Molecular organization of the human CD3 gene family on chromosome 11q23

The genes encoding three invariant components of the human T-cell antigen receptor, the CD3 , , and chains, are located on human chromosome 11 at band q23. We isolated cosmid clones containing the human CD3 and chain genes in vectors designed for rapid and efficient chromosome walking. The human CD3 gene was located in the region immediately downstream of the CD3 and genes using synthetic oligonucleotide probes and the localization of this gene confirmed by DNA sequencing. Detailed restriction mapping of the CD3 locus demonstrated that all three CD3 subunits are encoded within 60 kb of DNA with the CD3 gene located 26 kb downstream of the CD3 and genes. Analysis of genomic DNA on pulsed field gels using probes isolated from these cosmid clones defined a physical map of 750 kb spanning the CD3 locus on human chromosome 11g23. The CD3 genes thus comprise a multigene family encoding cell surface components important for transmembrane signaling on T lymphocytes. The arrangement of these genes suggest that they may share common regulatory elements for the control of gene expression during T-cell ontogeny.     

Lectin cytochemical evaluation of somatosensory neurons and their peripheral and central processes in rat and man

Summary Paraffin sections of the trigeminal nerve root of the rat, and human spinal nerve root and trigeminal ganglion were stained with a battery of lectin-horseradish peroxidase conjugates to localize and characterize glycoconjugate (GC) in situ. In the rat the myelin sheath of the peripheral segment contained GC with sialic acid most probably linked to the penultinate disaccharide galactose(1 4)-N-acetylglucosamine (Gal(1 )-GlcNAc), and complex type N-glycosidic side chains. The myelin sheath in the central segment differed in containing little if any of the GC named above and in containing GC with terminal -Gal linked to N-acetylgalactosamine (GalNAc), terminal GalNAc and fucose. Schwann cells stained for GC with GlcNAc or mannose whereas oligodendroglia stained for GC with the terminal disaccharide Gal-(1 3)-GalNAc and N-glycosidic side chains, especially in presumed Golgi zones, but also in processes continued as the outer myelin sheath. The human myelin sheath in the central segment differed from that of the rat in not staining with lectins specific for fucose and terminal GalNAc. Sialic acid and terminal -Gal were seen in the human central segment but these sugars appeared to bind to astroglial structures rather than to the myelin sheath as in the rat. Astrocytes in both rat and man were stained by two fucose-binding lectins. Several lectins revealed affinity for GC in the neurilemmal sheath, and staining of this structure was stronger in the human specimens. Neurons in the human trigeminal ganglion ranged from unstained to strongly positive for fucoconjugate in cytoplasmic bodies and plasmalemma. Positive ganglion cells gave rise to unmyelinated fibers which also stained for fucoconjugate. Remak fibers and their extensions into the substantia gelatinosa of the human spinal cord stained strongly for content of fucose.The stronger lectin affinity for N-glycosidic core sugars in the peripheral as compared with the central segment suggests that lectins localize P_o protein in peripheral myelin. The reactivity for several sugars in the central segment can possibly be attributed to myelin-associated glycoprotein (MAG) of central myelin, but lectin staining for GalNAc shows in addition a biochemically unrecognized GC with O-glycosidic linked oligosaccharides in myelin. The lectin cytochemistry indicates that the 170 K Dalton glycoprotein with PNA affinity obtained from rat sciatic nerves occurs in nodes of Ranvier.This research was supported by NIH Grants AM-10956, HL-29775 and United Health and Medical Research Foundation of South Carolina, Inc. Grant No. 79     

Effect of pH on microbial degradation of leaf litter in seven streams of the English Lake District

Summary Rates of degradation of alder, oak and grass leaf packs with associated microbial populations were measured in seven streams pH 6.8–4.9. Streams were chosen from upland and lowland sites of the same river for contrasts in pH, water chemistry and riparian vegetation. The most important factor governing rates of degradation is the physical and chemical nature of the leaf material. At pH 6.8 rates of degradation, k, and microbial colonization were higher than at pH 5.5: k on alder x6; on oak x2; on grass x2. At lowland sites, pH 6.8, higher decay rates were associated with high levels of microbial colonization including c.14 spp of aquatic hyphomycete fungi—regardless of riparian vegetation. Decay rates were similar at upland sites, pH 6.8 and 6.6, involving high levels of colonization by fewer fungal species and fewer bacteria—regardless of riparian vegetation-though grass was barely degraded at upland sites of any pH. At pH 5.5, slow decay rates were associated with low levels of microbial colonization and few fungal species. Largest microbial populations at low pH associated with riparian trees did not lead to markedly increased decay rates. Factors of water chemistry at low pH appear to inhibit microbial metabolism. The implications of these findings for stream invertebrates active in the winter is discussed.     

Evaluating a ‘Free Ion Activity Model’ applied to metal uptake by <Emphasis Type="Italic">Lolium perenne</Emphasis> L. grown in contaminated soils.

We investigated several formulations of the free ion activity model (FIAM) as a means of describing plant uptake of soil Cd and Zn from contaminated soils. Lolium perenne was grown on a range of urban and metal-spiked agricultural soils selected to provide a wide range of Cd and Zn concentrations, pH values and other physico-chemical properties. Plants were grown under controlled conditions and above-ground biomass was harvested at regular intervals. Concentrations of Cd and Zn in the grass were compared with estimates of metal capacity (total or radio-labile metal content in the soil) and intensity (metal concentration in the soil solution or free divalent ion activity). The results suggested that capacity terms alone were poor predictors of plant metal uptake (r² values between 0.001 and 0.43), while metal ion intensity provided quite reasonable predictions of the variation observed for several harvests of the grass (r²=0.60–0.87). Soil solution-to-plant transfer factors were highly pH-dependent which may suggest significant competition between trace metals and protons for sorption sites on roots. However, resolution of this question was confounded because of the strong co-variance between pH and p(M²⁺) in the soil pore water. Thus the influence of pH could not be separated from the effect of changing metal ion activity on uptake rate. Other possible effects on metal uptake such as dilution from increased biomass during growth and competition for uptake between different metal ions (Zn vs. Cd), or with Ca²⁺, appeared to play very minor roles in determining bioavailability. Several formulations of the FIAM failed to provide a consistently superior prediction of metal uptake when compared to purely empirical regression with pH and p(M²⁺) within the range of the data used to parameterise the models.     

Cellular redox state and activating protein-1 are involved in ascorbate effect on calcitriol-induced differentiation

Summary Ascorbate has been related to the differentiation of several mesenchymal cells including haematopoietic cells. We have previously demonstrated that ascorbate enhances the activity of 1,25-dihydroxyvitamin D₃ (1,25(OH)₂D₃) on monocytic differentiation of HL-60 cells. Here, we show that ascorbate-mediated modification of cellular redox state and AP-1 (activating protein-1) DNA binding during early phases are related to the enhancing effect of ascorbate on differentiation. Ascorbate, but not its fully oxidized form, dehydroascorbate, or an ascorbate analogue with a low rate of oxidation, ascorbate-2-phosphate, enhanced the differentiation induced by 1,25(OH)₂D₃, modified cytosolic reactive oxygen species levels and mitochondrial redox potential (_m), and modulated AP-1 DNA binding in HL-60 cells. Ascorbate itself increased AP-1 binding to DNA in noninduced cells, whereas it inhibited AP-1 binding in 1,25(OH)₂D₃-induced cells. However, ascorbate increased the mRNA levels ofc-jun, junB. andc-fos in 1,25(OH)₂D₃-induced cells. Taken together, these results suggest that the enhancing effect of ascorbate on HL-60 differentiation induced by 1,25(OH)₂D₃ is related to its effect on the cellular redox state and the modulation of AP-1 activity.Abbreviations 1,25-(OH)₂D₃ 1,25-dihydroxyvitamin D₃ - _m mitochondrial transmembrane potential - AP-1 activating protein-1 - Asc-2-P ascorbate-2-phosphate - DHA dehydroascorbate - DiOC₆(3) 3,3-dihexyloxacarboxyanine iodide - EMSA electromobility shift assay - NBT nitroblue tetrazolium - ROS reactive oxygen species     

Factors Controlling the Geochemical Partitioning of Trace Metals in Estuarine Sediments

The geochemical partitioning of trace metals in sediments is of great importance in risk assessment and remedial investigation. Selected factors that may control the partitioning behavior of Cu, Pb and Zn in non-sulfidic, estuarine sediments were examined with the use of combined sorption curve—sequential extraction analysis. This approach, which has not been previously used to examine estuarine sediments, allowed determination of sorption parameters for Cu, Pb and Zn partitioning to individual geochemical fractions. Partitioning behavior in sulfidic sediments was also determined by sequentially extracting Cu, Pb, and Zn from synthetic sulfide minerals and from natural sediment and pure quartz sand after spiking with acid-volatile sulfide (AVS). Trace metal sorption to the “carbonate” fraction (pH 5, NaOAc extraction) increased with metal loading due to saturation of sorption sites associated with the “Fe-oxide” (NH₂OH·HCl extraction) and “organic” (H₂O₂ extraction) fractions in non-sulfidic sediments. Freundlich parameters describing sorption to the “Fe-oxide” and “organic” fractions were controlled by the sediment Fe-oxide and organic carbon content, respectively. Sequential extraction of Cu from pure CuS, AVS-spiked sediment and AVS-spiked quartz sand showed that AVS-bound Cu was quantitatively recovered in association with the “organic” fraction. However, some AVS-bound Pb and Zn were recovered by the NH₂OH·HCl step (which has been previously interpreted as “Fe-oxide” bound metals) in the sequential extraction procedure used in this study. This indicates that the sequential extraction of Pb and Zn in sulfidic sediments may lead to AVS-bound metals being mistaken as Fe-oxide bound species. Caution should therefore be exercised when interpreting sequential extraction results for Pb and Zn in anoxic sediments.     

Sialic acid-binding lectins: submolecular specificity and interaction with sialoglycoproteins and tumour cells

We examined the specificity of limulin,Limax flavus agglutinin (LFA) andSambucus nigra agglutinin I (SNA I) at the submolecular level of sialic acid, and characterized their interactions with a panel of structurally distinct sialoglycoproteins. In haemagglutination inhibition assays NeuAc--glycosides were stronger inhibitors for limulin and LFA than nativeN-acetylneuraminic acid (NeuAc). TheN-acetyl of NeuAc was crucial for binding to both lectins. N-thioacetylated NeuAc lost affinity for LFA, but still bound to limulin. Thus, distinct intermolecular interactions are involved in binding of sialic acid to the lectins. The glyceryl side chain was required for interaction with LFA, but not with limulin. SNA I specifically bound NeuAc2 6Gal1 4Glc, but not monomeric sialic acids. Limulin and LFA strongly interacted with O-chain glycoproteins, whereas SNA I preferred N-chain proteins that carry NeuAc2 6 residues. The lectins were compared with those fromCepaea hortensis andTachypleus tridentatus (TTA) and to wheat-germ agglutinin, and were then used to probe tumour cell lines for cell surface sialylation. With the exception of TTA, all lectins interacted with the tumour cells. Limulin distinguished between the low (Eb) and highly (ESb) metastatic mouse lymphoma lines by selectively agglutinating sialidase-treated ESb cells.Abbreviations BSM bovine submaxillary mucin - CHA I Cepaea hortensis agglutinin I - LFA Limax flavus agglutinin - NeuAc N-acetylneuraminic acid - OSM ovine submaxillary mucin - SNA I Sambucus nigra agglutinin I - THP Tamm-Horsfall protein - TTA Tachypleus tridentatus agglutinin     

Heavy-metal accumulation in crops grown on sewage sludge amended with metal salts

Poplar (Populus euramericana Robusta), oats (Avena sativa L. Leander), maize (Zea mays L. Ona 36), English ryegrass (Lolium perenne L.), butter head lettuce (Lactuca sativa L. Reskia), spinach (Spinacia oleracea L. Subito) and French beans (Phaseolus vulgaris Prelude), were grown in pots with pure sewage sludge (pH 6.7), amended with Cd, Cr, Cu, Ni, Pb and Zn acetates, either added singly or in combination, to study metal effects on plant growth and metal uptake. Phytotoxic metal doses varied with metal and plant species, increasing in the order Cd<Ni<Cu<Zn<Cr and Pb. The threshold dose of toxic metals applied in combination was generally lower than that of metals given singly. Addition of Cd, Ni and Zn was clearly reflected in the respective plant concentrations. This was much less so for Cu, whereas Cr and Pb concentrations were not affected in most plant species. Critical plant (leaf) metal concentrations were lower for metals applied in combination than for single metals. Because of such phenomena the use of critical levels as a diagnostic tool for determining potential multiple metal toxicity is limited.     

Regulation of α-amylase activity in Amaranthus caudatus seeds by methyl jasmonate, gibberellin A3, benzyladenine and ethylene

Methyl jasmonate (JA-Me) at 10^–3 Minhibited Amaranthus caudatus seed germination anddecreased -amylase activity. Exogenous gibberellin A₃(GA₃) and ethylene, but not benzyladenine (BA), increased activity ofthe enzyme in the presence of JA-Me, with ethylene being the most effective. Theinhibitor of ethylene action, 2,5-norbornadiene (NBD) inhibited seed germinationand decreased -amylase activity. The inhibitory action of JA-Me onAmaranthus caudatus seed germination is associated with theinhibition of -amylase activity. Exogenous GA₃ and ethylenecontrol both -amylase activity and seed germination in the presence of JA-Me.     

Cytosolic cAMP-dependent protein kinase of Polysphondylium violaceum: developmental regulation and properties

Summary The cellular slime mould Polysphondylium violaceum contains a cAMP-dependent protein kinase resembling the mammalian type I enzyme. The appearance of this enzyme is developmentally regulated. The level of kinase activity is very low in vegetative cell and increases more than tenfold during differentiation.The catalytic subunit of this cAMP-dependent protein kinase has a native molecular weight of 60–80 kDa, an isoelectric point of 5.7 and an apparent K_m for ATP and Kemptide of 50 and 13.4 µM respectively. It is characterised by its sensitivity to a synthetic inhibitor specific for cAMP-dependent protein kinase. The regulatory subunit has a molecular weight of 50 kDa.Abbreviations HEPES N-2-Hydroxyethylpiperazine-N-2-ethane sulphonic acid - EDTA ethylenediamine tetraacetic acid - EGTA ethyleneglycol-bis-(ßaminoethyl ether)-N,N,N,N-tetraacetic acid - SDS sodium dodecyl sulphate     

The skeletal isotopic composition as an indicator of ecological and physiological plasticity in the coral genus<Emphasis Type="Italic"> Madracis</Emphasis>

Three species of the reef coral genus Madracis display skeletal isotopic characteristics that relate to depth, colony topography, and consequently to coral physiology. The joint interpretation of skeletal ¹³C and ¹⁸O provides information on the ecological plasticity and adaptation to depth of a coral species. Isotopic results are most easily understood in terms of kinetic effects, which reduce both ¹⁸O and ¹³C below isotopic equilibrium values, and metabolic effects, which only influence the skeletal ¹³C. Madracis mirabilis is adapted to depths shallower than 20 m, and shows the greatest range in kinetic effects and the strongest metabolic ¹³C enrichments caused by symbiont photosynthesis. Madracis formosa lives deeper than 40 m, and shows a reduced range of kinetic effects and relatively weak metabolic ¹³C enrichments. Madracis pharensis inhabits depths from 5 to >60 m, and does not attain the strength of kinetic effects of either of the other two species, apparently because it is not quite as well adapted to rapid growth at either extreme.     

Enzymatic studies on ninhydrin-positive compounds exuded by root tips of pea seedlings

Summary The action of L-amino-acid oxydase, D-amino-acid oxydase and L-glutamic decarboxylase on ninhydrin-positive compounds present in root-tip exudates and root-scraping extracts of pea seedlings, is described. Results of Fukuda et al. are confirmed as to the presence of D-alanine in -glutamyl-alanine.The possible inhibitory effects of -L-glutamyl-D-alanine and of Unknown Y (most probably 2-alanyl-isoxazolin-5-one) on bacterial growth are discussed. re]19760809     

Starch hydrolysing enzymes in the developing barley grain

Summary The enzymes -amylase (-1, 4-glucan 4-glucanohydrolase, 3.2.1.1), -amylase (-1,4-glucan maltohydrolase, 3.2.1.2) and phosphorylase (-1,4-glucan: orthophosphate glucosyltransferase, 2.4.1.1) were assayed in whole grains of barley throughout the maturation period. -amylase and phosphorylase had peaks of activity between 25 and 30 days after anthesis. On the other hand the activity of -amylase in both the available and latent forms reached a maximum value at 35 days after anthesis which did not decrease thereafter. -amylase activity was also assayed throughout development in the endosperm, aleurone, testa pericarp and embryo. Latent -amylase reached a constant maximum value in endosperm at 35 days but available -amylase reached a peak of activity at 25 days and then declined to zero at 45 days. Only latent -amylase was associated with the aleurone layer and activity rose to a maximum value at 35 days. The testa pericarp had mainly latent -amylase whose activity fell from an early maximum at 21 days to zero at 35 days. No hydrolytic activity was associated with the embryo. The phosphorylase activity was low and mainly associated with the endosperm fraction.     

Stoffwechsel von Isoflavonen und Cumöstanen in Zell- und Callussuspensionskulturen von Phaseolus aureus Roxb.

Summary The isoflavone daidzein, the coumestanes coumestrol and soyagol as well as 2,4,4-trihydroxychalcone were isolated from callus and cell suspensions of root tip tissue from Phaseolus aureus Roxb. Upon prolonged culturing callus suspensions gradually became cell suspensions, a process which was accompanied by a decrease in the accumulation of phenolics. Upon transfer of the cells into 3 different media containing -indolyl acetic acid, kinetin or -naphthalene acetic acid, a drastic increase in the amount of coumestrol was measured.The data are discussed in relation to the observed differentiation of the cultures after application of the various hormones. The cultures were shown to metabolize daidzein and other phenylpropanoid compounds. A pronounced binding of daidzein to polymeric, ethanol-insoluble material is discussed in relation to our earlier findings in isoflavone metabolism.     

The good,the bad and the ugly: science,aesthetics and environmental assessment

The question is raised, whether there are peculiarly scientific values which can be applied in environmental assessment. The use of the expression scientific interest is traced from its 19th century origins to modern British statutes. It is argued that attempts to replace expert judgements by objective scientific criteria (e.g., indices of biodiversity) can never be completely successful. In particular, interest is an aesthetic atribute particularly valued by scientists but incapable of precise measurement. While science provides the best framework for informed judgements on conservation issues, the judgements of scientists are inevitably distinct from their experimental results. Judgements rest on ethical and aesthetic values such as importance and interest, which are essential constituents of the scientific sub-culture, but which are not uniquely scientific.     

Hyperproduction of a recombinant fusion protein of Staphylococcus aureus V8 protease in Escherichia coli and its processing by OmpT protease to release an active V8 protease derivative

The expression of a recombinant fusion protein including Staphylococcus aureus V8 protease was studied by using Escherichia coli as the host strain. When the mature V8 protease was expressed as a fusion protein with a truncated E. coli \-galactosidase (\-gal97S4D), we could not obtain a sufficient amount of the enzyme because of the toxicity resulting from the expressed protease activity. Synthesis of V8 protease was increased by constructing a sandwich-type fusion protein consisting of \-gal197S4D, a V8 protease derivative with the 56 C-terminal amino acids deleted (V856) and a truncated aminoglycoside-3'-phosphotransferase. This fusion protein was successfully produced as inactive inclusion bodies. To release the V856 protease from the fusion protein, we developed a novel processing method using an endogeneous E. coli OmpT protease, which can recognize the dibasic amino acid residues located in the linker peptides of the fusion protein. After solubilizing the inclusion bodies with urea, the V856 protein was automatically released from the fusion protein by the OmpT protease, which was coprecipitated with the inclusion bodies. The V856 protease thus obtained showed the same enzymatic activity as that of the native V8 protease. We demonstrate in this study that the N-terminal prepro sequence and the C-terminal repeated sequence of this enzyme are not necessary for its enzymatic activity and protein folding.     

The responses of one class of neurons in the optic tract of crayfish (Procambarus) to monochromatic light

Summary Spectral sensitivity curves for four sustaining neurons in the optic tracts of Procambarus clarkii were determined under dark-adapted and chromatic light-adapted conditions. The _max in the dark-adapted state is at 570 to 575 nm, and shifts to longer wavelengths in the violet-light-adapted state (Fig. 4). Red-light-adaptation suppresses the sensitivity of the yellow-green receptors of the eye and alters the discharge pattern of the sustaining neurons, thereby exposing an input with a _max at 445 nm from the blue-sensitive receptors (Fig. 5). Such data raise the possibility that sustaining neurons may carry information that functions in color vision.This work was supported by a predoctoral fellowship FO1-GM-31,230 and then a training grant 2TO1-GM00836 to D.L.T. and grant NB-05423 to J.L.L. and Biomedical Sciences Support Grant 5-S05 FR-07091, all from the U.S.P.H.S.     

Effects of Cu,Pb and Zn on two Potamogeton species grown under field conditions

Two common macrophyte species, Potamogeton perfoliatus L. and Potamogeton pectinatus L. were grown for 12 weeks at shallow depths in sediments contaminated with 1250 or 2500 g Pb or Cu and/or Zn (gDW sediment)^-1. Control experiments were run at background levels of 4, 13, and 38 g Pb, Cu and Zn (gDW sediment)^-1, respectively. Effects of heavy metals on biomass production and metal uptake and distribution in plants are presented in relation to total amount and plant-available fraction of metals in the sediment.All three studied metals gave reduced biomass production, and the toxicity of the metals decreased in the order Zn>Cu>Pb. The root/shoot biomass ratio increased for P. pectinatus, but decreased for P. perfoliatus with metal treatment. The content of any single metal was higher in shoots than in roots of plants grown on sediments not contaminated with that specific metal, but addition of that metal increased the proportion in roots. The uptake by plants of any of the heavy metals increased with increased metal addition. The magnitude of the plant-available fraction of metals of untreated sediment was Zn>Cu>Pb, and increased in contaminated sediments. Addition of Cu decreased both the plant-available fraction and the total concentration of Zn in the sediment, while increased the uptake of Zn by the plants. The opposite was found for Cu when Zn was added. P. pectinatus accumulated about twice as much Cu as P. perfoliatus. On the other hand, the concentration of Pb was higher in P. perfoliatus than in P. pectinatus, and was negligible in P. pectinatus when cultivated in untreated sediments.     

Phytoplankton diversity and cyanobacterial dominance in a hypereutrophic shallow lake with biologically produced alkaline pH

In this work, we have characterized the diversity of phytoplanktonic species in a highly alkaline and hypereutrophic shallow lake, Santa Olalla (southwestern Spain), the evolution of their relative abundances, and that of several physicochemical parameters over 2 years. In the absence of an external input of alkaline water, Santa Olallas stable high pH (average pH 9.52, with several maxima >10.5) is explained by an extremely high photosynthetic primary productivity. A variety of phytoplankton species was observed even during pH maxima. These included several species of green algae, diatoms, and euglenoids and several cyanobacteria from the orders Nostocales and Chroococcales. Quantitatively, cyanobacteria dominated. A blooming event due to Aphanothece clathrata was observed at one pH maximum, during which the diversity as measured by the Shannon-Weaver index was extremely low. Santa Olallas cyanobacteria are alkaliphilic and/or extremely alkalitolerant and appear to be responsible for the generation and maintenance of stable high-pH conditions in their environment.Communicated by W.D. Grant     

Temperature functions of thermal death in yeasts and their relation to the maximum temperature for growth

Summary Parameters of thermal death were determined in 10 strains of yeast species whose maximum temperatures for growth (T _max) ranged from 22 to 49°C. Arrhenius plots of the specific thermal death rates (k _d) formed a positional sequence at the level of the experimental points that corrresponded in all but one case to the sequence of the respective T _max values. Extrapolated k _d values at higher or lower temperatures no longer formed this sequence.The correlation of the temperature functions with T _max could be characterized in terms of a new activation parameter, for which the name thermal death activation constant is introduced. It has the following form: T.D.A. – S where H and S are respectively the apparent heat and entropy of activation of thermal death and n is the number of degrees above T _max (expressed in °K) at which the T.D.A. constant exists.Seven mesophilic yeasts had a T.D.A. constant between 72 and 79 calxmol^-1 degree^-1 at n values between 1 and 4°. This suggested that the destructive process that limits k _d in these strains is of the same species as one that contributes to the establishment of T _max. Two psychrophilic yeasts apparently had a similar T.D.A. constant but at a high n value (about 12.5°C) which suggested that in these strains T _max is governed by a destructive process unrelated to the one that underlies thermal death. The strain of the nearly thermophilic Hansenula angusta (T _max 49°C) did not fit in either group.The significance of the T.D.A. constant is discussed and expressions for H and S in terms of bond activation parameters are proposed.