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Biomedical subjects

R B Wilson

Publications and source records attributed to R B Wilson.

At least 55 records · Page 3Linked to original sources
PubMed1993

A Chinese hamster ovary cell line with a temperature-conditional defect in receptor recycling is pleiotropically defective in lysosome biogenesis.

We have previously described the isolation of a Chinese hamster ovary cell line, TfT1.11, that has a pleiotropic, temperature-conditional defect in receptor recycling (Cain, C. C., Wilson, R. B., and Murphy, R. F. (1991) J. Biol. Chem. 266, 11746-11752). These cells show a rapid loss of cell surface receptors upon temperature shift due to a reduction in the rate of receptor recycling. We show here that, in addition to altered receptor recycling, TfT1.11 cells show three defects in lysosome biogenesis. At the nonpermissive temperature, they 1) redistribute at least one lysosomal enzyme from lysosomes to endosomes, 2) fail to transfer fluid-phase material from early endosomes to later compartments, and 3) fail to accumulate fluid-phase markers due to increased efflux of internalized material. The results suggest that the processes of recycling from the endosome and movement of material from endosomes to lysosomes are tightly linked.

Animals↗
PubMed1993

The pde2 gene of Saccharomyces cerevisiae is allelic to rca1 and encodes a phosphodiesterase which protects the cell from extracellular cAMP.

The high affinity cAMP phosphodiesterase, encoded by PDE2, is an important component of the cAMP-dependent protein kinase signaling system in Saccharomyces cerevisiae. An unexpected phenotype of pde2 mutants is sensitivity to external cAMP. This trait has been found independently for rca1 mutants and has been used to monitor the effects of cAMP on several biological processes. We demonstrate here that RCA1 is identical to PDE2. Further analysis of the phenotype of pde2 deletions reveal that exogenously added cAMP results in an increase in the internal level of cAMP. This increase slows down the rate of cell division by increasing the length of the G1 phase of the cell cycle and leads to increased cell volume. Also, cells with a disrupted PDE2 gene previously arrested by nutrient starvation rapidly lose thermotolerance when incubated with exogenous cAMP. From these observations we propose that a role of the PDE2-encoded phosphodiesterase may be to help insulate the internal cAMP pools from the external environment. This protective role might also be important in other eukaryotic organisms where cAMP is a key second messenger.

3',5'-Cyclic-AMP Phosphodiesterases↗
PubMed1993

Pulmonary response, in vivo, to silicon carbide whiskers.

Fischer rats were exposed to silicon carbide whiskers (SiCW), boron carbide whiskers (BCW), silicon carbide platelets (SiCP), or crocidolite asbestos separately administered by intratracheal instillation. SiCW proved to be the most toxic material within the test group. Dramatic increases in alveolar macrophage populations within 1 week of exposure to SiCW persisted for at least 28 days, evidence of the chronic inflammation observed in necropsies during the first months of the study. The most common finding in histological preparations of tissues taken from animals 18 months after exposure to SiCW was a high incidence (frequency > 0.85) of multiple pulmonary granulomas which occasionally occluded airways. Lesions associated with crocidolite were similar to those found with SiCW. Equivalent treatment with BCW and SiCP produced no significant histological changes within 18 months of exposure.

Administration, Inhalation↗
PubMed1993

Accessibility and distribution of intraerythrocytic antigens of Plasmodium-infected erythrocytes following mild glutaraldehyde fixation and detergent extraction.

Malarial antigens on the surface of infected erythrocytes have been described by many investigators. However, few of these antigens have been unambiguously demonstrated to be exposed on the surface of erythrocytes. This study demonstrates that mild glutaraldehyde fixation results in the cytoplasmic face of the host membrane becoming accessible to antibody under conditions that normally do not expose the cytoplasmic face of uninfected erythrocytes. These results indicate that caution should be used in interpreting data on the membrane disposition of malarial antigens. Detergent extraction of the glutaraldehyde-fixed erythrocytes results in an increased permeabilization such that malarial antigens on the parasite surface and within the cytoplasm of the infected erythrocyte are accessible to antibody. The accessibility of these antigens was demonstrated by both immunofluorescence and two-color flow cytometry. The antigens within the host cytoplasm were not diffuse but associated with patchy aggregates. Analysis of the antigens associated with the cytoplasmic aggregates by immunoelectron microscopy indicated that they were not associated with membrane-bound compartments. The fixation and permeabilization protocol described herein will have useful applications for the characterization and analysis of malarial antigens.

Animals↗
PubMed1991

Isolation by fluorescence-activated cell sorting of Chinese hamster ovary cell lines with pleiotropic, temperature-conditional defects in receptor recycling.

We have isolated several Chinese hamster ovary cell lines with temperature-sensitive defects in the recycling of receptors after endocytosis. These cell lines were selected using fluorescence-activated cell sorting for retention of a pulse of labeled transferrin after a chase in the presence of unlabeled transferrin. One of these cell lines, TfT1.11, was selected for further characterization. In TfT1.11 the trapping of transferrin within the cells is paralleled by a loss of cell surface transferrin receptors. Within 4 h after the shift from 33 to 41 degrees C the surface binding of transferrin is reduced to 18% of parental cells at 41 degrees C. The trapping of transferrin and the loss of transferrin receptor from the cell surface are caused by a temperature-conditional 5.5-fold decrease in the initial rate of transferrin recycling. TfT1.11 cells also rapidly lose 89% of their ability to take up alpha 2-macroglobulin after the temperature shift to 41 degrees C. These data indicate that the TfT1.11 cell line has a pleiotropic defect in receptor recycling.

Adenosine Triphosphate↗
PubMed1991

Psittacine proventricular dilatation syndrome in an umbrella cockatoo.

Psittacine proventricular dilatation syndrome (macaw wasting disease) is a fatal disease of the gastrointestinal tract and, sometimes, the CNS. The disease most often affects macaws and is thought to be of viral origin. An Umbrella cockatoo was referred with signs of neurologic dysfunction. Other unusual findings included flaccid crop, distended duodenum, and acute weight loss. Because treatment has not been reported to be effective, the affected cockatoo was euthanatized to limit spread of the disease.

Animals↗
PubMed1991

Repression of immunoglobulin enhancers by the helix-loop-helix protein Id: implications for B-lymphoid-cell development.

It has been proposed that the helix-loop-helix (HLH) protein Id serves as a general antagonist of cell differentiation by inhibiting bHLH (HLH with an adjacent stretch of basic amino acids) proteins specifically required for developmental programs (such as MyoD). We show here that ectopic expression of Id represses in vivo activity of the bHLH protein E2-5 (encoded by the E2A gene) and of both the immunoglobulin heavy-chain (IgH) and kappa-light-chain gene enhancers to which E2-5 binds. Id does not affect the activity of the bHLH-zip protein, TFE3, which also binds these enhancers. We examined a large panel of B-cell lines that represent different stages of lymphoid development and found only two that express Id mRNA. The cell lines Ba/F3 and LyD9 have been categorized previously as early B-lymphoid-cell progenitors. Unlike their more mature B-lymphoid-cell counterparts, Ba/F3 and LyD9 cells do not express I mu sterile transcripts, which are indicative of IgH enhancer activity. Moreover, Ba/F3-derived nuclear extracts lack E2-box-binding activity, indicating the absence of free bHLH proteins, and transfected Ba/F3 cells fail to support the activity of the IgH enhancer. Hence, expression of Id correlates inversely with bHLH protein activity and enhancer function in vivo. These results suggest that Id may play a role early in B-lymphoid-cell development to regulate transcription of the IgH locus.

3T3 Cells↗
PubMed1991

The Saccharomyces cerevisiae SRK1 gene, a suppressor of bcy1 and ins1, may be involved in protein phosphatase function.

The Saccharomyces cerevisiae SRK1 gene, when expressed on a low-copy shuttle vector, partially suppresses the phenotype associated with elevated levels of cyclic AMP-dependent protein kinase activity and suppresses the temperature-sensitive cell cycle arrest of the ins1 mutant. SRK1 is located on chromosome IV, 3 centimorgans from gcn2. A mutant carrying a deletion mutation in srk1 is viable. SRK1 encodes a 140-kDa protein with homology to the dis3+ protein from Schizosaccharomyces pombe. The ability of SRK1 to alleviate partially the defects caused by high levels of cyclic AMP-dependent protein kinase and the similarity of its encoded protein to dis3+ suggest that SRK1 may have a role in protein phosphatase function.

Amino Acid Sequence↗
PubMed1990

Filtration of platelet-poor plasma specimens yields platelet-free plasma. Application for batch analysis of beta-thromboglobulin and platelet factor 4.

The authors investigated filtration of platelet-poor plasma through a 0.2-micron filter for the analysis of beta-thromboglobulin (beta-TG) and platelet factor 4 (PF4) in frozen specimens. Platelets were detected by electron micrograph in the platelet-poor plasma preparation without filtration. No platelets or platelet fragments were seen after filtration. Freezing the unfiltered specimens resulted in a significant increase in the beta-TG and PF4 concentrations, presumably because of release of these proteins from the platelet alpha-granules during freezing and thawing. There was no change in the PF4 levels and only a slight decrease in beta-TG levels after freezing of the filtered specimens. The filtered samples could be frozen for later batch analysis, resulting in considerable cost savings to the laboratory.

Blood Platelets↗
PubMed1990

Endosomes can undergo an ATP-dependent density increase in the absence of dense lysosomes.

On the basis of evidence that lysosomal enzymes and membrane proteins are present in endosomes, we have previously suggested that the production of lysosomes involves maturation rather than vesicle fusion (Roederer, M., R. Bowser, R. F. Murphy, J. Cell. Physiol. 131, 200-209 (1987)). Since the appearance of endocytosed material in lysosomes is associated with an increase in buoyant density from that of endosomes, a prediction of the model is that endosomes should be capable of undergoing such an increase in vitro. We observe that under appropriate conditions, isolated endosomes containing [125I]EGF can undergo an increase in density in vitro to that of dense lysosomes, mimicking the density change which occurs in vivo. This occurs in the absence of dense lysosomes with which to fuse. The density increase requires ATP and can be efficiently inhibited in vitro by the presence of benzylamine, suggesting that vesicular acidification is required. Since low pH has previously been shown to induce formation of a matrix by lysosomal enzymes in vitro (Buckmaster, M. J., A. L. Ferris, B. Storrie, Biochem. J. 249, 921-923 (1988)), we propose that a mechanism by which endosomes and/or lysosomes increase their density is a low pH induced aggregation of vesicle contents which decreases the osmotic pressure inside the vesicle. Together with previous data, the results provide highly suggestive evidence that the pathway to lysosomes includes a maturation of the postsorting compartment into what has classically been termed a lysosome.

Adenosine Triphosphate↗
PubMed1990

Cholesterol:phospholipid ratio is elevated in platelet plasma membrane in patients with hypertension.

The cholesterol:phospholipid ratio was measured in platelet plasma membrane, red blood cell (RBC) membranes, low density lipoprotein (LDL) and whole plasma in patients with primary hypertension and in matched normal controls. The cholesterol:phospholipid ratio was raised in the platelet membrane from hypertensive patients compared with that from normal controls (0.65 +/- 0.03 vs 0.53 +/- 0.02: mean +/- SEM; P less than 0.01). The ratio observed in RBC membranes, LDL and whole blood was similar in the two groups. If this abnormality in the lipid composition of platelet plasma membrane is present in other cells it could account for some of the changes in cell membrane function that have been described in hypertension.

Adult↗
PubMed1989

A dipeptide insertion in domain I of exotoxin A that impairs receptor binding.

Deletions within the structural exotoxin A gene of 27 or 119 amino acids in domain I of the mature polypeptide, or of 88 or 105 amino acids in domains I and II, resulted in the synthesis of exotoxin A (ETA) polypeptides that were not secreted from Pseudomonas aeruginosa hosts but were localized in the cell membrane. Insertions of a hexanucleotide sequence, either pCGAGCT or pCGAATT, at TaqI sites within the gene resulted in variant exotoxin A polypeptides which were secreted normally. pCGAGCT causes insertion of either Glu-Leu or Ser-Ser in the amino acid sequence of the toxin, while pCGAATT causes insertion of either Glu-Phe or Asn-Ser dipeptides. Although the cytotoxicity of eight variants was unimpaired, that of four others was reduced, and one variant which had a Glu-Phe insert between residues 60 and 61 (ETA-60EF61) was 500-fold less cytotoxic than wild-type exotoxin A. Purified ETA-60EF61 dissociated much faster from mouse LMTK- cells than wild-type ETA, suggesting that the insertion impaired the ability of ETA-60EF61 to interact with exotoxin A receptors. The location of the insert is within a major concavity on the surface of domain I of the exotoxin A molecule, suggesting that this concavity is important for toxin-receptor interaction.

ADP Ribose Transferases↗
PubMed1989

Tonsillar granular cell tumour in a cat.

Granular cell tumour was diagnosed in a cat based on light and electron microscopic findings. Immunohistochemical findings for S-100 protein and neuron-specific enolase were negative, unlike its human counterpart.

Animals↗
PubMed1989

Choroid plexus carcinoma in a dog.

Choroid plexus carcinoma was diagnosed in a 5-year-old female mixed breed dog which was euthanized due to progressive neurologic disease. Diagnosis of the tumour was based on gross and light microscopic findings following a complete necropsy. The chemical staining patterns in the case are compared with human choroid plexus tumours. The criteria for the distinction between benign and malignant variants of choroid plexus tumours are discussed.

Animals↗