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R Jahn

Publications and source records attributed to R Jahn.

At least 163 records · Page 9Linked to original sources
PubMed1990

Phospholipid binding by a synaptic vesicle protein homologous to the regulatory region of protein kinase C.

Neurotransmitters are released at synapses by the Ca2(+)-regulated exocytosis of synaptic vesicles, which are specialized secretory organelles that store high concentrations of neurotransmitters. The rapid Ca2(+)-triggered fusion of synaptic vesicles is presumably mediated by specific proteins that must interact with Ca2+ and the phospholipid bilayer. We now report that the cytoplasmic domain of p65, a synaptic vesicle-specific protein that binds calmodulin contains an internally repeated sequence that is homologous to the regulatory C2-region of protein kinase C (PKC). The cytoplasmic domain of recombinant p65 binds acidic phospholipids with a specificity indicating an interaction of p65 with the hydrophobic core as well as the headgroups of the phospholipids. The binding specificity resembles PKC, except that p65 also binds calmodulin, placing the C2-regions in a context of potential Ca2(+)-regulation that is different from PKC. This is a novel homology between a cellular protein and the regulatory domain of protein kinase C. The structure and properties of p65 suggest that it may have a role in mediating membrane interactions during synaptic vesicle exocytosis.

Amino Acid Sequence↗
PubMed1990

Energy dependence and functional reconstitution of the gamma-aminobutyric acid carrier from synaptic vesicles.

The energy dependence of gamma-aminobutyric acid (GABA) uptake was characterized in rat brain synaptic vesicles and in proteoliposomes reconstituted with a new procedure from vesicular detergent extracts. The proteoliposomes displayed high ATP-dependent GABA uptake activity with properties virtually identical to those of intact vesicles. GABA uptake was similar at chloride concentrations of 0 and 150 mM, i.e. conditions under which either the membrane potential (delta psi) or the pH difference (delta pH) predominates. Delta psi was gradually dissipated by increasing the concentration of SCN-. GABA uptake was reduced by 10 mM SCN-, showing less sensitivity to delta psi reduction than glutamate uptake but more than dopamine uptake. Dissipation of delta pH with NH+4 abolished GABA uptake at pH 7.3, whereas no significant inhibition occurred at pH 6.5. In contrast, dopamine uptake was inhibited more strongly, even at pH 6.5, and glutamate uptake was not reduced in either condition. We conclude that GABA uptake is driven by both components of the proton electrochemical gradient, delta pH and delta psi, and that this is different from the uptake of both dopamine and glutamate, which is more strongly dependent on delta pH and delta psi, respectively. Thus, our data suggest that GABA uptake is electrogenic and occurs in exchange for protons.

Adenosine Triphosphate↗
PubMed1990

Bacteriorhodopsin drives the glutamate transporter of synaptic vesicles after co-reconstitution.

Active accumulation of neurotransmitters by synaptic vesicles is an essential component of the synaptic transmission cycle. Isolated vesicles show energy-dependent uptake of several transmitters by processes which are apparently mediated by a proton electrochemical potential across the vesicle membrane. Although this energy gradient is probably generated by a proton ATPase, the functional separation of ATP cleavage and transmitter uptake activity has only been shown clearly for monoamine transport. We report here that the light-driven proton pump, bacteriorhodopsin, can replace the endogenous proton ATPase in proteoliposomes reconstituted from vesicular detergent extracts. The system shows light-dependent uptake of glutamate with properties very similar to those observed in intact vesicles, e.g. chloride dependence or stimulation by NH4+. Our experiments show that the proton pump and the glutamate transporter are separate entities and provide a powerful tool for further characterization of the glutamate carrier.

Amino Acid Transport System X-AG↗
PubMed1990

Immunohistochemical localization of synaptophysin (p38) in the pineal gland of the Mongolian gerbil (Meriones unguiculatus).

Synaptophysin (protein p38), a major integral membrane glycoprotein of small presynaptic vesicles, was localized immunohistochemically in semithin sections of the superficial pineal gland of the Mongolian gerbil (Meriones unguiculatus). Synaptophysin immunoreactivity could be detected in all pinealocytes, which were visualized with antibodies directed against neuron-specific enolase (NSE) in adjacent sections. No p38 immunoreactivity was discernible in the interstitial glial cells, which showed a heterogeneous pattern of immunostaining for the intermediate filament proteins glial fibrillary acidic protein (GFAP) and vimentin. Pinealocytes exhibited considerable intercellular differences in the densities of immunostaining. The various degrees of synaptophysin immunoreactivities in pinealocytes were not correlated with the densities of NSE immunostaining. Nerve terminals and varicosities displayed stronger immunoreactivities than pinealocytes. They were particularly numerous in the perivascular spaces. It is not clear whether this distribution indicates an innervation of pineal capillaries in addition to the functionally important innervation of pinealocytes. Several highly p38-positive dots of variable size were a conspicuous feature throughout the gland. By the consecutive semithin-thin section technique, they could be identified as processes of pinealocytes, filled with accumulations of small clear vesicles. Obviously, these vesicles represent the major site of synaptophysin immunoreactivity in pinealocytes. In the gerbil, similar vesicles have been ascribed a role in the secretory activity of the gland, and/or in the transport of calcium. The intercellular differences in the degrees of p38 immunostaining may, therefore, reflect different states of a specific cellular activity. The presence of synaptophysin in pinealocytes of the normal pineal, including the deep portions of the gland, emphasizes the paraneuronal character of these cells.

Animals↗
PubMed1990

Amino acid neurotransmission: spotlight on synaptic vesicles.

The vesicle hypothesis describing quantal release of neurotransmitter at the cholinergic neuromuscular junction was introduced in 1956. Since then, the concept of vesicular storage and release of acetylcholine has become firmly established and extended to include other synapses and neurotransmitters. However, for the amino acids, which are the major class of neurotransmitters in the mammalian CNS, there was no direct experimental evidence of the participation of synaptic vesicles in neurotransmission. This area of research has now moved out of the shadows and this article discusses recent findings which indicate that amino acid neurotransmitters are accumulated and stored by synaptic vesicles in presynaptic nerve endings.

Amino Acids↗
PubMed1990

rab3 is a small GTP-binding protein exclusively localized to synaptic vesicles.

rab3, a low molecular weight GTP-binding protein, is primarily expressed in brain, where it is present in soluble and membrane-bound forms. Membrane-bound rab3 in brain is exclusively localized on synaptic vesicles, the secretory organelles of the synapse that store and release neurotransmitters. rab3 is also expressed in endocrine tissues such as the adrenal medulla, where it is found together with other synaptic vesicle proteins on microvesicles distinct from chromaffin granules. The tight binding of rab3 to membranes correlates with hydrophobic modifications that are different in the membrane-bound and soluble forms of rab3. The results demonstrate the exclusive targeting of a small GTP-binding protein to secretory vesicles of a subset of the regulated pathway of secretion.

Amino Acid Sequence↗
PubMed1990

P29: a novel tyrosine-phosphorylated membrane protein present in small clear vesicles of neurons and endocrine cells.

A novel membrane protein from rat brain synaptic vesicles with an apparent 29,000 Mr (p29) was characterized. Using monospecific polyclonal antibodies, the distribution of p29 was studied in a variety of tissues by light and electron microscopy and immunoblot analysis. Within the nervous system, p29 was present in virtually all nerve terminals. It was selectively associated with small synaptic vesicles and a perinuclear region corresponding to the area of the Golgi complex. P29 was not detected in any other subcellular organelles including large dense-core vesicles. The distribution of p29 in various subcellular fractions from rat brain was very similar to that of synaptophysin and synaptobrevin. The highest enrichment occurred in purified small synaptic vesicles. Outside the nervous system, p29 was found only in endocrine cell types specialized for peptide hormone secretion. In these cells, p29 had a distribution very similar to that of synaptophysin. It was associated with microvesicles of heterogeneous size and shape that are primarily concentrated in the centrosomal-Golgi complex area. Secretory granules were mostly unlabeled, but their membrane occasionally contained small labeled evaginations. Immunoisolation of subcellular organelles from undifferentiated PC12 cells with antisynaptophysin antibodies led to a concomitant enrichment of p29, synaptobrevin, and synaptophysin, further supporting a colocalization of all three proteins. P29 has an isoelectric point of approximately 5.0 and is not N-glycosylated. It is an integral membrane protein and all antibody binding sites are exposed on the cytoplasmic side of the vesicles. Two monoclonal antibodies raised against p29 cross reacted with synaptophysin, indicating the presence of related epitopes. P29, like synaptophysin, was phosphorylated on tyrosine residues by endogenous tyrosine kinase activity in intact vesicles.

Animals↗
PubMed1990

Synaptophysin: a substrate for the protein tyrosine kinase pp60c-src in intact synaptic vesicles.

Expression of pp60c-src, the first well defined proto-oncogene product, is developmentally regulated and tissue-specific, with neuronal tissues displaying high amounts of the c-src encoded pp60c-src kinase activity. In the central nervous system pp60c-src is preferentially expressed in regions characterized by a high content of grey matter and elevated density of nerve terminals. In this study we show for the first time a direct interaction between pp60c-src and synaptophysin as a physiological target protein in neurons by demonstrating that endogenous pp60c-src is able to phosphorylate synaptophysin (p38). p38 is a major constituent of the synaptic vesicle membrane protein and is thought to play a key role in the exocytosis of small synaptic vesicles and possibly small clear vesicles in neuroendocrine cells.

Animals↗
PubMed1990

Synaptic vesicles: key organelles involved in neurotransmission.

1. This article summarizes some of the recent advances in the understanding of structural and functional properties of isolated small synaptic vesicles (SSV) from mammalian brain. 2. SSV contain a set of integral membrane proteins which are highly specific for this organelle and which occur on all SSV of the central and peripheral nervous system irrespective of their transmitter content. In contrast, these proteins are absent from the membrane of peptide-containing large dense-core vesicles indicating that the two types of organelle have a different membrane composition. The availability of antibodies for these proteins has allowed the evaluation of the purity of vesicle preparations which is instrumental for functional studies. 3. Recent advances in the study of neurotransmitter uptake have revealed that SSV contain specific carrier systems for glutamate and GABA. They are different from the transporters of the plasma membrane, and are dependent on the energy of a proton electrochemical gradient. The uptake of glutamate has been characterized in some detail and the mechanistic and physiological implications of these findings are discussed.

Animals↗
PubMed1990

[A case report of welder's lung].

The case of a 57-year-old man is reported, who worked as a plasma welder about 15 years. Chronic recurrent bronchitis was diagnosed 12 years after beginning of this work. The patient changed the place of his employment but the course of disease was uncommonly progressive. He died of right ventricular heart failure 10 years later. One year before death, the chronic obstructive pulmonary lesions were acknowledged as occupational disease. The post-mortem histological and chemical analysis of lung tissue yielded a mixed dust pneumoconiosis. These findings supported additionally the diagnosis of occupational disease.

Dust↗
PubMed1989

Characterization and partial purification of a chloride- and calcium-dependent glutamate-binding protein from rat brain.

A glutamate-binding protein was solubilized from rat brain synaptic plasma membranes using sodium cholate. Its properties were characterized after addition of exogenous phospholipids and formation of proteoliposomes. Glutamate binding was dependent on calcium and chloride ions with maximal binding at concentrations of 10(-5) M calcium and 10 mM chloride ions. The effects of the two ions were synergistic rather than additive. In addition, glutamate binding was not affected by inhibitors specific for N-methyl-D-aspartate and kainate receptor subtypes, but was inhibited by quisqualate (Ki = 50 microM) and DL-2-amino-4-phosphonobutyrate (Ki = 1.3 mM). Furthermore, binding was abolished by 100 microM 4,4'-diisothiocyanatostilbene-2,2'-disulfonic acid and 1 mM dithiothreitol. These properties resemble those of the chloride- and calcium-dependent binding site. Starting from the detergent extract, the glutamate-binding protein was purified 123-fold using fractionated ammonium sulfate precipitation, chromatography on hydroxyapatite and on DEAE-Sephacel as sequential purification steps. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of the purified protein fraction showed two major bands migrating with Mr values of 51,000 and 105,000. The properties of the partially purified binding protein were similar to those of the detergent extract. Glutamate binding to the partially purified protein is not due to a sequestration process or product binding to N-acetylated alpha-linked dipeptidase. Thus, the functional role of the binding protein remains to be established.

Animals↗
PubMed1989

Transmembrane topography and evolutionary conservation of synaptophysin.

Synaptophysin is the major integral membrane protein of small synaptic vesicles. Its primary structure deduced from rat and human complementary DNA sequences predicts that synaptophysin contains four transmembrane regions and a carboxyl-terminal domain having a novel repetitive structure. To elucidate the transmembrane organization of this protein in the synaptic vesicle, five antipeptide antibodies were raised. The site-specific antibodies were used to map the cognate sequences to the cytoplasmic or intravesicular side of the synaptic vesicle membrane by determining the susceptibility of the epitopes to proteolysis. The results confirm a topographic model for synaptophysin in which the protein spans the vesicle membrane four times, with both the amino and carboxyl terminus being cytoplasmic. In addition, the evolutionary conservation of the synaptophysin domains was addressed as a function of their membrane localization. To this end the primary structure of bovine synaptophysin was determined. Sequence comparisons between bovine, rat, and human synaptophysin revealed that only the intravesicular loops showed a significant number of amino acid substitutions (22%), while the transmembrane regions and cytoplasmic sequences were highly conserved (3% substitutions). These results depict synaptophysin as a protein with multiple membrane spanning regions whose functional site is likely to reside in highly conserved intramembranous and cytoplasmic sequences.

Amino Acid Sequence↗
PubMed1989

Synaptobrevin: an integral membrane protein of 18,000 daltons present in small synaptic vesicles of rat brain.

A protein with an apparent mol. wt of 18,000 daltons (synaptobrevin) was identified in synaptic vesicles from rat brain. Some of its properties were studied using monoclonal and polyclonal antibodies. Synaptobrevin is an integral membrane protein with an isoelectric point of approximately 6.6. During subcellular fractionation, synaptobrevin followed the distribution of small synaptic vesicles, with the highest enrichment in the purified vesicle fraction. Immunogold electron microscopy of subcellular particles revealed that synaptobrevin is localized in nerve endings where it is concentrated in the membranes of virtually all small synaptic vesicles. No significant labeling was observed on the membranes of peptide-containing large dense core vesicles. In agreement with these results, synaptobrevin immunoreactivity has a widespread distribution in nerve terminal-containing regions of the central and peripheral nervous system as shown by light microscopy immunocytochemistry. Outside the nervous system, synaptobrevin immunoreactivity was found in endocrine cells and cell lines (endocrine pancreas, adrenal medulla, PC12 cells, insulinoma cells) but not in other cell types, for example smooth muscle, skeletal muscle and exocrine pancreas. Thus, the distribution of synaptobrevin is similar to that of synaptophysin, a well-characterized membrane protein of small vesicles in neurons and endocrine cells.

Animals↗
PubMed1989

Synaptophysin is targeted to similar microvesicles in CHO and PC12 cells.

Synaptophysin, an integral membrane protein of small synaptic vesicles, was expressed by transfection in fibroblastic CHO-K1 cells. The properties and localization of synaptophysin were compared between transfected CHO-K1 cells and native neuroendocrine PC12 cells. Both cell types similarly glycosylate synaptophysin and sort it into indistinguishable microvesicles. These become labeled by endocytic markers and are primarily concentrated below the plasmalemma and at the area of the Golgi complex and the centrosomes. A small pool of synaptophysin is transiently found on the plasma membrane. In CHO-K1 cells synaptophysin co-localizes with transferrin that has been internalized by receptor-mediated endocytosis. These findings suggest that synaptophysin in transfected CHO-K1 cells and neuroendocrine PC12 cells is directed into a pathway of recycling microvesicles which, in CHO cells, is shown to coincide with that of the transferrin receptor. They further indicate that fibroblasts have the ability to sort a synaptic vesicle membrane protein. Our results suggest a pathway for the evolution of small synaptic vesicles from a constitutively recycling organelle which is normally present in all cells.

Adrenal Gland Neoplasms↗