Rous sarcoma virus-transformed baby hamster kidney cells express higher levels of asparagine-linked tri- and tetraantennary glycopeptides containing [GlcNAc-beta (1,6)Man-alpha (1,6)Man] and poly-N-acetyllactosamine sequences than baby hamster kidney cells

The alterations in complex-type N-linked oligosaccharides that can occur when an animal cell line is transformed by two dissimilar viruses were examined by comparing the N-linked oligosaccharides of baby hamster kidney (BHK) cells, metabolically radiolabeled with [2-3H]mannose, to the same class of oligosaccharides from BHK cells separately transformed by Rous sarcoma virus (RS-BHK), an RNA retrovirus, and polyoma virus (PY-BHK), a DNA papovavirus. Based on experiments that utilized serial lectin affinity chromatography, glycosidase digestions, and methylation analyses, both RS-BHK and PY-BHK cells demonstrated a significant increase in the relative amounts of tri- and tetraantennary complex-type N-linked oligosaccharides containing the branching sequence, [GlcNAc-beta(1,6)Man-alpha(1,6)Man], compared to the nontransformed BHK cells. In addition, almost all of the poly-N-acetyllactosamine sequence, [GlcNAc-beta(1,3)-Gal-beta(1,4)], was expressed on the tri- and tetraantennary N-linked oligosaccharides from BHK and RS-BHK cells that contain the sequence, [GlcNAc-beta(1,6)Man-beta(1,6)Man]. The increase in the relative amounts of this latter sequence in the transformed cells, therefore, most likely results in an increase in the amount of poly-N-acetyllactosamine sequence on the N-linked glycopeptides of these cells. The analysis of the degree of sialylation of the complex-type N-linked oligosaccharides from BHK and RS-BHK cells by ion exchange chromatography revealed no apparent differences, and in both of these cell types approximately 3% of the glycopeptide fraction radiolabeled with mannose was recovered in a highly negatively charged fraction that was identified by keratanase digestion to be keratan sulfate.     

The v-sis/PDGF-2 transforming gene product localizes to cell membranes but is not a secretory protein

The v-sis transforming gene encodes the woolly monkey homologue of human platelet-derived growth factor (PDGF) polypeptide 2. After its synthesis on membrane bound polyribosomes, the glycosylated precursor dimerizes in the endoplasmic reticulum and travels through the Golgi apparatus. At the cell periphery, the precursor is processed to yield a dimer structurally analogous to biologically active PDGF. Small amounts of two incompletely processed forms are detectable in tissue culture fluids of simian sarcoma virus (SSV) transformants. However, the vast majority remains cell associated. Thus, this growth factor-related transforming gene product is not a classical secretory protein. These findings define possible cellular locations where the transforming activity of the sis-PDGF-2 protein may be exerted.     

Augmentation of lung antineutrophil elastase capacity with recombinant human alpha-1-antitrypsin

To evaluate the potential use of recombinant DNA-produced alpha-1-antitrypsin (alpha-1-AT) to augment the lung antineutrophil elastase defenses in alpha-1-AT deficiency, we compared the kinetics of intravenously administered recombinant produced alpha-1-AT (r alpha-1-AT) and purified normal human plasma alpha-1-AT (p alpha-1-AT) in the blood and lung of rhesus monkeys. The r alpha-1-AT was produced in yeast transformed with an expressing plasmid containing a full-length human alpha-1-AT complementary deoxyribonucleic acid and purified to greater than 99% homogeneity. The r alpha-1-AT has a molecular weight of 45,000, no carbohydrates, and is identical in sequence to normal plasma alpha-1-AT except for an additional N-terminal acetylmethionine. Despite its lack of carbohydrates, the r alpha-1-AT inhibited human neutrophil elastase with an association rate constant similar to that of p alpha-1-AT. Rhesus monkeys were infused intravenously with 120 mg/kg of r alpha-1-AT (n = 13) or p alpha-1-AT (n = 12) and the serum, urine, and lung epithelial lining fluid (ELF) concentrations of these molecules quantified at various intervals.(ABSTRACT TRUNCATED AT 250 WORDS)     

Recognition sites for a membrane-derived DNA binding protein preparation in the E. coli replication origin

Summary The DNA binding protein B' preparation, isolated from the membrane of E. coli, recognizes two sites, one of which is locatd in the minimum oriC (35–270 bp) and the other between base pairs 417 and 488. Recognition is only possible when restriction fragments containing these sites are in single-stranded state. At the first site the strand reading 3OH-5P in the direction of the E. coli genetic map is recognized, at the second site the 5P-3OH strand.     

Stomatal movement in Zea mays: Shuttle of potassium and chloride between guard cells and subsidiary cells

Summary When stomates of Zea mays open K and Cl migrate from the subsidiary cells into the guard cells; when the stomates close both elements return to the subsidiary cells. Subsidiary cells function as reservoirs for K and Cl. Import of K and Cl into the guard cells and loss of both elements from the guard cells become observable 1 or 2 min after light is turned on or off, both when histochemical methods and the electron-probe microanalyzer are used for detection. Each stomatal complex of maize contains on the average 10±3×10^-13 gram equivalents (eq) of K and 4±1×10^-13 eq of Cl. Guard cells accumulate K in the light and CO₂-free air at an average rate of 10×10^-15 eq K per minute, and Cl at approximately half that rate.     

INCREASE OF GLUCOSE AND HIGH ENERGY PHOSPHATE RESERVE IN THE BRAIN AFTER HYDROCORTISONE1

Abstract— Young mice treated with hydrocortisone (50 mg/kg) subcutaneously for 10 days showed a doubling of brain glucose. Brain phospho-creatine, glucose-6-phosphate, and ATP increased slightly. Brain glycogen and lactate were unchanged. Total energy reserve of the brain was 23 per cent higher than the control value. Liver glycogen was increased 47 per cent; liver and blood glucose levels were 11 per cent lower than in control animals. Since the animals showed no evidence of sedation, these findings suggest a facilitated transport of glucose from blood into the brain under the influence of hydrocortisone. Other possible explanations include an inhibition of the hexose monophosphate shunt and a proportionate decrease in both the oxidative and glycolytic pathways of the brain, but it was concluded that these explanations are less likely.     

Area-sampling Technique for Quantitative Pharyngeal Cultures

An area-sampling device for obtaining quantitative samples of the oropharyngeal bacterial flora is described and illustrative data are presented.     

ENZYME-LABELED ANTIBODIES FOR THE LIGHT AND ELECTRON MICROSCOPIC LOCALIZATION OF TISSUE ANTIGENS

Enzymes, either acid phosphatase or horseradish peroxidase, were conjugated to antibodies with bifunctional reagents. The conjugates, enzymatically and immunologically active, were employed in the immunohistochemical localization of tissue antigens utilizing the reaction product of the enzymatic reaction as the marker. Tissues reacted with acid phosphatase-labeled antibodies directed against basement membrane were stained for the enzyme with Gomori's method, and those reacted with peroxidase-labeled antibody were stained with Karnovsky's method. The reaction products of the enzymes localized in the basement membrane. Unlike the preparations of the fluorescent antibody technique, enzyme-labeled antibody preparations were permanent, could be observed with an ordinary microscope, and could be examined with the electron microscope. In the latter, specific localization of antibody occurred in the basement membrane and in the endoplasmic reticulum of cells known to synthesize basement membrane antigens. The method is sensitive because of the amplifying effect of the enzymatic activity. The ultrastructural preservation and localization were better with acid phosphatase-labeled antibody than with peroxidase-labeled antibody, but acid phosphatase conjugated antibody was unstable and difficult to prepare. Peroxidase-antibody conjugates were stable and could be stored for several months at 4°C, or indefiniely in a frozen state.