Search PubMed⌕ Search

Biomedical subjects

R H Scheller

Publications and source records attributed to R H Scheller.

At least 217 records · Page 12Linked to original sources
PubMed1986

A neuropeptide precursor expressed in Aplysia neuron L5.

Aplysia abdominal ganglion neuron L5 is immunoreactive with an antiserum generated against the tetrapeptide Phe-Met-Arg-Phe-amide (FMRFamide); however, the specificity of this immune reagent is limited to the sequence Arg-Phe-amide. We isolated cDNA clones homologous to mRNAs specifically expressed in L5 and demonstrated that these clones do not hybridize to a previously characterized gene encoding FMRFamide. The nucleotide sequence of one of these clones, L5-67, does not encode any FMRFamide peptides but does reveal a Gly-Lys-Arg cleavage site following the amino acids Arg-Phe. This data predicts that neuron L5 expresses a peptide ending in Arg-Phe-amide, consistent with the FMRFamide immunoreactivity.

Amino Acid Sequence↗
PubMed1986

Egg-laying hormone genes of Aplysia: evolution of the ELH gene family.

Evolution of the egg-laying hormone (ELH) gene family was examined in the genus Aplysia using genomic Southern blotting, gene cloning, and immunocytochemical techniques to identify and characterize homologous sequences. Most of the species examined have fewer than the 4-5 ELH-related genes present in the A. californica genome (Mahon et al., 1985; Scheller et al., 1983). In A. parvula there are 2 ELH genes, and unlike A. californica, no sequences were found to encode the A or B peptides. The 2 A. parvula ELH genes share at least 90% DNA sequence homology, while the homology between the A. parvula and A. californica ELH genes is 71%. The structural organization of the A. parvula ELH precursor is quite similar to the ELH precursor of A. californica, with all but one of the potential proteolytic cleavage sites conserved. The overall amino acid homology between the A. parvula and A. californica ELH precursors is 66%; however, the alpha and beta bag cell peptides, as well as ELH, are more highly conserved, suggesting that these peptides have important physiological and behavioral roles within both Aplysia species. Immunocytochemical studies indicate that the A. parvula ELH genes are expressed in 2 bag-cell-like clusters of about 40 neurons each in the abdominal ganglion. There does not appear to be an atrial gland in A. parvula; however, ELH-immunoreactive peripheral neurons and their processes are observed along the perimeter of the large hermaphroditic duct.

Animals↗
PubMed1985

Expression of the L11 neuropeptide gene in the Aplysia central nervous system.

Neuron L11 in the abdominal ganglion of Aplysia californica is thought to be both cholinergic and peptidergic. In previous studies, we isolated a cDNA clone encoding the precursor for an L11 secreted protein(s) by differentially screening an abdominal ganglion cDNA library. We now report the isolation of genomic clones encoding the L11 cDNA sequences. Analysis of these clones reveals that the gene is present in a single copy per haploid genome. RNA blotting and cDNA cloning demonstrate that the L11 gene is expressed not only in the abdominal ganglion but in the head ganglia as well. To define the positions of cells expressing this gene and to follow their processes, we raised antibodies to synthetic peptides defined by the cDNA sequence. Histochemistry revealed about 100 neurons containing immunoreactive material. These cells arborize in the neuropil and are distributed throughout the central nervous system, representing about 0.5% of the Aplysia central neurons. In addition, cells in the abdominal ganglion send processes to the mantle floor at the base of the gill via the genital and branchial nerves. Our data suggest that this network of cells expresses the single L11 peptide gene.

Animals↗
PubMed1985

Low molecular weight proteins of Aplysia neurosecretory cells.

The proteins of identified cells from the Aplysia californica central nervous system were labeled with radioactive amino acids and fractionated on SDS acrylamide gels containing 6 M urea. Most of the large cells contain prominent, cell-specific protein products in the molecular weight range between 3 and 30 KD. The molecular weights of the largest specific prevalent protein products are in good agreement with the predicted molecular weights of precursors as determined from an analysis of cDNA clones homologous to mRNA's specifically expressed in several of these neurons. Biologically active peptides have been found in many of these cells. These data, and other indirect evidence suggests that the synthesis of a large amount of a particular protein in this molecular weight range is indicative of the synthesis of a neurosecretory product. We conclude that most, if not all, large neurons in the Aplysia central nervous system are peptidergic.

Animals↗
PubMed1985

Aplysia neurons express a gene encoding multiple FMRFamide neuropeptides.

The neuroactive peptide Phe-Met-Arg-Phe-NH2 (FMRF-amide) has a variety of effects on both mammalian and invertebrate tissues; moreover, FMRFamide-like immunoreactivity is found throughout the animal kingdom. Here we describe the isolation and characterization of a cDNA clone from an Aplysia abdominal ganglion cDNA library that encodes a precursor protein that may give rise to as many as 19 individual FMRFamide peptides. Nearly all of the FMRF sequences are flanked on the amino terminus by Lys-Arg residues and on the carboxy terminus by Gly-Lys residues, suggesting that the single lysine residues function to signal cleavage by processing enzymes. The gene is present in a single copy per haploid genome and gives rise to multiple transcripts, at least some of which appear to arise through alternate RNA splicing. Immunohistochemical analysis suggests that the peptide is present in many neurons throughout the Aplysia nervous system and that these neurons send processes to a variety of different tissues.

Amino Acid Sequence↗
PubMed1985

The small cardioactive peptides A and B of Aplysia are derived from a common precursor molecule.

We have identified cells in the central nervous system of the marine mollusc Aplysia that react with antibody raised against the small cardioactive peptide B (SCPB). Antisera to this neuropeptide stained a subset of central neurons that include the large identified buccal neurons, B1 and B2. The distribution of SCP-containing neurons was used in a strategy to isolate a cDNA clone encoding the precursor protein for the peptide. RNA from neurons B1 and B2 and from cells that did not stain with SCPB antisera was used to direct the synthesis of radiolabeled cDNA probes. A cDNA clone complimentary to mRNA specifically expressed in the B1 and B2 cells was isolated by differentially screening a buccal cDNA library with these probes. The cloned cDNA segment is 1394 nucleotides in length and contains a 408-base-pair open reading frame. The predicted precursor protein is composed of 136 amino acids and has a characteristic hydrophobic leader sequence. The sizes of the precursor protein with and without this leader sequence agree with in vivo and in vitro labeling studies. The amino acid sequences for SCPB and a related peptide, SCPA, are present and are flanked by known proteolytic processing sites.

Amino Acid Sequence↗
PubMed1985

Neuropeptides in identified Aplysia neurons.

Extensive electrophysiological experiments on Aplysia neurons have resulted in an understanding of simple behaviors in terms of the activities of a single identified neurons. Beginning with the work of Kupfermann & Kandel, neuropeptides in Aplysia have become increasingly implicated as chemical agents that control or affect behavior. Several neuropeptides have been isolated and characterized; recently, the genes that code for several of these neuropeptides have been isolated. Studies of neuropeptide gene expression and the behaviors affected thereby have been bridged in the egg-laying hormone neuroendocrine system. The role of polyproteins in coordinating complex, fixed-action patterns is beginning to emerge. The continued investigation of this neuroendocrine system, and the other cell-specific polyproteins that have been characterized more recently, promises to yield further insights into the roles of neuropeptides in governing behavior.

Animals↗
PubMed1985

Biochemical and immunocytological localization of molluscan small cardioactive peptides in the nervous system of Aplysia californica.

High pressure liquid chromatography (HPLC) followed by bioassay on isolated snail hearts were used to locate two related peptides, termed small cardioactive peptides A and B (SCPA and SCPB) in each of the central ganglia of Aplysia. The peptides are most concentrated in the buccal ganglia, the ganglia involved in the control of feeding movements. Immunocytology with antisera raised to conjugated SCPB stained three groups of neurons in the buccal ganglia. One group consisted of relatively small neurons that were tightly clustered. The second group was comprised of larger neurons that were more scattered. The third group was made up of several neurons including the two largest in the ganglia, identified cells B1 and B2. B1 and B2 and other neurons in this group innervate the gut by way of the esophageal nerve. HPLC-bioassay of single, individually dissected B1 or B2 neurons demonstrated that the two peptides are present in a single cell. For B2, but not B1, choline injected into the cell body was converted to the conventional transmitter, acetylcholine. This indicates that, in addition to the two peptides, B2 also contains choline acetyltransferase, and raises the possibility that acetylcholine and the SCPs may act as co-transmitters in B2. Strong immunocytological staining of fibers and varicosities was observed in the neuropilar region of the cerebral, pleural, pedal, and abdominal ganglia. In addition to the buccal ganglia, immunoreactive neurons were observed in all of the other central ganglia. The high concentration of the SCPs and the relatively large number of immunoreactive neurons in the buccal ganglion suggest a particularly important role of these peptides specifically in feeding behavior. However, the widespread occurrence of the SCPs in fibers and neuronal cell bodies throughout the nervous system suggests that these peptides also may have additional behavioral functions in Aplysia.

Aplysia↗
PubMed1985

Structure and expression of the egg-laying hormone gene family in Aplysia.

Transcription of the egg-laying hormone (ELH) gene family was examined by characterizing homologous cDNA clones from abdominal ganglion and atrial gland cDNA libraries. All cDNAs contain an exon that spans the coding region (exon III) and one or two additional exons. The tissue-specific expression of the ELH gene family was confirmed by the observation that exon III encodes the ELH precursor protein in the bag cell transcripts and either the A or B precursor proteins in the atrial gland transcripts. The cDNA clones also contain 5' untranslated exons not present in the previously isolated genomic clones. One type of transcript has a 40-base pair segment, designated exon I, contiguous with exon III. A second type of transcript has an additional 149 base pairs of DNA, designated exon II, located between exons I and III. Several genomic clones containing exons I and II were isolated. DNA sequence analysis reveals that exons I and II are directly linked and that they are separated from exon III by an intervening sequence of at least 5 kilobases (kb). Consensus sequences for a putative promotor region and also for RNA splicing and polyadenylation were identified. From this work we can describe a prototype ELH gene complete with identified sequences necessary for the proper initiation of transcription and the subsequent processing of the transcript.

Aplysia↗
PubMed1984

Neuropeptides: mediators of behavior in Aplysia.

The Aplysia neuroendocrine system is a particularly advantageous model for cellular and molecular studies because of the relatively small number and large size of its component neurons. Recombinant DNA techniques have been used to isolate the genes that encode the precursors of peptides expressed in identified neurons of known function. The organization and developmental expression of these genes have been examined in detail. Several of the genes encode precursors of multiple biologically active peptides that are expressed in cells which also contain classical transmitters. These studies, as well as immunohistochemical studies and the use of intracellular recording and voltage clamp techniques are the first steps toward revealing the mechanisms by which neuropeptides govern simple behaviors.

Animals↗
PubMed1984

A cDNA clone encoding neuropeptides isolated from Aplysia neuron L11.

Single nerve cells can use more than one substance as extracellular chemical messengers. Classical transmitters have been shown to coexist in the same neuron and possibly even in the same vesicle as neuroactive peptides. Furthermore, multiple neuroactive peptides, which are thought to be coreleased, are often encoded in the same precursor assuring stoichiometric synthesis. The precise organization of multiple message systems and the physiological significance of the coexistence is poorly understood. The abdominal ganglion of the gastropod mollusc Aplysia contains a number of identified neurons that are cotransmitter candidates. One such cell, L11, is cholinergic and probably also uses biologically active peptides. Differential screening with labeled cDNA was used to isolate cDNA clones expressed specifically in the bag cells and abdominal ganglion neurons L11 or R15. Analysis of an L11-specific clone suggests that it encodes a 14.7-kDa protein that is the precursor for the secreted peptides. The poly(A)+ RNA transcript is approximately equal to 1.2 kilobases and there are 1-3 copies of this gene in the Aplysia haploid genome.

Amino Acid Sequence↗
PubMed1984

Two introns define functional domains of a neuropeptide precursor in Aplysia.

Biologically active peptides are synthesized as parts of precursor proteins which are proteolytically processed to generate active molecules. The structure of the gene encoding peptides expressed in Aplysia neurons R3-14 suggests that two intervening sequences split the transcript into functional domains. The first exon encodes the 5' untranslated region, the second exon the signal sequence and the bulk of the negative charge of the precursor protein while the third exon encodes the remainder of the precursor and the 3' untranslated region.

Animals↗
PubMed1984

Antibodies to synthetic peptides defined by cDNA cloning reveal a network of peptidergic neurons in Aplysia.

We previously isolated and characterized a cDNA clone specifically expressed in neurons R3 to R8 and R14 of the Aplysia abdominal ganglion (Nambu, J.R., R. Taussig, A.C. Mahon, and R.H. Scheller (1983) Cell 35: 47-56). The cDNA nucleotide sequence and the inferred protein amino acid sequence suggest that this gene encodes the precursor for neuroactive peptides used by these cells. Peptides corresponding to three regions of the precursor were synthesized, coupled to a protein carrier, and used to generate antibodies. These antibodies stain a set of cell bodies, R3 to R14, and their processes in the abdominal ganglion; no other cells in the nervous system or the periphery are immunoreactive. R3 to R14 send numerous fine immunoreactive processes into the vascularized sheath that surrounds the ganglion. Each of these cells also has a large axon which exits the ganglion via the branchial nerve and terminates on the heart. In addition, R14 is anatomically distinct from R3 to R13 in that it sends additional immunoreactive processes to the vasculature near the ganglion. Immunoreactive processes and varicosities were observed on the efferent vein of the gill, the abdominal ganglion artery, and the anterior aorta. These data are consistent with previous studies suggesting that one or more neuropeptides released from R3 to R14 function as modulators of cardiovascular physiology.

Abdomen↗
PubMed1983

In situ hybridization to study the origin and fate of identified neurons.

Egg-laying behavior in Aplysia is mediated by a set of peptides, including egg-laying hormone (ELH), which are released by a cluster of identified neurons, the bag cells. A family of neuropeptide genes which includes the gene encoding ELH along with two additional genes encoding the A and B peptides thought to initiate the egg-laying process has been isolated and their nucleotide sequence has been determined. In situ hybridization and immunofluorescence was used to explore the origin and distribution of the neurons that express this family of genes. The ELH genes are expressed, not only in the bag cells, but in an extensive system of neurons distributed in four of the five ganglia of the central nervous system. The genes for ELH are expressed in these cells early in the animal's life cycle. As a result, it was possible to use in situ hybridization to trace the cells expressing ELH to their site of origin. The cells originate outside the central nervous system in the ectoderm of the body wall and appear to migrate to their final locations within the central nervous system by crawling along strands of connective tissue.

Age Factors↗
PubMed1983

Gene isolation with cDNA probes from identified Aplysia neurons: neuropeptide modulators of cardiovascular physiology.

The Aplysia abdominal ganglion neurons, R3-R14, modulate cardiovascular activity. In vitro translations of poly(A)+ RNA from these cells suggest that they contain a prevalent mRNA encoding a 14 kd protein. Utilizing differential screening techniques with 32P-labeled cDNA synthesized from the poly(A)+ RNA of identified neurons, we isolated the corresponding gene. The Aplysia haploid genome contains a single copy of this sequence, which is interrupted by two large introns and spans approximately 7 kb of genomic DNA. The R3-R14 neurons specifically express this gene, resulting in the synthesis of a 1.25 kb mRNA not found in other abdominal ganglion cells or in the head ganglia. The gene was shown to encode a 13.5 kd precursor, which is proteolytically cleaved into at least three peptides with molecular weights of 5.0, 3.3, and 1.3 kd. These peptides and glycine are thought to act as chemical messengers in the central nervous system and peripherally.

Amino Acid Sequence↗
PubMed1983

A single gene encodes multiple neuropeptides mediating a stereotyped behavior.

Egg laying in Aplysia is characterized by a stereotyped behavioral array which is mediated by several neuroactive peptides. We have sequenced two genes encoding the A and B peptides thought to initiate the egg-laying process, as well as a gene encoding egg-laying hormone (ELH) which directly mediates the behavioral array. The three genes share 90% sequence homology and are representatives of a small multigene family. Each gene encodes a protein precursor in which the active peptides are flanked by internal cleavage sites providing the potential to generate multiple small peptides. Each of the three genes consists of sequences homologous to A or B peptide as well as ELH. Although these genes share significant nucleotide homology, they have diverged such that different member genes express functionally related but nonoverlapping sets of neuroactive peptides in different tissues.

Animals↗
PubMed1983

The molecular basis of a neuroendocrine fixed action pattern: egg laying in Aplysia.

We describe a gene family that encodes the proteins controlling the egg-laying behavior of Aplysia. The family evolved by duplication and divergence from a common ancestral gene. The ELH gene family is expressed in the atrial gland, in the bag cells, and in a small network of neurons in the central ganglia. The bag cells and the atrial gland express distinct members of the family that encode different precursor proteins. These contain one or more biologically active peptides that can be released by proteolytic cleavage. The bag cell precursor releases several peptides that have multiple sites of action and may generate different components of the egg-laying behavior. Coordination of the full stereotyped behavior is achieved by simultaneous release of these peptides from the bag cell processes.

Amino Acid Sequence↗