PubMed Health⌕ Search

Biomedical subjects

R H Scheller

Publications and source records attributed to R H Scheller.

At least 181 records · Page 10Linked to original sources
PubMed1990

Dale's hypothesis revisited: different neuropeptides derived from a common prohormone are targeted to different processes.

In the bag cells of Aplysia californica, the prohormone of egg-laying hormone is processed by means of endoproteolytic cleavage into two sets of peptides. The amino-terminal region of the prohormone gives rise to the bag cell peptides (alpha, beta, and gamma). These serve an autocrine function; they are autoexcitatory on the bag cells and also act locally to alter the firing patterns of neurons in the abdominal ganglion. The carboxyl-terminal portion of the prohormone gives rise to the egg-laying hormone. This peptide acts as a hormone on nearby neurons and by means of the circulation on peripheral tissues to bring about egg-laying. We have previously reported that the first cleavage of the prohormone, which occurs in the trans-Golgi network, results in two intermediates that are sorted into distinct vesicle classes prior to further processing. Here we show that these distinct vesicles are localized to separate processes, thus spatially segregating autocrine and hormonal release sites. The findings of segregation indicate that neurons need not always release the same set of chemical messengers from all of their endings.

Animals↗
PubMed1990

Sorting within the regulated secretory pathway occurs in the trans-Golgi network.

Bioactive peptides cleaved from the egg-laying hormone precursor in the bag cell neurons of Aplysia are sorted into distinct dense core vesicle classes (DCVs). Bag cell prohormone processing can be divided into two stages, an initial cleavage occurring in a late Golgi compartment, which is not blocked by monensin, and later cleavages that occur within DCVs and are blocked by monensin. Prohormone intermediates are sorted in the trans-Golgi network. The large soma-specific DCVs turn over, while the small DCVs are transported to processes for regulated release. Thus, protein trafficking differentially regulates the levels and localization of multiple biologically active peptides derived from a common prohormone.

Animals↗
PubMed1990

Organization and expression of the Drosophila FMRFamide-related prohormone gene.

The Drosophila FMRFamide gene is comprised of two exons separated by a 2.8-kb intervening sequence. The first exon consists of the 5' untranslated region and is spliced to the initiator methionine codon. The second exon encodes the prohormone and the 3' untranslated region of the mRNA. The promoter region contains a TATA box 30 nucleotides upstream from a consensus transcription start site. Immunohistochemical studies using antibodies generated against synthetic peptides and in situ hybridization histochemistry define a set of about 40 neurons in the brain and ventral ganglia that express the prohormone gene.

Amino Acid Sequence↗
PubMed1990

A unique neurofilament from Torpedo electric lobe: sequence, expression, and localization analysis.

A set of cDNA clones encoding a protein highly homologous to the mammalian middle-size class of neurofilaments (NF-M) was characterized. The amino acid similarity between the Torpedo and rat NF-M approaches 90% in the amino-terminal "rod-like" domain and is significantly lower in the carboxy-terminal tail. The Torpedo protein contains 13 tandem repeats of a unique six amino acid core, containing a Lys-Ser-Lys putative phosphorylation site. Surprisingly, the 3' untranslated region contains stretches of 80-90% nucleic acid homology with the mammalian, but not with the chicken sequences. This homology is greater than much of the coding region, suggesting that the 3' untranslated region of the message has an important functional role, perhaps governing RNA stability or localization. This Torpedo NF-M mRNA is expressed specifically in the electric lobe and was not detected in other tissues, including brain and spinal cord. A polyclonal antibody generated against a fusion protein synthesized in E. coli detects a 150-kDa protein in the electric lobe and organ, as well as a small amount of material in the brain. Cytochemical studies reveal immunoreactivity in electromotor neuron axons and terminals. Specific expression of neurofilament genes in subsets of central neurons may be important in determining the morphology and functional characteristics of specific neuronal subtypes.

Amino Acid Sequence↗
PubMed1990

Subcellular fractionation of prohormone processing products in the bag cell neurons.

Multiple biologically active peptides arising from a common prohormone are sorted into distinct classes of dense core vesicles within the bag cell neurons of Aplysia californica. In this study, pulse-chase analysis, combined with subcellular fractionation on Percoll gradients, are used to define the location of the prohormone processing events within the secretory pathway. Initial cleavage of the prohormone occurs in a light cellular compartment associated with the Golgi apparatus. The amino-terminal processing intermediate then accumulates in a denser compartment containing small dense cores enclosed in membranous sacs, as well as larger immature vesicles. After 4 h, amino-terminal products are found primarily in a much denser compartment which consists of large and small dense core vesicles. These large and small vesicles can be separated from each other using Percoll gradient centrifugation and are found to be enriched in amino- and carboxy-terminal products, respectively. Lastly, membrane association experiments suggest differential binding to membranes, or integral membrane proteins, as a possible mechanism for sorting of amino- and carboxy-terminal products.

Animals↗
PubMed1990

Multiple GTP-binding proteins from cholinergic synaptic vesicles.

Cholinergic synaptic vesicles purified from the electric organ of the marine ray, Discopyge ommata, contain 2 different size classes of GTP-binding proteins: one or more with an apparent molecular weight (MW) between 37 and 41 kDa, and 3 major and at least 2 minor proteins with MW between 20 and 29 kDa. These GTP-binding proteins were detectable using the alpha 32P-GTP overlay technique and covalent modification with bacterial toxins. The higher MW GTP-binding proteins are ADP-ribosylated by pertussis toxin and 2 of the lower MW GTP-binding proteins are sensitive to botulinum toxin.

Adenosine Diphosphate Ribose↗
PubMed1990

Distinct patterns of expression of two VAMP genes within the rat brain.

VAMPs are synaptic vesicle-specific proteins composed of a carboxy-terminal hydrophobic membrane anchor and an approximately 100 amino acid domain oriented towards the cytoplasm. In rat, two 77% homologous VAMP genes are expressed in the CNS. To precisely localize the neurons expressing these 2 forms of VAMP, we have used RNA blotting and in situ hybridization histochemistry with RNA probes specific for the 3' untranslated regions of the transcripts. These experiments revealed that the 2 genes are expressed in distict, but slightly overlapping, patterns in the rat brain. VAMP-1 expression is localized to a limited number of nuclei, particularly those involved in modulating somatomotor functions, while VAMP-2 expression is more ubiquitous, being found in nuclei associated with autonomic, sensory, and integrative roles. These data suggest that the specific structural features of individual VAMPs may play an important role in synaptic vesicle metabolism.

Animals↗
PubMed1989

A dense core vesicle protein is restricted to the cortex of granules in the exocrine atrial gland of Aplysia california.

We have generated a monoclonal antibody (mAb) 5E10 which recognizes an antigen localized to dense core vesicles (DCVs) in the atrial gland of Aplysia californica. mAb5E10 immunoprecipitates an abundant 57-kDa glycoprotein (atrial gland granule-specific antigen, AGSA) which is a soluble component of atrial gland DCVs. Electron microscopy reveals that AGSA immunoreactivity is restricted to the region between the dense core, which contains neuropeptide immunoreactivity, and the membrane of atrial gland DCVs. AGSA was purified by immunoaffinity chromatography, and the amino acid sequences of both N-terminal and internal cyanogen bromide fragments were determined. This information was used to isolate a 2.8-kilobase cDNA which encodes a 47-kDa protein. The predicted amino acid sequence contains the micro-sequenced peptides, an N-terminal hydrophobic signal sequence, and four N-linked glycosylation sites, but does not contain any significant homologies to database sequences. Northern blots and light level immunocytochemistry demonstrate that the AGSA gene is specifically expressed in the atrial gland. The identification of a protein localized to the cortex of DCVs suggests that this region has a specialized role in the function of these vesicles.

Amino Acid Sequence↗
PubMed1989

A bag cell neuron-specific antigen localizes to a subset of dense core vesicles in Aplysia californica.

The bag cell neurons of Aplysia govern egg-laying through the release of a number of bioactive peptides which are processed from a common precursor. Immunoelectron microscopic studies suggest that sorting at the trans-Golgi segregates peptides from the amino terminal and carboxy terminal of the precursor into distinct classes of dense-cored vesicles (DCVs). Here we identify a novel bag cell-specific antigen (4F6 antigen) using monoclonal antibodies (MAbs). Immunoprecipitations and Western blots demonstrate that the MAb4F6 specifically recognizes a protein of 80 kDa and does not react with the egg-laying hormone precursor, processing intermediates or final products. The 4F6 antigen is localized in a subset of DCVs which also contain peptides derived from the amino terminus of the precursor. These results further demonstrate the complexity of vesicular sorting in the bag cells and also identify a novel tissue specific antigen localized to DCVs.

Animals↗
PubMed1989

Two vesicle-associated membrane protein genes are differentially expressed in the rat central nervous system.

Vesicle-associated membrane protein (VAMP) 1 is a 120-amino acid protein which co-purifies with cholinergic synaptic vesicles from the marine ray Torpedo californica. We used the Torpedo gene to isolate two independent classes of VAMP cDNA clones from rat brain. Nucleotide sequence analysis of the cDNAs predicts proteins which are 84 and 75% homologous to Torpedo VAMP-1. The amino-terminal 24-28 amino acid residues which comprise the proline-rich head are only about 50% homologous between the different VAMPs, yet the proline-rich character is maintained. The 69 amino acids which comprise the hydrophilic core are highly homologous to Torpedo VAMP-1, with only 2 amino acid substitutions in rat VAMP-1 and 6 in rat VAMP-2. The carboxyl-terminal 23 amino acids of all of the VAMP proteins maintain the hydrophobic character necessary to serve as a membrane anchor. Both VAMP transcripts are expressed differentially in the rat central nervous system. Whereas VAMP-2 is more highly expressed in the whole brain, VAMP-1 is expressed at a higher level in the spinal cord.

Amino Acid Sequence↗
PubMed1989

Large dense cored vesicles are enriched in neuropeptide processing intermediates in the Aplysia bag cells.

The bag cell neurons in the marine snail Aplysia synthesize large amounts of the egg-laying hormone (ELH) prohormone. The ELH precursor is proteolytically processed into 9 peptides making this a useful system for studying prohormone processing and the sorting of proteins destined for the secretory pathway. The peptides derived from the ELH prohormone are differentially packaged into four distinct classes of dense cored vesicles (DVCs). Dense cored vesicles in the large class are greater than 250 nm in diameter, contain the 6 peptides derived from the aminoterminus of the prohormone and are localized to the cell soma and not the neuronal processes. Here we demonstrate that the large DCVs are enriched in prohormone processing intermediates. In addition, many of the large DCVs do not contain acid phosphatase activity suggesting they are an organelle distinct from the lysosomes and that different classes of DCVs may subserve unique functions within the secretory pathway.

Animals↗
PubMed1989

VAT-1: an abundant membrane protein from Torpedo cholinergic synaptic vesicles.

Expression screening was used to isolate cDNA clones encoding a synaptic vesicle membrane protein, VAT-1, which is specifically expressed in the electric lobe of marine rays. The predicted protein has a molecular weight of 41,572 daltons and contains several hydrophobic regions. An antibody raised against a fusion protein synthesized in E. coli recognizes an abundant 42 kd protein that copurifies largely with synaptic vesicles. Trypsin digestion of intact and lysed vesicles as well as membrane extractions suggests that VAT-1 is an integral membrane protein. The VAT-1 RNA is localized to the electromotor nucleus, and the fusion protein antibody stains the electric organ, demonstrating that the protein is transported to nerve terminals. These studies define a novel synaptic vesicle protein that is likely to play a central role in the functions mediated by specific classes of synaptic vesicles.

Acetylcholine↗
PubMed1989

Isolation and characterization of two homologous cDNA clones from Torpedo electromotor neurons.

Two homologous cDNA clones were isolated from a Torpedo california electric lobe lambda gt11 expression library using a polyclonal antiserum directed against proteins associated with synaptic vesicles. Northern blotting reveals an 8- to 9-kb transcript in the electric lobe and the spinal cord, but not in the brain or other non-neuronal tissues. Antibodies generated against a fusion protein synthesized in Escherichia coli reacted with a 85- to 90-kD species in the neurons of the electric lobe. The immunoreactivity is associated with microsomal membranes and can be extracted readily with high salt. Immunohistochemical studies demonstrated a sparse punctate staining pattern in the cell body which colocalized with a subpopulation of post-Golgi vesicles.

Amino Acid Sequence↗
PubMed1989

Modulation of ionic currents in Aplysia motor neuron B15 by serotonin, neuropeptides, and second messengers.

Both 5-HT and the 9 amino acid neuropeptide SCPb modulate 3 ionic currents in B15, enhancing a voltage-dependent inward sodium current, decreasing an outward potassium current and increasing an inward rectifying potassium current. In contrast, FMRFamide decreases a voltage-dependent inward sodium current and increases an outward potassium current. We have also investigated the roles of several second-messenger systems that may be mediating the effects of these modulators. Bath application of membrane permeable analogs of cAMP enhance the voltage-dependent inward sodium current and both 5-HT and SCPb increase cAMP levels in B15, suggesting that cAMP may be mediating part of the observed effects of these transmitters on B15. Experiments with phorbol ester, a protein kinase inhibitor, and a phospholipase inhibitor suggest that the phospholipase C/protein kinase C cascade may decrease an outward potassium current. Thus, 5-HT and SCPb may activate multiple second-messenger systems to modulate 3 ionic currents in B15. Additional studies suggest that a cascade involving arachidonic acid may be involved in mediating part of the FMRFamide responses in B15. These studies are beginning to define molecular mechanisms whereby a neuron differentially modulates multiple ionic currents in response to distinct chemical messengers.

Animals↗