Oscillations of membrane potential in L cells

Summary Effects of divalent cations on oscillations of membrane potentials (i.e., spontaneous repetitive hyperpolarizing responses) and on hyperpolarizing responses induced by electrical stimuli as well as on resting potentials were studied in large nondividing L cells. Deprivation of Ca²⁺ from the external medium inhibited these hyperpolarizing responses accompanying slight depolarization of the resting potential. Sr²⁺ or Mn²⁺ applied to the external medium in place of Ca²⁺ was able to substitute for Ca²⁺ in the generation of hyperpolarizing responses, while Mg²⁺, Ba²⁺ or La³⁺ suppressed hyperpolarizing responses. The addition of A23187 to the bathing medium or intracellular injection of Ca²⁺, Sr²⁺, Mn²⁺ or La³⁺ induced membrane hyperpolarization. When the external Ca²⁺, Sr²⁺ or Mn²⁺ concentration was increased, the resting potential also hyperpolarized, in a saturating manner. The amplitude of maximum hyperpolarization produced by high external Ca²⁺ was of the same order of magnitude as those of hyperpolarizing responses and was dependent on the external K⁺ concentration. In the light of these experimental observations, it was deduced that the K⁺ conductance increase associated with the hyperpolarizing excitation is the result of an increase in the intracellular concentration of free Ca²⁺ mainly derived from the external solution.     

Amino acids in a carbonaceous chondrite from Antarctica

Summary A carbonaceous chondrite from the Antarctic, referred to as the Allan Hills meteorite 77306, appears to be free from terrestrial organic contamination. The presence of both protein and non-protein amino acids and an equal abundance of D- and L-enantiomers of amino acids, is testimony to the extraterrestrial nature of these compounds.     

Relationships between nucleic acid,nitrogen, and growth rate of tobacco cells in suspension culture

Summary Changes in the amount of nucleic acid and nitrogen, and the relationships between these amounts and the growth rate of tobacco cells (Nicotiana tabacum L. cv. Bright Yellow-2) at different initial nitrogen concentrations in the medium, were examined in batch cultures. During culture in basal medium, the amount of intracellular nucleic acid expressed per unit of dry biomass was 36.3 mg RNA g^–1 cell and 8.1 mg DNA g^–1 cell at the beginning of batch culture. These values increased 2.5 fold for RNA and 1.5 fold for DNA during the exponential growth phase and then gradually decreased with the decline in the growth rate. Similar changes were also observed in the medium containing less nitrogen. The specific growth rate, (day^–1), of the culture corresponded to the magnitude of the intracellular RNA content (mg RNA g^–1 cell), and the linear relationship, RNA=38+23 was obtained. In addition, there were remarkable positive correlations between the total and protein nitrogen, and during the cultures. The mononucleotide composition of total RNA (AMP+UMP)/(GMP+CMP) which was suggested to be a convenient index of metabolic activity was nearly constant (0.78 to 0.80) during tobacco cell culture in the basal medium.     

Effect of piperonyl butoxide and diphenylamine on lipid peroxidation in ozonated chloroplasts

Lipid peroxide (LPO) formation was remarkable when isolatedtobacco chloroplasts were bubbled with high concentrations ofozone, though the fatty acid composition and the fractionationpattern of glycolipids and phospholipids in the chloroplastlipids changed little after ozone fumigation of the leaves.Piperonyl butoxide (PB), a potent protectant against ozone injury,strongly inhibited LPO formation in ozonated chloroplasts. PBalso prevented ozone-induced decreases in the amounts of linolenicand linoleic acids in the chloroplast lipids. These resultssuggest that PB inhibition of LPO formation may be involvedin the protective mechanism against ozone phytotoxicity. However,the mode of PB action differed on some points from that of diphenylamine,which is an antioxidant and also effective against ozone injury.The mode of PB action is discussed. ¹ Present address: The Central Research Institute, Japan Tobacco& Salt Public Corporation, Umegaoka, Midori-ku, Yokohama227, Japan. (Received July 5, 1976; )     

Redistribution of intramembrane particles of human erythrocytes induced by HVJ (Sendai virus): A prerequisite for the virus-induced cell fusion

Aggregation of intramembrane particles of human erythrocytes was found to be induced by HVJ (Sendai virus) under conditions which lead to cell fusion. Degree of polyerythrocyte formation was compared under a variety of conditions with extent of cluster formation observed with the same preparations. Both structural changes of the membranes, ie, fusion and clustering of the particles, behaved very similarly under widely different virus-to-cell ratios and over the time course of cell fusion. Furthermore, by inclusion of high concentrations of antispectrin antibodies within the ghosts, inhibition of clustering of intramembrane particles and hindrance of virus-induced cell fusion were found to occur simultaneously. Antibodies by themselves did not induce aggregation of particles under isotonic conditions, whereas particle clustering could be induced under hypotonic conditions at antibody concentrations causing partial cross-linking of spectrin molecules. In conclusion, clustering of intramembrane particles seems to be required for virus-induced fusion of human erythrocytes.     

Studies on the mechanism of the conversion of cytosol δ-aminolevulinic acid synthase to the mitochondrial enzyme in the liver of the allylisopropylacetamide-treated rat

δ-Aminolevulinic acid (ALA) synthase was partially purified from liver cytosol fraction of rats treated with allylisopropylacetamide (AIA). The cytosol ALA synthase showed an apparent molecular weight of 320,000. The cytosol ALA synthase of this size dissociates into at least three protein components when subjected to sucrose density gradient centrifugation in the presence of 0.25 m NaCl: one is the catalytically active protein with an s value of about 6.4 or a molecular weight of 110,000, and the other two are catalytically inactive binding proteins showing s values of about 4 and 8, respectively. Recombination of the 6.4 S protein and the 4 S protein yielded a protein complex with an apparent molecular weight of 170,000 and recombination of all three protein components resulted in formation of the original cytosol ALA synthase. The cytosol ALA synthase also loses its binding proteins when treated with various proteases; thus, the enzyme-active protein obtained after papain digestion was very similar, if not identical, to mitochondrial ALA synthase. When treated with trypsin, however, the cytosol ALA synthase was converted to an enzyme showing an apparent molecular weight of 170,000, which probably represents the complex of the mitochondria-type enzyme and the 4 S binding protein. The cytosol ALA synthase tends to aggregate to form a dimer with an apparent molecular weight of 650,000–700,000. The aggregated form of the cytosol ALA synthase was less susceptible to trypsin digestion. Hemin strongly stimulated dimer formation of the cytosol ALA synthase and the aggregate produced by contact with hemin was very tight and did not easily dissociate into its respective protein components by sucrose gradient centrifugation or even after treatment with trypsin. The possible mechanisms of the conversion of cytosol ALA synthase to the mitochondrial enzyme and also of the inhibition by hemin of the intracellular translocation of ALA synthase are discussed.     

The role of virulence antigens (VW) in the protection of mice againstYersinia pestis infection

Subcutaneous infection withYersinia enterocolitica harboring plasmid responsible for Ca²⁺ dependence at 37°C induced cell-mediated protective immunity against a lethal challenge withYersinia pestis; the isogenic derivative strain cured from this plasmid subverted the immunity in mice. This is the first identification of the antigen(s) responsible for the induction of cell-mediated protective immunity against the facultatively intracellular bacteria.     

The asparagine-linked sugar chains of the glycoproteins in rat erythrocyte plasma membrane—Fractionation of oligosaccharides liberated by hydrazinolysis and structural studies of the neutral oligosaccharides

The asparagine-linked sugar chains of the plasma membrane glycoproteins of rat erythrocytes were released as oligosaccharides by hydrazinolysis and labeled by NaB³H₄ reduction. The radioactive oligosaccharides were separated into a neutral and at least four acidic fractions by paper electrophoresis. The neutral oligosaccharide fraction was separated into at least 11 peaks upon Bio-Gel P-4 column chromatography. Structural studies of them by sequential exoglycosidase digestion in combination with methylation analysis revealed that they were a mixture of three high mannose-type oligosaccharides and at least 11 complex type oligosaccharides with Manα1 → 6(Manα1 → 3)Manβ1 → 4GlcNAcβ1 → 4(±Fucα1 → 6)GlcNAc as their cores and Galβ1 → 4GlcNAc, Galβ1 → 3Galβ1 → 4GlcNAc, and various lengths of Galβ1 → 4GlcNAc repeating chains in their outer chain moieties. Most of the complex-type Oligosaccharides were biantennary, and the tri- and tetraantennary Oligosaccharides contain only the Galβ1 → 3Galβ1 → 4GlcNAc group in their outer chain moieties.