Sucrose transport into the phloem of Ricinus communis L. seedlings as measured by the analysis of sieve-tube sap

Careful cutting of the hypocotyl of Ricinus communis L. seedlings led to the exudation of pure sieve-tube sap for 2–3 h. This offered the possibility of testing the phloem-loading system qualitatively and quantitatively by incubating the cotyledons with different solutes of various concentrations to determine whether or not these solutes were loaded into the sieve tubes. The concentration which was achieved by loading and the time course could also be documented. This study concentrated on the loading of sucrose because it is the major naturally translocated sieve-tube compound. The sucrose concentration of sieve-tube sap was approx. 300 mM when the cotyledons were buried in the endosperm. When the cotyledons were excised from the endosperm and incubated in buffer, the sucrose concentration decreased gradually to 80–100 mM. This sucrose level was maintained for several hours by starch breakdown. Incubation of the excised cotyledons in sucrose caused the sucrose concentration in the sieve tubes to rise from 80 to 400 mM, depending on the sucrose concentration in the medium. Thus the sucrose concentration in the sieve tubes could be manipulated over a wide range. The transfer of labelled sucrose to the sieve-tube sap took 10 min; full isotope equilibration was finally reached after 2 h. An increase of K⁺ in the medium or in the sieve tubes did not change the sucrose concentration in the sievetube sap. Similarly the experimentally induced change of sucrose concentration in the sieve tubes did not affect the K⁺ concentration in the exudate. High concentrations of K⁺, however, strongly reduced the flow rate of exudation. Similar results were obtained with Na⁺ (data not shown). The minimum translocation speed in the sieve tubes in vivo was calculated from the growth increment of the seedling to be 1.03 m·h^-1, a value, which on average was also obtained for the exudation system with the endosperm attached. This comparison of the in-vivo rate of phloem transport and the exudation rate from cut hypocotyls indicates that sink control of phloem transport in the seedlings of that particular age was small, if there was any at all, and that the results from the experimental exudation system were probably not falsified by removal of the sink tissues.Abbreviations PTS 3-hydroxy-5,8, 10-pyrenetrisulfonate     

A similar protein portion for two exoglucanases secreted by Saccharomyces cerevisiae

Exoglucanase (exo-1,3-β-D-glucan glycohydrolase, EC 3.2.1.56) activity secreted by Saccharomyces cerevisiae into the culture medium was separated by ion exchange chromatography into two glycoprotein isoenzymes which contributed 10% (exoglucanase I) and 90% (exoglucanase II) towards the total activity. Analysis of the “in vitro” deglycosylated products by polyacrylamide gel electrophoresis under native or denaturing conditions indicated that the protein portions of both exoglucanases exhibited identical mobility, each one consisting of two polypeptides with M _r of 47000 and 48000. The same profile was shown by the exoglucanase secreted in the presence of tunicamycin. Antibodies raised against the protein portion of exoglucanase II did react with both native exoglucanases and their deglycosylated products with a pattern indicative of immunological identity. Digestion of the “in vitro” deglycosylated products of both exoglucanases with Staphylococcus aureus V-8 protease or trypsin generated the same proteolytic fragments in each case. Only exoglucanase II was secreted by protoplasts. These and previously reported results indicate that the protein portions of both isoenzymes may be the product of the same gene (or a family of related genes), and that exoglucanase I is a product of enzyme II, modified by a process occurring beyond the permeability barrier of the cell.     

Localization of the nerve growth factor receptor on fetal human schwann cells in culture

Previous studies have established that Schwann cells (SC) in culture express an NGF receptor. In this study, cultures of fetal human SC were established from fetal nerves and various light microscopic (LM) and electron microscopic (EM) techniques were used to localize the NGF receptor on the SC. Results indicate that NGF receptor is localized to the plasma membrane of the SC. Quantitative digital analysis determined that the distal portion of the SC process had high concentrations of NGF receptor. The possible functional significance of this latter observation is discussed in terms of SC migration and ensheathment of axons.     

Histochemical detection of monoamine oxidase and alcohol dehydrogenase activities in the Syrian hamster Harderian glands: existence of a sexual dimorphism

Summary Monoamine oxidase (MAO) and alcohol dehydrogenase (AD) activities were studied histochemically in the Syrian hamster Harderian gland using tryptamine as substrate and Nitroblue Tetrazolium as the final electron acceptor. No dark: light-related changes were observed. Male type I secretory cells showed an intense MAO reaction. Female type I cells exhibited a moderate MAO activity. Both male and female glands showed a moderate/intense AD-positive reaction. Male type II cells were lacking MAO and AD activities. MAO activity found in the hamster Harderian glands corresponded mainly to MAO type A since treatment with chlorgyline (0.01, 0.1 and 0.5mm) totally inhibited it. The possible role of these two enzymes in Harderian gland indolalkylamine metabolism is discussed.     

Size variation in Brachionus plicatilis resting eggs

The effect of temperature and salinity on resting egg size of two Brachionus plicatilis (Rotifers) clones was investigated. Clones were selected according to their different behaviour in laying resting eggs: one clone ejects them, whereas they remain inside the females body in the other clone. The difference in resting eggs size between the two clones is noticeable, although the difference is not as great as that between female body size. An important temperature-salinity interaction on resting egg size has been observed. The general inverse relationship between size and temperature is only true at lower temperatures. At high temperatures size varies around the mean although could be greater than at intermediate temperatures. This is more evident at the intermediate salinity tested which is considered to be the closest to the optimum in our experiments. This pattern of variation suggests that mean size is bigger than expected, in relation to temperature and salinity, when these factors have values close to the extremes of their range, normally found in nature, and to which adaptative mechanisms can evolve. Size is bigger at the salinity — temperature low - low and high - high combinations which are the most commonly found in the temperate environments.     

Dicarboxylic acid anhydrides as dissociating agents of protein-containing structures

Dissociation of protein-containing structures by modification of protein amino groups with dicarboxylic acid anhydrides is a mild procedure which, in some cases, offers advantages over treatment with alternative dissociating agents, such as urea, guanidine hydrochloride, detergents, high ionic strength, and extremes of pH: In addition to dissociating multimeric proteins and protein aggregates, dicarboxylic acid anhydrides are effective dissociating agents for membrane-bound proteins and nucleoprotein particles. With most dicarboxylic acid anhydrides reviewed, the introduced reagent residues can be eliminated under moderate acid conditions, which allows the purification of unmodified individual components, and the use of disassembly-reconstitution systems valuable for investigating the structural and functional roles played by the individual components of complex particles:Each reagent can be suitable for a particular purpose, depending on the required specificity of the modification and stability of the modified groups: The stability of the acylated amino groups ranges from the very stable succinylated amino groups to the very labile acylation obtained with dimethylmaleic anhydride: Between these extremes, the stability of the modified amino groups decreases stepwise in the following order: maleic, exo-cis-3,6-endoxo-⁴-tetrahydrophthalic, citraconic, and 3,4,5,6-tetrahydrophthalic anhydride. With respect to the selectivity of the produced modification, little or no modification of hydroxyamino acid and cysteine residues has been observed with dimethylmaleic, exo-cis-3,6-endoxo-⁴-tetrahydrophthalic, and 3,4,5,6-tetrahydrophthalic anhydrides: With the other reagents, the extent of modification of hydroxyamino acid residues increases in the order citraconic, maleic and succinic anhydride: Citraconic and maleic anhydrides can produce irreversible modification of cysteine residues, the reactivity of sulfhydryl groups being higher with maleic anhydride:     

Analysis of polyadenylated RNA from brain synaptosomes and mitochondria

We have isolated RNA from sheep brain synaptosomes and mitochondria separated by an aqueous two-phase system composed of dextran and poly(ethylene glycol). RNA was fractionated through oligo(dT)-cellulose columns and analyzed by electrophoresis through agarose slab gels containing methylmercuric hydroxide and stained with ethidium bromide. The electrophoretic patterns of the poly(A)-containing RNA fraction from synaptosomes and mitochondria are very similar although some high molecular weight RNA species, clearly visible in the synaptosomal fraction, are scarcely detected in the mitochondrial preparations. The electrophoretic analysis of a cleaner RNA preparation from digitonin-treated free mitochondria (mitoplasts) showed that all the poly (A)-RNA species of the synaptosomal preparation are also present in mitoplast. These results strongly suggest that all the discrete poly(A)-RNA species identified in brain synaptosomes are of mitochondrial origin.     

Ca2+ ionophores inhibit superoxide generation by chemotactic peptide in rabbit neutrophils and the correlation with intracellular calcium

Summary Effects of Ca²⁺ ionophores, A23187 and lasalocid, on superoxide anion generation by chemotactic peptide, N-formyl-methionyl-leucyl-phenylalanine methyl ester, in rabbit peritoneal exudate neutrophils were studied. The ionophores by themselves did not activate superoxide anion generation in these neutrophils. When preincubated with the cells for 2 min, both the ionophores inhibited superoxide generation induced by chemotactic peptide. The inhibition was present even in the absence of extracellular Ca²⁺ and the inhibition was better then. Lasalocid produces a dose-dependent chlortetracycline fluorescence decrease response in neutrophils loaded with chlortetracycline. This response is independent of extracellular Ca²⁺ concentration and is related to release of Ca²⁺ from intracellular storage sites. The dose-range at which lasalocid gives this response is same as the dose-range at which it causes inhibition of superoxide response. It may be concluded that the inhibition of superoxide generation by these ionophores is correlated to intracellular Ca²⁺ modulation.Abbreviations FMLP Formyl-Methionyl-Leucyl-Phenylalanine methyl ester     

Developmental expression of 44-kDa bone phosphoprotein (osteopontin) and bone γ-carboxyglutamic acid (Gla)-containing protein (osteocalcin) in calcifying tissues of rat

New aspects of the distribution and developmental appearance of the 44-kDa bone phosphoprotein (44K BPP, also called sialoprotein I or osteopontin) and bone gamma-carboxyglutamic acid (Gla)-containing protein (BGP, also called osteocalcin) during osteogenesis and dentinogenesis were investigated with immunocytochemical techniques using monospecific, affinity-purified polyclonal antibodies. Sections from newborn rat incisors and from various bone anlagen of newborn animals and fetuses were processed for detection of 44K BPP or BGP antigenicity. In addition, histochemical reactions for detection of alkaline phosphatase or calcium salts were performed on a number of the sections. The 44K BPP appears to be synthesized and secreted by chondrocytes only in the areas of cartilage-to-bone transition; these cells could participate indirectly in the process of bone formation by providing a suitable scaffold onto which primary marrow osteoblasts attach and spread. During osteogenesis, 44K BPP is found in bone-forming cells almost concomitantly with the appearance of alkaline phosphatase and before osteoid deposition, whereas BGP is still absent during early stages of mineralization. We hypothesize that this dramatic difference between the developmental appearance of 44K BPP and BGP reflects the delayed expression of the BGP gene relative to that of 44K BPP. In long-term cultures of bone marrow from adult mice, some fibroblastic cells expressed the 44K BPP phenotype; these cells could represent early osteogenic progenitor cells. Some experiments also suggested that, as with BGP, 44K BPP or an immunologically related protein is synthesized by some odontoblasts and secreted into predentin, prior to the onset of mineralization.     

Fermentation parameters of Peptostreptococcus productus on gaseous substrates (CO,H2/CO2)

Intrinsic growth and substrate uptake parameters were obtained for Peptostreptococcus productus, strain U-1, using carbon monoxide as the limiting substrate. A modified Monod model with substrate inhibition was used for modeling. In addition, a product yield of 0.25 mol acetate/mol CO and a cell yield of 0.034 g cells/g CO were obtained. While CO was found to be the primary substrate, P. productus is able to produce acetate from CO₂ and H₂, although this substrate could not sustain growth. Yeast extract was found to also be a growth substrate. A yield of 0.017 g cell/g yeast extract and a product yield of 0.14 g acetate/g yeast extract were obtained. In the presence of acetate, the maximum specific CO uptake rate was increased by 40% compared to the maximum without acetate present. Cell replication was inhibited at acetate concentrations of 30 g/l. Methionine was found to be an essential nutrient for growth and CO uptake by P. productus. A minimum amount of a complex medium such as yeast extract (0.01%) is, however, required.