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S L Snyder

Publications and source records attributed to S L Snyder.

At least 19 recordsLinked to original sources
PubMed2004

Characterization of the yeast tricalbins: membrane-bound multi-C2-domain proteins that form complexes involved in membrane trafficking.

In a survey of yeast genomic sequences encoding calcium- and phospholipid-binding C2 domains, three homologous genes were identified that encode proteins that each have three C2 domains and an apparent transmembrane domain near the N terminus. The name tricalbins is suggested for these proteins, corresponding to the open reading frames YOR086c (TCB1), YNL087w (TCB2), and YML072c (TCB3). An antiserum was raised against the C-terminal portion of tricalbin 2 and used on Western blots to demonstrate that the corresponding protein is expressed in yeast and appears as a high-molecular-weight band at 130 kDa with smaller fragments at 39 kDa and 46 kDa. A fusion protein consisting of full length tricalbin 2 fused to the green fluorescent protein was expressed in cells and found to traffic from the cell surface to intracellular vesicles near the vacuole. A two-hybrid interaction screen with the C-terminal portion of tricalbin 2 indicated that tricalbin 2 binds the C-terminal portions of tricalbins 1 and 3 suggesting that the tricalbins may form heterodimers in vivo. Tricalbin 2 also interacted with the activation domain of the pleiotropic drug resistance transcription factor Pdr1p. Combinatorial disruptions of the tricalbin genes revealed that tcb2 single mutants or tcb1, tcb3 double mutants have an altered vacuole morphology and are hypersensitive to cycloheximide. A screen for single-copy suppressors of the cycloheximide sensitivity of tricalbin mutants yielded RSP5, which encodes a C2-domain-containing, ubiquitin-conjugating ligase essential for receptor-mediated and fluid phase endocytosis. The results suggest that the tricalbins function as multimers in membrane-trafficking events and may provide insights into the roles of multi-C2-domain proteins, such as the synaptotagmins, in other organisms.

Amino Acid Motifs↗
PubMed1998

The copines, a novel class of C2 domain-containing, calcium-dependent, phospholipid-binding proteins conserved from Paramecium to humans.

In an attempt to identify proteins that might underlie membrane trafficking processes in ciliates, calcium-dependent, phospholipid-binding proteins were isolated from extracts of Paramecium tetraurelia. The major protein obtained, named copine, had a mass of 55 kDa, bound phosphatidylserine but not phosphatidylcholine at micromolar levels of calcium but not magnesium, and promoted lipid vesicle aggregation. The sequence of a 920-base pair partial cDNA revealed that copine is a novel protein that contains a C2 domain likely to be responsible for its membrane active properties. Paramecium was found to have two closely related copine genes, CPN1 and CPN2. Current sequence data bases indicate the presence of multiple copine homologs in green plants, nematodes, and humans. The full-length sequences reveal that copines consist of two C2 domains at the N terminus followed by a domain similar to the A domain that mediates interactions between integrins and extracellular ligands. A human homolog, copine I, was expressed in bacteria as a fusion protein with glutathione S-transferase. This recombinant protein exhibited calcium-dependent phospholipid binding properties similar to those of Paramecium copine. An antiserum raised against a fragment of human copine I was used to identify chromobindin 17, a secretory vesicle-binding protein, as a copine. This association with secretory vesicles, as well the general ability of copines to bind phospholipid bilayers in a calcium-dependent manner, suggests that these proteins may function in membrane trafficking.

Amino Acid Sequence↗
PubMed1996

Identification, localization, and functional implications of an abundant nematode annexin.

Cultures of the nematode C. elegans were examined for the presence of calcium-dependent, phospholipid-binding proteins of the annexin class. A single protein of apparent mass on SDS-polyacrylamide gels of 32 kD was isolated from soluble extracts of nematode cultures on the basis of its ability to bind to phospholipids in a calcium-dependent manner. After verification of the protein as an annexin by peptide sequencing, an antiserum to the protein was prepared and used to isolate a corresponding cDNA from an expression library in phage lambda gt11. The encoded protein, herein referred to as the nex-1 annexin, has a mass of 35 kD and is 36-42% identical in sequence to 10 known mammalian annexins. Several unique modifications were found in the portions of the sequence corresponding to calcium-binding sites. Possible phosphorylation sites in the NH2-terminal domain of the nematode annexin correspond to those of mammalian annexins. The gene for this annexin (nex-1) was physically mapped to chromosome III in the vicinity of the dpy-17 genetic marker. Two other annexin genes (nex-2 and nex-3) were also identified in chromosome III sequences reported by the nematode genomic sequencing project (Sulston, J., Z. Du, K. Thomas, R. Wilson, L. Hillier, R. Staden, N. Halloran, P. Green, J. Thierry-Mieg, L. Qiu, et al. 1992. Nature (Lond.). 356:37-41). The nex-1 annexin was localized in the nematode by immunofluorescence and by electron microscopy using immunogold labeling. The protein is associated with membrane systems of the secretory gland cells of the pharynx, with sites of cuticle formation in the grinder in the pharynx, with yolk granules in oocytes, with the uterine wall and vulva, and with membrane systems in the spermathecal valve. The presence of the annexin in association with the membranes of the spermathecal valve suggests a novel function of the protein in the folding and unfolding of these membranes as eggs pass through the valve. The localizations also indicate roles for the annexin corresponding to those proposed in mammalian systems in membrane trafficking, collagen deposition, and extracellular matrix formation.

Amino Acid Sequence↗
PubMed1994

Identification of the major chromaffin granule-binding protein, chromobindin A, as the cytosolic chaperonin CCT (chaperonin containing TCP-1).

Chromobindin A is a multisubunit complex ATPase that binds to chromaffin granule membranes in a calcium-dependent manner and requires ATP for release from the membrane (Martin, W. H., and Creutz, C. E. (1987) J. Biol. Chem. 262, 2803-2810). Here we report that the seven previously characterized subunits of chromobindin A cross react with antisera specific to subunits of CCT, the chaperonin containing TCP-1 (Kubota, H., Hynes, G., Carne, A., Ashworth, A., and Willison, K. (1994) Curr. Biol. 4, 89-99). The chromobindin A subunits previously called chromobindins 12, 13, 14, 15, 16, 18, and 19 cross-react specifically with subunits beta, delta, theta, alpha, zeta, xi, and gamma, respectively, of CCT. Additional similarities in subunit molecular weights, isoelectric points, and the morphologies of the two protein complexes as determined by electron microscopy support identification of chromobindin A as an adrenal medullary form of CCT. The chromobindin A/CCT complex was found to bind at least 7-fold more efficiently to affinity columns of chromaffin granule membranes than of adrenal medullary cytosol proteins, suggesting a specific interaction occurs between the complex and membrane components. The results indicate that the previously described characteristics of chromobindin A are likely to be relevant to the functions of CCT and suggest that the adrenal medullary form of CCT may play a role in the activities of secretory vesicle membranes.

Animals↗
PubMed1994

Nurses and clinical trials.

The article explores the Food and Drug Administration (FDA) Clinical Trial Process for new products in the United States and some unlimited roles nurses provide in this process. Discussions of recent regulatory issues concerning nurses caring for patients in clinical trial are discussed.

Clinical Trials as Topic↗
PubMed1994

SCID diagnosis of panic disorder in psychogenic seizure patients.

The authors administered the Structured Clinic Interview for DSM-III-R (SCID) to 20 outpatients with nonepileptic seizures documented by video-EEG. Fourteen (70%) had one or more non-somatoform DSM-III-R diagnoses. All 14 met criteria for panic disorder. Comorbid mood, psychotic, substance abuse, and eating disorders were also noted. Meticulous use of the SCID, with extensive follow-up, may have resulted in enhanced detection of panic disorder in patients who do not spontaneously report panic symptoms. Panic attacks may play a more important role in nonepileptic seizures than has been generally recognized, especially in outpatients with a chronic course of illness.

Adult↗
PubMed1993

New technique for improving hemostasis in aortic root replacement with composite graft.

By using both aortic annular and supraannular aortic wall tandem suture lines, we have, in our experience, markedly improved hemostasis in aortic root replacement with composite conduits of two types. Using this technique, 4 of 5 patients required 2 units or less of transfused red blood cells. Two patients did not receive transfusions. The technique consists of using interrupted mattress sutures to anchor the lower part of the valve sewing ring to the aortic annulus. The upper part of the sewing ring is then anchored with a running suture line to the cut edge of the supraannular aortic wall.

Adolescent↗
PubMed1992

Effects of the expression of mammalian annexins in yeast secretory mutants.

The hypothesis that calcium-dependent membrane-binding proteins of the annexin family can influence intracellular membrane trafficking was tested by expressing five mammalian annexins in wild-type yeast cells (Saccharomyces cerevisiae) and in 13 yeast secretory (sec) mutants. Expression of human synexin (annexin VII) inhibited the growth of sec2, sec4 and sec15 mutants at a semi-permissive temperature. These three sec mutants are defective in the final step in the secretory pathway, the process of exocytosis. The inhibition of growth correlated with reduced viability and increased accumulation of internal invertase in these mutants when expressing synexin. Bovine endonexin (annexin IV) partially suppressed the growth defect of a sec2 mutant incubated at a semi-permissive temperature. Human synexin, human lipocortin (annexin I), and murine p68 (annexin VI) reduced the lag time associated with adaptation of sec2 mutants to galactose-containing medium. These interactions suggest that the annexins may influence specific steps in membrane trafficking associated with cell growth, secretion and plasma membrane remodelling.

Annexins↗
PubMed1991

Calcium-dependent secretory vesicle-binding and lipid-binding proteins of Saccharomyces cerevisiae.

Yeast (Saccharomyces cerevisiae) cytosol was examined for the presence of calcium-dependent membrane- or lipid-binding proteins that might play fundamental roles in membrane-associated phenomena in stimulated cells. A complex group of proteins was isolated from late log phase cultures of yeast strain YP3 on the basis of calcium-dependent association with yeast secretory vesicles isolated from the temperature-sensitive sec6-4 secretory mutant. The masses of the major proteins in this group were 32, 35, 47, 51, 55, 60 and 120 kDa. A similar group of proteins was isolated by calcium-dependent association with bovine brain lipids enriched in the predominant acidic phospholipids of the yeast secretory vesicles. The 47 kDa protein was highly purified when commercial yeast cake was used as the source of yeast cytosol. The 32 kDa and 60 kDa proteins were demonstrated to reassociate with lipids at calcium concentrations of 100 microM or higher, while no association was promoted by 2 mM-magnesium. The 47 kDa protein could be removed from lipids by reducing the calcium concentration to between 1 and 32 microM. The sequences of peptides isolated from digests of several of these proteins indicate that they are novel proteins but are insufficient to judge the possible homology of these proteins with mammalian membrane-binding proteins. The sequence data may be adequate to permit isolation and modification of the corresponding genes in order to assess the possible function of this class of proteins in stimulated cells.

Amino Acid Sequence↗
PubMed1991

Preparing patients for cancer chemotherapy: effect of coping preparation and relaxation interventions.

Sixty cancer chemotherapy patients were randomly assigned to one of four treatments: (a) relaxation training with guided relaxation imagery (RT), (b) general coping preparation package (PREP), (c) both RT and PREP, or (d) routine clinic treatment only. All patients were assessed on self-report, nurse observation, family observation, and physiological measures and were followed for five sequential chemotherapy treatments. Results indicate that the PREP intervention increased patients' knowledge of the disease and its treatment, reduced anticipatory side effects, reduced negative affect, and improved general coping. RT patients showed some decrease in negative affect and vomiting, but not as great as in past studies. The data suggest that a relatively simple, one-session coping preparation intervention can reduce many different types of distress associated with cancer chemotherapy and may be more effective than often-used behavioral relaxation procedures.

Adaptation, Psychological↗
PubMed1991

Radioprotective properties of DNA methylation-disrupting agents.

5-Azacytidine and sodium butyrate, two DNA methylation-disrupting agents, were tested for radioprotective properties on V79A03 cells. Both compounds can activate genes not previously expressed (e.g. metallothionein). 5-Azacytidine treatment (3 microM, 24 h) caused a 50% decrease in the 5-methylcytosine content of V79A03 DNA whereas sodium butyrate treatment (1 mM, 24h) resulted in a 700% increase in 5-methylcytosine content. Additionally, 5-azacytidine treatment resulted in the increased survival of V79A03 cells, with treatment 24 h prior to exposure to gamma radiation providing a dose reduction factor of 1.8. Sodium butyrate treatment did not result in a significant increase in survival. These results indicate that the hypomethylation of genomic DNA prior to exposure to gamma radiation correlates with an increase in survival of V79A03 cells, possibly due to the activation of the enzymes involved in repair.

5-Methylcytosine↗
PubMed1989

18 S ribosomal RNA is degraded during ribosome maturation in irradiated HeLa cells.

The effects of ionizing radiation (137Cs) on processing of ribosomal RNA (rRNA) were studied by pulse-labeling HeLa S3 cells with [3H]uridine immediately prior to irradiation. The 45 S rRNA precursor, and its two major daughter species, 28 and 18 S rRNA, were separated by gel electrophoresis and the extent of radiolabel incorporation into each was determined at various times after irradiation. This approach permitted kinetic analysis of processing of the 45 S rRNA which had been predominantly synthesized (radiolabeled) prior to irradiation. Since they both derive from the same 45 S pre-rRNA transcript, 28 and 18 S rRNA are produced with a stoichiometry of 1:1, as observed in control cells in the present studies. However, within 1 h following 10 Gy an altered stoichiometry of 28 S:18 S rRNA was apparent, reaching 1.6:1 by 5-7 h following irradiation. This alteration was also observed following the higher dose of 20 Gy, but not following exposures of 5 Gy or less. The 18 S portion of the 45 S pre-rRNA is transcribed prior to the 28 S portion. Consequently, an increase in the 28 S/18 S ratio can only be due to degradation of the 18 S species during or after processing. This alteration may represent a response to radiation-induced growth arrest, by reducing the number of newly synthesized ribosomes that would otherwise be required for cell propagation.

Cesium Radioisotopes↗
PubMed1989

The effect of gamma radiation on DNA methylation.

The effect of 60Co gamma radiation on DNA methylation was studied in four cultured cell lines. In all cases a dose-dependent decrease in 5-methylcytosine was observed at 24, 48, and 72 h postexposure to 0.5-10 Gy. Nuclear DNA methyltransferase activity decreased while cytoplasmic activity increased in irradiated (10 Gy) V79A03 cells as compared to controls. No DNA demethylase activity was detected in the nuclei of control or irradiated V79A03 cells. Additionally, gamma radiation resulted in the differentiation of C-1300 N1E-115 cells, a mouse neuroblastoma line, in a dose- and time-dependent manner. These results are consistent with the hypothesis that (1) genes may be turned on following radiation via a mechanism involving hypomethylation of cytosine and (2) radiation-induced hypomethylation results from decreased intranuclear levels of DNA methyltransferase.

5-Methylcytosine↗
PubMed1989

Radiation-induced association of beta-glucuronidase with purified nuclei from irradiated MOLT-4 and HeLa cells.

Beta-glucuronidase, a lysosomal marker enzyme, associates with purified nuclei from HeLa and MOLT-4 cell lines in a radiation dose-dependent manner, up to 300 cGy in MOLT-4 cells, and 1000 cGy in HeLa cells. In MOLT-4 cells (200-cGy exposure), there is a significant increase in beta-glucuronidase activity detected in the nuclear fraction 24 h postirradiation with a maximum association occurring at 72 h. In HeLa cells (1000-cGy exposure), a significant association is first detected 24 h postirradiation with a maximum association at 48 h. The association is not the result of nonspecific contamination occurring during nuclei purification since nuclei from irradiated cells show no greater levels of plasma membrane marker and mitochondrial marker than controls. The nature of the association remains unclear, but activity is not removed by detergents used in the nuclei isolation procedure, and incubation of the nuclei with EDTA reverses the association only modestly. Exposure of nuclei from irradiated cells to anisotonic buffers also results in only a small decrease in beta-glucuronidase activity associated with the nuclei. These observations suggest that lysosomal hydrolases become intimately associated with the nuclei of irradiated cells.

Cell Line↗
PubMed1988

Pattern of repeating aromatic residues in synexin. Similarity to the cytoplasmic domain of synaptophysin.

Synexin was isolated from bovine liver by high resolution cation exchange chromatography and fragmented with cyanogen bromide or trypsin. Peptides were isolated and their amino acid sequences partially determined. Twenty percent of the synexin sequence was determined in one contiguous sequence of 61 residues and a nonoverlapping sequence of 20 residues. The sequence is characterized by a hexapeptide repeat of the form YPXXXX occurring eight times in series, with phenylalanine substituting for tyrosine in two positions. The intervening amino acids (X) are predominantly proline, glycine and alanine. This pattern of periodic aromatic residues suggests the presence of a novel secondary structure and is similar to repeats present in synaptophysin, gliadin and type II keratin.

Amino Acid Sequence↗
PubMed1988

Antibody responses to liposome-associated antigen.

The humoral antibody response of CAF1 mice to low doses (1-100 micrograms) of egg albumin (EA) encapsulated in or covalently bound to the surface of liposomes was studied for three routes of administration. The liposome immunoadjuvant effect observed was found to depend on the location of the antigen, either on the liposome surface or entrapped inside the liposome, and on the number of immunizations. Following a single immunization, the highest antibody titers were elicited with liposomes having EA conjugated to their surface, regardless of the route of administration. For multiple immunizations given i.v. or i.p. EA conjugated to the surface of liposomes was also superior to either free or liposome-encapsulated EA. However, the antibody response to EA bound to the surface of liposomes was not enhanced as compared to free EA following multiple subcutaneous immunizations.

Animals↗