Biomedical subjects

S W Rogers

Publications and source records attributed to S W Rogers.

At least 37 records · Page 2Linked to original sources

PubMed1998

Glutamate receptor GluR1 expression is altered selectively by chronic audiogenic seizures in the Frings mouse brain.

The audiogenic seizure-susceptible mouse, Frings, is genetically susceptible to sound-induced seizures and provides a reliable model of reflex epilepsy that lasts throughout the life span of the animal. We used immunohistochemistry to examine if the expression of the non-N-methyl-D-aspartate glutamate receptor (GluR) subunits GluR1, GluR2, or GluR3 were altered subsequent to multiple seizures. Following a regimen of one seizure per day for 3 weeks, GluR1 immunoreactivity, but not GluR2 or GluR3, was substantially elevated in the outer shell of the nucleus accumbens in 21 of 31 chronically seized Frings mice. No other brain regions such as the hippocampus exhibited any qualitative changes in expression of these subunits. In 9 of the 21 Frings mice exhibiting increased GluR1, but in none of the controls, bilateral structural lesions were observed in the lateral hypothalamus. These results support a model where highly localized changes in the expression of GluR1 occur in response to repeated audiogenic seizure.

Acoustic Stimulation↗

PubMed1997

Identification of amino acids in the glutamate receptor, GluR3, important for antibody-binding and receptor-specific activation.

We reported (Twyman, R. E., Gahring, L. C., Speiss, J., and Rogers, S. W. (1995) Neuron 14, 755-762) that antibodies to a subregion of the glutamate receptor (GluR) subunit GluR3 termed GluR3B (amino acids 372-395), act as highly specific GluR agonists. In this study we produced additional rabbit anti-GluR3B-specific antibodies, ranked them according to their ability to function as GluR agonists and characterized the immunoreactivity using deletion and alanine substitution mutagenesis. These anti-GluR3B antibodies bound to a subset of the residues in GluR3B (amino acids 372-386), of which glutamate 375, valine 378, proline 379, and phenylalanine (Phe) 380 were preferred. The level of GluR activation correlated with the binding of antibody to Phe-380, which suggests that immunoreactivity directed toward Phe-380 is an index for the anti-GluR agonist potential. Since the identity of this residue varies between respective GluR subunits, this suggested that this residue may be important for imparting antibody subunit specificity. To test this possibility, the alanine in GluR1 was converted to a phenylalanine, which extended the subunit specificity from GluR3 to the modified GluR1. We conclude that antibody contacts with key residues in the GluR3B region define a novel GluR subunit-specific agonist binding site and impart subunit-specific immunoreactivity.

Alanine↗

PubMed1997

Interleukin-1alpha in the brain is induced by audiogenic seizure.

We examined the expression of the sleep-inducing cytokine interleukin-1alpha (IL-1alpha) in the brains of audiogenic seizure-susceptible mice subsequent to the induction of sound-induced seizure. Animal models of epilepsy often require lesioning or trauma that may nonspecifically alter IL-1alpha expression. To avoid this, we employed the Frings mouse strain; a model of auditory-evoked reflex epilepsy. Frings mice were exposed to a high-intensity sound stimulus to induce a tonic extension seizure, and the expression of IL-1alpha transcripts in different brain regions was measured thereafter. Compared to control animals, IL-1alpha transcripts were elevated 6 to 8 h postseizure in the hypothalamus, but not hippocampus, by a dexamethasone-sensitive pathway. Similar results were obtained from the genetically distinct DBA/2J audiogenic seizure-susceptible mouse strain. These findings demonstrate that the expression of IL-1alpha is altered following generalized seizure activity, induced by noninvasive sensory stimulation, in a brain-region-specific manner.

Acoustic Stimulation↗

PubMed1997

Monoclonal antibodies to barley aleurain and homologs from other plants.

Barley aleurain is contained within a specific type of vacuole characterized by acidic pH and the presence of other hydrolytic enzymes. The aleurain-containing vacuole is distinct from protein storage vacuoles, and anti-aleurain antibodies serve as markers for this organelle in barley cells. Aleurain is a unique type of cysteine protease, and other plant species have genes for homologs whose sequences are highly conserved, but little is known about these homologs at the protein level. Seven monoclonal antibodies to barley aleurain were isolated, which bind to and define aleurain homologues in Arabidopsis, Petunia, and tobacco cell extracts. Interestingly, in addition to 29-32 kDa aleurain homologs, Petunia extracts contain a protein of approximately 50 kDa and tobacco extracts a protein of approximately 40 kDa that are recognized by multiple different monoclonal antibodies, indicating an unexpected diversity to the aleurain protein family. Among the group of antibodies are some that efficiently immunoprecipitate metabolically labeled aleurain from barley cell extracts, and some that efficiently label aleurain in immunofluorescence assays using root tip cells. These antibodies should be useful for plant cell biologists who study vacuole biogenesis and function and sorting of proteins to specific vacuolar compartments, in barley as well as other plant species.

Amino Acid Sequence↗

PubMed1997

Molecular cloning and further characterization of a probable plant vacuolar sorting receptor.

BP-80 is a type I integral membrane protein abundant in pea (Pisum sativum) clathrin-coated vesicles (CCVs) that binds with high affinity to vacuole-targeting determinants containing asparagine-proline-isoleucine-arginine. Here we present results from cDNA cloning and studies of its intracellular localization. Its sequence and sequences of homologs from Arabidopsis, rice (Oryza sativa), and maize (Zea mays) define a novel family of proteins unique to plants that is highly conserved in both monocotyledons and dicotyledons. The BP-80 protein is present in dilated ends of Golgi cisternae and in "prevacuoles," which are small vacuoles separate from but capable of fusing with lytic vacuoles. Its cytoplasmic tail contains a Tyr-X-X-hydrophobic residue motif associated with transmembrane proteins incorporated into CCVs. When transiently expressed in tobacco (Nicotiana tabacum) suspension-culture protoplasts, a truncated form lacking transmembrane and cytoplasmic domains was secreted. These results, coupled with previous studies of ligand-binding specificity and pH dependence, strongly support our hypothesis that BP-80 is a vacuolar sorting receptor that trafficks in CCVs between Golgi and a newly described prevacuolar compartment.

Amino Acid Sequence↗

PubMed1997

Autoantibodies to glutamate receptor subunit GluR2 in nonfamilial olivopontocerebellar degeneration.

We describe a 63-year-old man with a 5-year history of progressive sporadic olivopontocerebellar atrophy (OPCA) who exhibits high serum titers of IgM autoantibodies to the neuronal glutamate receptor subunit GluR2. Immunohistochemistry revealed intense staining of mouse cerebellar Purkinje cells and cells in the pontine nuclei and olivary complex. Glutamate receptor currents were activated in a subset of cultured mouse neurons by an anti-GluR2 IgM fraction, and they were blocked by the competitive AMPA-type glutamate receptor antagonist CNQX and by a synthetic peptide to a specific epitope region of GluR2 (AA 369-393). The patient was treated with nine courses of plasmapheresis with little improvement of symptomatology. However, IgM titers to GluR2 decreased approximately 8-fold and the serum functional activity decreased proportionally. These findings may suggest a role for autoimmunity to glutamate receptors in the pathophysiology of certain forms of progressive nervous system degeneration.

Animals↗

PubMed1996

Neuronal nicotinic receptor expression in sensory neurons of the rat trigeminal ganglion: demonstration of alpha3beta4, a novel subtype in the mammalian nervous system.

The identification of a family of neuronal nicotinic receptor subunit genes establishes the potential for multiple subtypes with diverse physiological functions. Virtually all of the high affinity nicotinic receptors measured to date in the rodent CNS are composed of alpha4 and beta2 subunits only. However, the demonstration of other subunit transcripts in a variety of central and peripheral nervous tissues suggests a greater degree of receptor subtype heterogeneity than so far has been elucidated. The purpose of the present studies was to determine at the mRNA and protein levels which neuronal nicotinic receptor subunits are expressed by sensory neurons of the rat trigeminal ganglion and in what combinations these gene products associate to form neuronal nicotinic receptor subtypes in this tissue. Radioreceptor binding analysis indicated that in the adult rat trigeminal ganglion there exist at least two nicotinic receptor binding sites with differing affinities for [3H]-epibatidine. In situ hybridization histochemical studies revealed the existence of mRNA encoding the alpha3, alpha4, alpha5, beta2, and beta4 subunits, but not the alpha2 subunit. Immunoprecipitation with subunit-specific antisera demonstrated that each of the subunits present in the ganglion at the mRNA level is a constituent of nicotinic receptors capable of binding 3H-epibatidine. Various applications of these approaches yielded strong evidence that, in addition to alpha4beta2, which is thought to be the predominant neuronal nicotinic receptor subtype in the rodent CNS, trigeminal sensory neurons express as the principal subtype alpha3beta4, which has not been demonstrated previously in mammalian nervous tissue.

Animals↗

PubMed1996

Nicotinic receptor subunits alpha 3, alpha 4, and beta 2 and high affinity nicotine binding sites are expressed by P19 embryonal cells.

Controlled exposure to retinoic acid (RA) induces the murine embryonal carcinoma cell line P19S18O1A1 (P19) to differentiate into a variety of cell types. One of the cell types exhibits neuronal-like morphology and expresses neuronal markers including neurofilament proteins, glutamate receptors, and the cholinergic enzymes choline acetyl-transferase and acetylcholinesterase. In this study we use Northern blot analysis, double-label immunocytochemistry, and single cell RNA analysis using polymerase chain reaction to show that RA-treated P19 cells with neuronal-like morphology also express neuronal nicotinic acetylcholine receptor (nAChR) subunits alpha 3, alpha 4, and beta 2. Greater than 80% of RA-treated P19 cells with a neuronal-like phenotype express nAChR alpha 4 subunit transcripts and both alpha 4 and beta 2 protein. The RA-induced expression of alpha 3 transcripts accounts for a comparably small number of nAChR-containing cells (< 20%) of which half coexpress alpha 4 transcripts. Expression of high-levels of alpha 4 RNA is dependent upon both cell-cell contact and RA exposure. The appearance of nAChR subunits also coincides with RA-induced expression of high affinity [3H]-nicotine binding receptors. The P19 cell line offers an inducible neuronal cell system to study mammalian neuronal nicotinic receptor expression and the development of high affinity nicotinic binding sites similar to those expressed in the mammalian central nervous system.

Animals↗

PubMed1996

Kainic acid induced excitotoxicity and cfos expression in fibroblasts transfected with glutamate receptor subunit, GluR1.

Glutamate receptors participate in the majority of fast excitatory neurotransmission in the mammalian brain. Excessive excitation of these receptors has been linked to neuronal dysfunction and death through a process termed excitotoxicity. In this study we demonstrate that transfection of a single non-NMDA glutamate receptor subunit, GluR1, into cultured fibroblasts is sufficient to confer kainic acid mediated excitotoxicity similar to that seen in neuronal cells. Death of transfected cells requires at least 24 h of continuous exposure to kainic acid and can be blocked with a glutamate receptor antagonist. Also, the induction of protooncogene cfos transcripts occurs 30 min following kainic acid administration, and Fos protein accumulated in the nucleus within 90 min. These observations suggest that the signaling system(s) required to initiate gene expression and kainic acid excitotoxicity from a neuronal ionotropic receptor to the nucleus is present in these nonneuronal cells. Finally, antibodies prepared to amino acids 185-449 of GluR1 are demonstrated to be useful for fluorescence-activated sorting of live cells transfected with a GluR1 expression vector. This supports the conclusion that this region of the protein is located extracellularly.

Animals↗

PubMed1996

The role of autoimmunity to glutamate receptors in neurological disease.

Understanding and experimentally approaching the processes that underlie the origin and progression of many severe neurological disorders presents a challenge to both clinical and basic researchers. We have found that the origin of some neurological diseases, including a rare form of childhood epilepsy and a neurodegenerative disease associated with peripheral cancers, may be related to production of physiologically active autoantibodies that are directed towards excitatory ionotropic glutamate receptors of the brain. This suggests that some neurological diseases may result from dysfunction of the immune system.

Autoantibodies↗

PubMed1996

PEST sequences and regulation by proteolysis.

In 1986, we proposed that polypeptide sequences enriched in proline (P), glutamic acid (E), serine (S) and threonine (T) target proteins for rapid destruction. For much of the past decade there were only sporadic experimental tests of the hypothesis. This situation changed markedly during the past two years with a number of papers providing strong evidence that PEST regions do, in fact, serve as proteolytic signals. Here, we briefly review the properties of PEST regions and some interesting examples of the conditional nature of such signals. Most of the article, however, focuses on recent experimental support for the hypothesis and on mechanisms responsible for the rapid degradation of proteins that contain PEST regions.

Amino Acid Sequence↗

PubMed1995

Glutamate receptor antibodies activate a subset of receptors and reveal an agonist binding site.

Two rabbits immunized with a portion of glutamate receptor (GluR) subunit GluR3 (amino acids 245-457) exhibited seizure-like behaviors, suggesting that antibodies to GluR3 may modulate neuronal excitability. Using whole-cell recording, rabbit GluR3 antisera were tested on cultured fetal mouse cortical neurons. In a subset of kainate-responsive neurons, miniperfusion of antisera and IgG evoked currents that were blocked by CNQX. Immunoreactivity to synthetic peptides prepared to subregions GluR3A (amino acids 245-274) and GluR3B (amino acids 372-395) was present in both rabbit sera. Peptide GluR3B, but not GluR3A, specifically blocked antisera- and IgG-evoked currents. Similar receptor activation and anti-GluR3 reactivity was present in sera from patients with active Rasmussen's encephalitis, an intractable pediatric epilepsy. Thus, antibodies to GluR3 define a region involved in agonist binding and specific receptor activation. These data suggest that antibodies to neuronal receptors can function as agonists and that autoantibodies to GluRs may be highly specific neurotoxicants in some neurological diseases.

6-Cyano-7-nitroquinoxaline-2,3-dione↗

PubMed1995

Comparison of the effects of N6-methyldeoxyadenosine and N5-methyldeoxycytosine on transcription from nuclear gene promoters in barley.

Methylation of deoxycytosine residues in plant nuclear DNA at CpG dinucleotides is generally assumed to suppress transcription, while deoxyadenosine methylation on recombinant reporter gene constructs transiently introduced into plant cells increases expression by uncharacterized mechanisms. A particle bombardment transient expression system was used in intact barley aleurone layers to quantitate the effects of CpG and A methylation on transcription from well-characterized hormone-regulated alpha-amylase promoters. Methylation of C in all CpG pairs had little effect on transcription. In contrast, the presence of methylated A residues in the sequence GATC scattered in the reporter plasmid outside of the promoter increased transcription two- to fivefold, regardless of the strength of the promoter, and proper hormonal regulation of transcription was maintained. The methyl-dA effect was similar when the amount of reporter construct DNA used was varied over a 20-fold range, beginning with an amount that gave only a small increment of expression above background. Similar enhancing effects for methyl-dA were seen with the CaMV 35S, maize Adh1, and maize ubiquitin promoters, though the magnitude varied for each individual promoter. Methyl-dA did not detectably affect plasmid DNA stability in aleurone cells in transient expression experiments because the enhancing effect of methyl-dA on expression was the same regardless of whether the assay was performed at 12 h or 40 h. Several proteins in wheat germ nuclear extracts bound preferentially to methylated DNA as assessed by gel retardation assays; one showed preferential binding to methyl-dA rather than methyl-CpG residues. The data indicate that the presence of methyl-dA in the vicinity of active promoters enhances transcription of nuclear genes in barley in a manner independent of the strength of the promoter. This effect may be mediated by a methyl-dA-binding protein.

5-Methylcytosine↗

PubMed1995

Distribution of the excitatory amino acid receptor subunits GluR2(4) in monkey hippocampus and colocalization with subunits GluR5-7 and NMDAR1.

Ionotropic excitatory amino acid (EAA) receptors are divided pharmacologically into three categories termed NMDA, AMPA/kainate, and high affinity kainate receptors. Each of these receptor subtypes is composed of a specific subset of subunits termed GluR1-4 (AMPA/kainate), GluR5-7, KA1-2 (high affinity kainate), and NMDAR1, 2 A-D (NMDA). Although colocalization of NMDA and non-NMDA receptors has been previously demonstrated electrophysiologically in rat, comprehensive analyses of subunit specific colocalization patterns have not been possible until the advent of appropriate antibodies. The present study investigates such immunocytochemical colocalization of several EAA receptor subunits within individual cells as well as dendritic spines in the monkey hippocampus. Double-label immunohistochemical experiments using antibodies which are specific for GluR2(4), GluR5-7, and NMDAR1 demonstrated that virtually all projection neurons in each subfield of the hippocampus contain subunits from the AMPA/kainate, kainate, and NMDA receptor families. In addition, confocal microscopy has demonstrated that individual spines may contain subunits representative of multiple EAA receptor families. Furthermore, detailed regional, cellular, and ultrastructural distribution patterns of the EAA receptor subunits GluR2 and GluR4 in monkey hippocampus are presented based on the use of a monoclonal antibody (mAb), 3A11, which was generated against the putative extracellular N-terminal domain of GluR2. Since this antibody recognizes only GluR2 in Western blots, and GluR2 as well as GluR4 in fixed transiently transfected cells, it has been designated anti-GluR2(4). Immunocytochemical labeling with mAb 3A11 revealed pyramidal cell somata and dendrites in each field of the hippocampus, as well as granule cells and polymorphic hilar cells in the dentate gyrus. Small cells with the morphologic characteristics of astroglia were also immunolabeled for GluR2(4) within the alveus and fimbria. Immunoreactivity at the ultrastructural level was localized to postsynaptic densities on dendritic spines and shafts and within the somatodendritic cytoplasm in all major hippocampal regions, as well as in a subset of dentate granule cell axons within the mossy fiber projection.

Aging↗

PubMed1995

Autoantibodies to neuronal glutamate receptors in patients with paraneoplastic neurodegenerative syndrome enhance receptor activation.

BACKGROUND: Paraneoplastic syndromes are "remote" complications of cancer characterized clinically by neurological disease. The sera and cerebrospinal fluid (CSF) from patients with paraneoplastic neurological syndromes (PNS) frequently contain autoantibodies to ill-defined neuronal antigens. We report here that neuronal glutamate receptors are targets for autoantibodies found in the serum from some patients with well-characterized PNS. MATERIALS AND METHODS: We have analyzed the serum from seven patients with well-characterized PNS for the presence of autoreactive antibodies to non-NMDA glutamate receptor subunits. Autoantibodies were assessed using Western blot, immunohistochemistry, and immunocytochemistry. Whole-cell electrophysiological recordings were used to examine the effect of antibodies on glutamate receptors expressed by cortical neurons in culture. RESULTS: Six of seven patients' serum contained autoantibodies to the non-NMDA glutamate receptor (GluR) subunits GluR1, GluR4, and/or GluR5/6. No patient had autoantibodies to GluR2, and only one patient exhibited weak immunoreactivity to GluR3. Electrophysiological analysis demonstrated that the serum from four of the six GluR-antibody-positive patients enhanced glutamate-elicited currents on cultured cortical neurons but had no effect on receptor function alone. Enhancement of glutamate-elicited currents was also produced by affinity-purified antibody to GluR5. CONCLUSIONS: The occurrence of autoantibodies to specific neuronal neurotransmitter subunits in the sera of patients with PNS and the ability of these autoantibodies to modulate glutaminergic receptor function suggest that some paraneoplastic neurological injury could result from glutamate-mediated excitotoxicity.

6-Cyano-7-nitroquinoxaline-2,3-dione↗

PubMed1994

Autoantibodies to glutamate receptor GluR3 in Rasmussen's encephalitis.

Rasmussen's encephalitis is a progressive childhood disease of unknown cause characterized by severe epilepsy, hemiplegia, dementia, and inflammation of the brain. During efforts to raise antibodies to recombinant glutamate receptors (GluRs), behaviors typical of seizures and histopathologic features mimicking Rasmussen's encephalitis were found in two rabbits immunized with GluR3 protein. A correlation was found between the presence of Rasmussen's encephalitis and serum antibodies to GluR3 detected by protein immunoblot analysis and by immunoreactivity to transfected cells expressing GluR3. Repeated plasma exchanges in one seriously ill child transiently reduced serum titers of GluR3 antibodies, decreased seizure frequency, and improved neurologic function. Thus, GluR3 is an autoantigen in Rasmussen's encephalitis, and an autoimmune process may underlie this disease.

Animals↗

PubMed1994

KEKE motifs. Proposed roles in protein-protein association and presentation of peptides by MHC class I receptors.

A stretch of 28 'alternating' lysine (K) and glutamate (E) residues is found in an activator of the multicatalytic protease. Such 'KEKE sequences' are also present in subunits of the multicatalytic protease, in subunits of the 26S protease and in a variety of chaperonins. We propose that KEKE regions promote association between protein complexes. Furthermore, they may contribute to the selection of peptides presented on MHC Class I receptors.

Amino Acid Sequence↗

PubMed1994

Selective RNA editing and subunit assembly of native glutamate receptors.

RNA editing and subunit assembly of ionotropic glutamate receptors (GluRs) were examined in an oligodendrocyte progenitor cell line, CG4, which expresses GluR2-GluR4, GluR6, GluR7, KA1, and KA2. AMPA-evoked currents rapidly desensitize, whereas kainate-evoked currents contain a steady-state component with a nearly linear current-voltage relation and a fast desensitizing component that is inwardly rectifying. The Q/R site is edited > 95% to the arginine codon in GluR2(Q607) mRNA, and < 5% in GluR6(Q621) mRNA. Immunoprecipitation experiments demonstrate that GluR6 and/or GluR7 subunits assemble with KA2, but not with GluR2-GluR4. These results indicate that oligodendrocyte progenitor cells selectively edit and assemble glutamate receptors into at least two functionally and structurally distinct heteromeric channels.

Animals↗