, PKC-mediated de-novo synthesis of components of the connective tissue or the extracellular matrix may be important in the pathomechanism of diabetic angiopathy, has to be proven.

Biotests using unicellular algae and ciliates for predicting long-term effects of toxicants.

Test systems for predicting long-term effects with the freshwater algae Chlamydomonas reinhardi and Scenedesmus subspicatus and the ciliate Tetrahymena pyriformis were evaluated with respect to the following reference chemicals: atrazine, bromacil, diuron, methyl parathion, lindane, 3,4-dichloroaniline, pentachlorophenol, cadmium, copper, and the volatile 1,2-dichloropropane. In growth-inhibition tests under static conditions the algae revealed a higher sensitivity to the toxicants than the ciliate except for lindane and methyl parathion. Comparison of the impairment of photosynthetic efficiency (EPR, NOEC 24 hr) with the inhibition of growth (NOEC 72 hr) of S. subspicatus revealed a higher sensitivity of the EPR parameter for inhibitors of the photosynthesis. A flowthrough system was developed for long-term tests and testing of volatile and instable substances. Under flowthrough conditions C. reinhardi was more susceptible to the chemicals than under static test conditions, except for pentachlorophenol. Due to the high volatility, 1,2-dichloropropane was only tested in the flowthrough system. The data obtained from these toxicity tests provide information about effects on organisms representing different levels of the aquatic food web, possessing differences in sensitivity against toxicants. The presented flowthrough system allows the testing of volatile and instable chemicals, problematic in static test systems, and the EPR parameter is suitable for the early characterization of chemicals acting as specific inhibitors of the photosynthetic electron transport chain.

Studies on leaching from spruce twigs and beech leaves.

Leaching, depending on the acidity of the eluent, was investigated for K, Mg and Mn. The curves for the leachate concentration versus time could be described by a power function. Leaching from old needles exceeded leaching from young needles to a high degree, but the ratios of the leached elements were almost equal. A method for continuous measurements of pH change in solutions contacting plant materials was developed. The results were in agreement with data on acidity changes in water falling through spruce or beech canopies.

cDNA structure and expression of calpactin, a peptide involved in Ca2(+)-dependent cell aggregation in sponges.

Aggregation of cells of the marine sponge Geodia cydonium is mediated by an aggregation factor (AF) particle of Mr 1.3 X 10(8). It is now reported that the AF particle is associated with calpactin, which was ascribed a role in the cell-adhesion process. In order to identify the sequence similarity to other members of the lipocortin family, the cDNA of sponge calpactin was cloned and found to display an 80% sequence similarity to vertebrate calpactin II but only a 47% similarity to calpactin I. The calpactin gene, which contains the consensus sequence coding for the amino acids G-T-D-E, was expressed in Escherichia coli and subsequently purified to a 37000-Mr polypeptide. Both the p32 and the p37 are provided with approximately two Ca2+ ions/molecule and the property to bind to phospholipids. The dissociation constant (calpactin-Ca2+) was in the absence of phospholipids in the range 500-700 microM-Ca2+ but in their presence about 20-30 microM-Ca2+. On the basis of (i) inhibition studies with antibodies to calelectrin and (ii) competition experiments with soluble phospholipids (both chemically defined as well as total homologous membrane lipids) we conclude that the AF-associated calpactin and plasma-membrane-bound phospholipid(s) are involved in cell-cell aggregation in sponges.

Role of phospholipase A2 in the stimulation of sponge cell proliferation by homologous lectin.

Using the Geodia cydonium system, we showed that after incubation of competent sponge cells in the presence of lectin, phospholipase A2 was released from the cells. The substrates for this enzyme, phosphatidylethanolamine and phosphatidylcholine, were identified in the extracellular material of sponge tissue. In addition, the phospholipase A2 inhibitor calelectrin was identified by immunobiochemical techniques; this molecule was associated with the aggregation factor. Reconstitution experiments strongly suggested that phospholipase A2 catalyzed the release of arachidonic acid, which is then taken up by the cells. Intracellularly, arachidonic acid was metabolized primarily to prostaglandin E2. Inhibition studies revealed that prostaglandin E2 is involved in the ultimate increase of DNA synthesis. These findings suggest that the phospholipase A2-arachidonic acid system is involved in the matrix-initiated signal transduction pathway in sponges.

[Status and development of peripartal mortality in East Germany]. / Stand und Entwicklung der peripartalen Mortalität in der DDR.

Based on records drawn up by the Special Commission on Maternal Death Prevention, the article gives a description of the development of the peripartal mortality in the GDR during the years 1952-1987. Special regard is paid to expert opinions on the avoidability of maternal deaths and the causes of maternal mortality determined by the Special Commissions. In addition, the article analyzes all peripartal deaths recorded in the years 1984-1987 by considering the respective ages and places of death.

The separation and characterization of two forms of Torpedo electric organ calelectrin.

Two methods for extracting calelectrin, a Ca2+-regulated membrane-binding protein from the electric organ of Torpedo marmorata, have been compared and the more promising one was modified to increase the yield to 7-8 mg.kg-1 wet weight of tissue, that is 4-5 times greater than the original method. The calelectrin so obtain could be resoloved into a minor component (designated L-calelectrin) eluted from an anion-exchange column at relatively low ionic strength (100 mM NaCl) and a major component (H-calelectrin) eluted at higher ionic strength (300 mM NaCl). The two forms were also separated by chromatography on a hydrophobic resin. Electrophoresis on cellulose acetate indicated that L-calelectrin had a lower mean isoelectric point that the H-form and polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulphate showed that under reducing conditions (presence of 5% beta-mercaptoethanol) both forms migrated as single species, the L-form having a lower apparent relative molecular mass (Mr 32,000) that the H-form (34,000). Under non-reducing conditions, there was no change in the migration of L-calelectrin but the H-form was resolved into two components of Mr 34,000 and 32,000. The addition of 2 mM Ca2+ had no effect on the migration of either form. Both forms were equally recognized by an anti-calelectrin antiserum and were microheterogeneous with respect to their isoelectric points (pH 4.3-5.5) in two-dimensional gel electrophoresis. Physical measurements were carried out on the major H-form. The Stokes radius was estimated to be 3nm, corresponding to an apparent Mr of 44,000. It was unaffected by changes in ionic strength, pH or Ca2+ concentration. Analytical ultracentrifugation gave a sedimentation constant of 2.9 S and an apparent Mr of 36,000. Measurements of circular dichroism indicated that 78% of the molecule was in the alpha-helix configuration and 22% in random coil. Ca2+ had no significant effect on the conformation.

Ca2+- and phospholipid-binding properties of Torpedo electric organ calelectrin.

The Ca2+-regulated lipid-binding properties of the H- and L-forms of calelectrin present in the electric organ of Torpedo marmorata have been compared. Binding of H-calelectrin required Ca2+ in millimolar concentrations, whereas that of L-calelectrin occurred in the micromolar range. Dissociation of H-calelectrin previously bound to lipids in the presence of 2 mM Ca2+ took place only when the Ca2+ concentration was reduced to micromolar concentrations. Binding was most effective to acidic phospholipids such as phosphatidylserine. Both forms of calelectrin promoted the aggregation of membrane vesicles in the presence of Ca2+.Mg2+, Na+ and K+ inhibited the Ca2+-induced binding to phospholipid, decreasing in effectiveness in that order. Binding was also inhibited by high pH. The surface activity and hydrophobicity index showed that H-calelectrin is a hydrophilic molecule. It may represent a less active, more highly phosphorylated "down-regulated" form of L-calelectrin. The role of calcium in H-calelectrin binding to lipid appeared to be consistent with the formation of a ternary complex of the protein, an acidic lipid and Ca2+, rather than with a direct interaction of lipid with hydrophobic sequences in H-calelectrin whose accessibility is Ca2+-regulated.

Antibodies to calcium/phospholipid binding protein (calelectrin) recognize neurons, astrocytes and Schwann cells in the nervous system of rat.

We report on the identification of proteins Mr 32 kDa, 34 kDa, 68 kDa in rat brain or sciatic nerve which cross-react (by immunoblotting) with antibodies to purified Ca2+/phospholipid binding protein calelectrin from Torpedo marmorata. Immunocytochemical staining revealed that anti-calelectrin antibodies labeled cell bodies and processes of cortical neurons and astrocytes in the central nervous system (CNS). In sciatic nerve, calelectrin-like immunoreactivity was expressed in the cytoplasm of Schwann cells of adult as well as newborn rat. Additional staining occurred at periaxonal sites in mature nerve. The localization of calelectrin-related proteins in distinct cell types of the mammalian nervous system suggests a cell-specific role of this new group of proteins in Ca2+-mediated membrane processes involved in neural function.

[Some aspects of family planning and population reproduction]. / Einige Aspekte zur Familienplanung und Bevölkerungsreproduktion.

PIP: This article on the decline in fertility in the German Democratic Republic focuses on whether social policy measures can increase the number of births to a level sufficient to ensure population replacement.^ieng

A consensus amino-acid sequence repeat in Torpedo and mammalian Ca2+-dependent membrane-binding proteins.

A group of calcium-binding proteins which bind to biomembranes has recently been identified in widely different cells and tissues (refs 1-7, reviewed in ref. 8). Three of these proteins (p70, p36 and p32.5) cross-react with antiserum to calelectrin, a Ca2+-binding protein (relative molecular mass 34,000 (34K] from the ray Torpedo marmorata, giving rise to their designation as calelectrin-related proteins. We now report that calelectrin, p36 and p32.5 contain a 17-amino-acid consensus sequence which is conserved and present in multiple copies. We suggest that this sequence may be common to other members of this new group of Ca2+-binding proteins and may underlie their unusual mode of combination with biomembranes.

Characterization of calelectrin, a Ca2+-binding protein isolated from the electric organ of Torpedo marmorata.

We report a fast (less than 1 day) and efficient (2-3 mg protein/100 g tissue) isolation method for calelectrin, a protein of Mr 34,000 in the electric organ of Torpedo marmorata that binds to membranes in the presence of Ca2+. Purified protein was used to investigate the nature of its interaction with membranes and with Ca2+. Calelectrin binds to liposomes composed of total extractable lipids from the electric organ in a Ca2+-dependent and -specific manner with half-maximal binding between 3 and 7 microM free Ca2+. This binding is totally inhibited by 1 mM mercaptoethanol. It is also shown that calelectrin directly binds Ca2+ in solution by two techniques: at 1 and 10 microM Ca2+ it binds 45Ca2+ as measured by gel permeation chromatography, and it contains saturable Tb3+-binding sites that are Ca2+-displaceable. An investigation of the protein's endogenous fluorescence shows that although it contains both tryptophan and tyrosine, there is no change in the apparent quantum yield as a function of Ca2+. Ca2+-dependent hydrophobic affinity chromatography of the total soluble proteins from Torpedo electric organ shows that Torpedo calelectrin, like calmodulin and mammalian calelectrins, is specifically retained in the presence of Ca2+ and eluted by EGTA. Calelectrin also contains high-affinity sites for hydrophobic fluorescence probes such as N-phenyl-1-naphthylamine, 2-CP-toluidinylnaphthalene-6-sulfonic acid, and 1-anilinonaphthalene-8-sulfonic acid, which again unlike calmodulin, show no changes as a function of Ca2+. We conclude that calelectrin is a Ca2+-binding protein whose binding to the lipid moieties of membranes is regulated by physiological change in the Ca2+ concentration.(ABSTRACT TRUNCATED AT 250 WORDS)

Isolation of mammalian calelectrins: a new class of ubiquitous Ca2+-regulated proteins.

In a new approach to isolating proteins which participate in the Ca2+-dependent regulation of membrane traffic in animal cells, two new Ca2+-binding proteins (Mr 67 000 and 32 500) have been identified in and purified from bovine liver, brain, and adrenal medulla. These proteins specifically and reversibly bind to chromaffin granule membranes at low Ca2+ concentrations (half-maximal binding at 5.5 microM Ca2+) and greatly potentiate the Ca2+-induced aggregation of these membranes at higher concentrations (above 10 microM). In the presence of ethylene glycol bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetate, the isolated proteins have Stokes radii of 3.40 nm (Mr 67 000) and 2.53 nm (Mr 32 500) as estimated by gel filtration and therefore occur as monomers. They are slightly acidic proteins with pI's of 5.85 and 5.60. In bovine tissues, both proteins and a third protein of Mr 35 000 cross-react immunologically with each other and with Torpedo calelectrin (Mr 34 000) and are therefore identified as mammalian calelectrins. In all tissues of Torpedo marmorata tested, only a single molecular mass form of calelectrin exists, whereas multiple forms of calelectrin exist in mammalian tissues, indicating gene duplication during evolution. We suggest that the evolutionary conservation and diversification, the high tissue concentrations, and the Ca2+-specific interactions of the calelectrins make them candidates for Ca2+-dependent regulators of membrane events in animal cells.