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C S Copeland

Publications and source records attributed to C S Copeland.

13 recordsLinked to original sources
PubMed1999

Spectacle-mounted prisms as an aid in face-down positioning after vitrectomy surgery.

PURPOSE: To report a modification in the design of bed spectacles to provide an aid for patients who undergo face-down positioning after vitrectomy with gas-fluid exchange and postoperative gas tamponade. METHODS: New equipment design. Bed spectacles that use prisms to move the image 90 degrees around the horizontal meridian were modified by rotating the prisms in the frame 180 degrees and adding nosepads and cable temples. The spectacles were given to 12 patients for use during postoperative face-down positioning. A questionnaire was used to evaluate the helpfulness of the spectacles during face-down positioning. RESULTS: Patients ranked the spectacles as "helpful" to "very helpful" with a mean and SD of 8.3+/-1.6 on a scale of 1 to 10, where 1 was "a hindrance" and 10 was "very helpful." Patients wore the spectacles an average of 5.7+/-4.5 hours a day (range, 1.5 to 15 hours). CONCLUSION: Modified bed spectacles appear to be a portable aid to patients who undergo face-down positioning while sitting, standing, or prone.

Gases↗
PubMed1995

Social interaction effects on restrained eating.

The eating behavior of college women scoring within either the above-average or middle ranges on the Restraint Scale was compared following brief social encounters with male students or other female students, or a no-encounter control situation. The eating of average restraint women was significantly depressed following interaction with a partner whom the subjects considered attractive. For high restraint women a nonsignificant tendency towards disinhibition of eating following interaction with an attractive other occurred. Personality ratings by partners indicated that high restraint women presented a distinct social persona to male, but not female, strangers in the brief experimental interaction.

Feeding Behavior↗
PubMed1995

Nuclear pore complex clustering and nuclear accumulation of poly(A)+ RNA associated with mutation of the Saccharomyces cerevisiae RAT2/NUP120 gene.

To identify genes involved in the export of messenger RNA from the nucleus to the cytoplasm, we used an in situ hybridization assay to screen temperature-sensitive strains of Saccharomyces cerevisiae. This identified those which accumulated poly(A)+ RNA in their nuclei when shifted to the non-permissive temperature of 37 degrees C. We describe here the properties of yeast strains carrying mutations in the RAT2 gene (RAT - ribonucleic acid trafficking) and the cloning of the RAT2 gene. Only a low percentage of cells carrying the rat2-1 allele showed nuclear accumulation of poly(A)+ RNA when cultured at 15 degrees or 23 degrees C, but within 4 h of a shift to the nonpermissive temperature of 37 degrees C, poly(A)+ RNA accumulated within the nuclei of approximately 80% of cells. No defect was seen in the nuclear import of a reporter protein bearing a nuclear localization signal. Nuclear pore complexes (NPCs) are distributed relatively evenly around the nuclear envelope in wild-type cells. In cells carrying either the rat2-1 or rat2-2 allele, NPCs were clustered together into one or a few regions of the nuclear envelope. This clustering was a constitutive property of mutant cells. NPCs remained clustered in crude nuclei isolated from mutant cells, indicating that these clusters are not able to redistribute around the nuclear envelope when nuclei are separated from cytoplasmic components. Electron microscopy revealed that these clusters were frequently found in a protuberance of the nuclear envelope and were often located close to the spindle pole body. The RAT2 gene encodes a 120-kD protein without similarity to other known proteins. It was essential for growth only at 37 degrees C, but the growth defect at high temperature could be suppressed by growth of mutant cells in the presence of high osmolarity media containing 1.0 M sorbitol or 0.9 M NaCl. The phenotypes seen in cells carrying a disruption of the RAT2 gene were very similar to those seen with the rat2-1 and rat2-2 alleles. Epitope tagging was used to show that Rat2p is located at the nuclear periphery and co-localizes with yeast NPC proteins recognized by the RL1 monoclonal antibody. The rat2-1 allele was synthetically lethal with both the rat3-1/nup133-1 and rat7-1/nup159-1 alleles. These results indicate that the product of this gene is a nucleoporin which we refer to as Rat2p/Nup120p.

Alleles↗
PubMed1995

Mutation or deletion of the Saccharomyces cerevisiae RAT3/NUP133 gene causes temperature-dependent nuclear accumulation of poly(A)+ RNA and constitutive clustering of nuclear pore complexes.

To identify genes whose products play potential roles in the nucleocytoplasmic export of messenger RNA, we isolated temperature-sensitive strains of Saccharomyces cerevisiae and examined them by fluorescent in situ hybridization. With the use of a digoxigen-tagged oligo-(dT)50 probe, we identified those that showed nuclear accumulation of poly(A)+ RNA when cells were shifted to the nonpermissive temperature. We describe here the properties of yeast strains bearing the rat3-1 mutation (RAT-ribonucleic acid trafficking) and the cloning of the RAT3 gene. When cultured at the permissive temperature of 23 degrees C, fewer than 10% of cells carrying the rat3-1 allele showed nuclear accumulation of poly(A)+ RNA, whereas approximately 70% showed nuclear accumulation of poly(A)+ RNA, whereas approximately 70% showed nuclear accumulation of poly(A)+ RNA after a shift to 37 degrees C for 4 h. In wild-type cells, nuclear pore complexes (NPCs) are distributed relatively evenly around the nuclear envelope. Both indirect immunofluorescence analysis and electron microscopy of rat3-1 cells indicated that NPCs were clustered into one or a few regions of the NE in mutant cells. Similar NPC clustering was seen in mutant cells cultured at temperatures between 15 degrees C and 37 degrees C. The RAT3 gene encodes an 1157-amino acid protein without similarity to other known proteins. It is essential for growth only at 37 degrees C. Cells carrying a disruption of the RAT3 gene were very similar to cells carrying the original rat3-1 mutation; they showed temperature-dependent nuclear accumulation of poly(A)+ RNA and exhibited constitutive clustering of NPCs. Epitope tagging of Rat3p demonstrated that it is located at the nuclear periphery and co-localizes with nuclear pore proteins recognized by the RL1 monoclonal antibody. We refer to this nucleoporin as Rat3p/Nup133p.

Amino Acid Sequence↗
PubMed1993

Nuclear pore complex antigens delineate nuclear envelope dynamics in vegetative and conjugating Saccharomyces cerevisiae.

In the yeast Saccharomyces cerevisiae, the nucleus undergoes dramatic shape changes during mitosis and mating. We have studied nuclear envelope dynamics during the processes of mitosis and conjugation using nuclear pore complexes as a marker for the nuclear envelope in wild-type cells and several cell-division-cycle (cdc) mutants. Three monoclonal antibodies are described that recognize nuclear pore complex-related antigens in S. cerevisiae. One of these antibodies, RL1, has been extensively characterized by Gerace and colleagues and recognizes nuclear pore complexes in mammalian and amphibian cells. By indirect immunofluorescence of yeast cells, all three antibodies yield a discontinuous nuclear rim stain. All three react with multiple nuclear-enriched proteins in immunoblots, including the nucleoporin protein encoded by the NSP1 gene. When the antibodies were used in immunofluorescence experiments on mating cells, the nuclear pore complex staining pattern proved to be a sensitive indicator of nuclear fusion. Nuclei with closely apposed spindle pole bodies and unfused nuclear envelopes could be readily distinguished. Marked shape changes were observed in nuclei during fusion and segregation of the diploid nucleus into the zygotic bud. In cdc14 and cdc15 mutants that arrest late in mitosis, the elongated nuclear envelope extension that stretches between daughter nuclei during telophase was preserved. In cytokinesis-defective mutants (cdc3, cdc10, cdc11 and cdc12), the elongated nuclear envelope was usually resolved into two daughter nuclei in the absence of cytokinesis. These results indicate that nuclear envelope division is mechanically distinguishable from chromosome segregation, nucleolar segregation and cytokinesis.

Amino Acid Sequence↗
PubMed1992

The NUF1 gene encodes an essential coiled-coil related protein that is a potential component of the yeast nucleoskeleton.

In an attempt to identify structural components of the yeast nucleus, subcellular fractions of yeast nuclei were prepared and used as immunogens to generate complex polyclonal antibodies. One such serum was used to screen a yeast genomic lambda gt11 expression library. A clone encoding a gene called NUF1 (for nuclear filament-related) was identified and extensively characterized. Antibodies to NUF1 fusion proteins were generated, and affinity-purified antibodies were used for immunoblot analysis and indirect immunofluorescence localization. The NUF1 protein is 110 kD in molecular mass and localizes to the yeast nucleus in small granular patches. Intranuclear staining is present in cells at all stages of the cell cycle. The NUF1 protein of yeast is tightly associated with the nucleus; it was not removed by extraction of nuclei with nonionic detergent or salt, or treatment with RNAse and DNAse. Sequence analysis of the NUF1 gene predicts a protein 945 amino acids in length that contains three domains: a large 627 residue central domain predicted to form a coiled-coil structure flanked by nonhelical amino-terminal and carboxy-terminal regions. Disruption of the NUF1 gene indicates that it is necessary for yeast cell growth. These results indicate that NUF1 encodes an essential coiled-coil protein within the yeast nucleus; we speculate that NUF1 is a component of the yeast nucleoskeleton. In addition, immunofluorescence results indicate that mammalian cells contain a NUF1-related nuclear protein. These data in conjunction with those in the accompanying manuscript (Yang et al., 1992) lead to the hypothesis that an internal coiled-coil filamentous system may be a general structural component of the eukaryotic nucleus.

Amino Acid Sequence↗
PubMed1989

Interactions of misfolded influenza virus hemagglutinin with binding protein (BiP).

We have characterized the association between the binding protein, BiP (also known as GRP 78), and misfolded forms of the influenza virus hemagglutinin precursor, HA0. BiP is a heat-shock-related protein that binds to unassembled immunoglobulin heavy chain and to a variety of misfolded proteins in the lumen of the ER. A small fraction (5-10%) of newly synthesized HA0 in CV-1 cells was found to be misfolded and retained in the ER. When glycosylation was blocked with tunicamycin, all of the HA0 produced was similarly misfolded. The misfolded HA0 was retained as relatively small (9-25-S) complexes associated with BiP. In these complexes the top domains of HA0 were correctly folded judging by their reactivity with monoclonal antibodies, but the polypeptides were cross-linked via anomalous interchain disulfides. The association with BiP was non-covalent and easily broken by warming to 37 degrees C or by adding ATP to the lysate. Pulse-chase experiments showed that HA0's self-association into complexes occurred immediately after synthesis and was followed rapidly by BiP association. The misfolded, BiP-associated HA0 was not transported to the plasma membrane but persisted as complexes in the ER for a long period of time before degradation (t1/2 = 6 h). The results suggested that BiP may be part of a quality control system in the ER and that one of its functions is to detect and retain misfolded proteins.

Animals↗
PubMed1988

Folding, trimerization, and transport are sequential events in the biogenesis of influenza virus hemagglutinin.

Results from several systems indicate that correct protein folding and subunit assembly correlate with the transport of membrane and secretory proteins from the endoplasmic reticulum (ER) to the Golgi complex. Because the site of oligomer assembly and its precise relationship to intracellular transport remain unclear, we have studied in detail the folding and trimerization of the influenza virus hemagglutinin (HA0) relative to its transport from ER to Golgi. Trimerization and transport were analyzed using several different methods, including transport inhibitors, temperature blocks, semi-intact cells, in vitro protein translocation, and immunocytochemistry. Taken together, the results clearly demonstrate that trimerization occurs at a point prior to exit from the ER. Before assembly, HA0 monomers were extensively folded and possessed intramolecular disulfide bonds, but monomers were not transported to the cis Golgi compartment. Thus, hemagglutinin progresses through at least two intermediate states before transport to the Golgi: highly folded monomers and trimers that have not yet left the ER.

Adenosine Triphosphate↗
PubMed1986

Assembly of influenza hemagglutinin trimers and its role in intracellular transport.

The hemagglutinin (HA) of influenza virus is a homotrimeric integral membrane glycoprotein. It is cotranslationally inserted into the endoplasmic reticulum as a precursor called HA0 and transported to the cell surface via the Golgi complex. We have, in this study, investigated the kinetics and cellular location of the assembly reaction that results in HA0 trimerization. Three independent criteria were used for determining the formation of quaternary structure: the appearance of an epitope recognized by trimer-specific monoclonal antibodies; the acquisition of trypsin resistance, a characteristic of trimers; and the formation of stable complexes which cosedimented with the mature HA0 trimer (9S20,w) in sucrose gradients containing Triton X-100. The results showed that oligomer formation is a posttranslational event, occurring with a half time of approximately 7.5 min after completion of synthesis. Assembly occurs in the endoplasmic reticulum, followed almost immediately by transport to the Golgi complex. A stabilization event in trimer structure occurs when HA0 leaves the Golgi complex or reaches the plasma membrane. Approximately 10% of the newly synthesized HA0 formed aberrant trimers which were not transported from the endoplasmic reticulum to the Golgi complex or the plasma membrane. Taken together the results suggested that formation of correctly folded quaternary structure constitutes a key event regulating the transport of the protein out of the endoplasmic reticulum. Further changes in subunit interactions occur as the trimers move along the secretory pathway.

Animals↗
PubMed1984

Bacillus Calmette-Guérin-activated murine macrophages kill syngeneic melanoma cells under strict anaerobic conditions.

We have studied the spontaneous killing of B5(59) melanoma cells by Bacillus Calmette-Guérin (BCG)-elicited macrophages under strictly anaerobic conditions to investigate the role of oxygen in macrophage-mediated cytotoxicity. The number of melanoma cells capable of forming colonies after aerobic or anaerobic incubation with BCG-macrophages was used as the index of cytotoxicity. The BCG-macrophages killed melanoma cells regardless of the amount of oxygen present. The killing observed was proportional to the ratio of effector cells added; a ratio of 25:1 effector to target cells was required to achieve nearly 90% cytotoxicity both aerobically and anaerobically. This cytotoxicity was not dependent on a diffusible macrophage product nor on alteration of the medium by macrophages, since tumor cells incubated in the same culture medium, but not in contact with a mixed population of tumor cells and macrophages, were not killed. These results also indicated that macrophage-mediated cytotoxicity was dependent on macrophage-tumor cell contact. The mechanism responsible for the oxygen-independent cytotoxicity is unknown at present.

Aerobiosis↗
PubMed1981

Regulation of human natural killing. I. The role of monocytes, interferon, and prostaglandins.

We have found that human endogenous natural killer activity as measured in a rapid 51Cr release assay is unaffected by the presence of monocytes. Moreover, stimulation of natural killer cells by poly I:C is independent of monocytes. In contrast, the presence of monocytes in a mixed population of mononuclear cells that have been stimulated by poly I:C suppresses NK activity. The suppression is shown to be partially reversible if indomethacin (10(-6) M) is added to the cultures during stimulation. Culture supernatants of monocytes stimulated with poly I:C are shown to contain PGE1 in a radioimmunoassay, and have NK inhibitory activity comparable to that obtained with exogenous PGE1 added to NK assays at a concentration range of 10(-7) to 10(-9) M. Culture supernatants from poly I:C-stimulated monocytes do not have detectable levels of antiviral activity. In contrast, plastic nonadherent cells stimulated with poly I:C produce significant levels of interferon (100 to 200 U/ml/2 x 10(6) cells) but almost undetectable amounts of PGE. Supernatants of nonadherent cells incubated with indomethacin (10(-6) M) alone augment NK activity. Taken together, the results suggest that stimulation of mononuclear cells with poly I:C is dependent on and regulated by the relative levels of interferon produced by lymphocytes and PGE produced by monocytes.

Cell Adhesion↗