Leakage Of Metabolites Research Articles

The biochemistry of virus-induced cell fusion. Changes in membrane integrity

1. Phospholipids prelabelled with [(14)C]acetate, [(32)P]phosphate, [(3)H]- or [(14)C]-choline or [(3)H]inositol are not significantly degraded during fusion of Lettrée cells mediated by Sendai virus, nor are carbohydrates prelabelled with [(3)H]fucose, [(14)C]galactose or [(3)H]glucosamine. Less than 1nmol of lysophosphatidylcholine/10(7) cells is formed during fusion. Diethyl p-nitrophenyl phosphate, which inhibits phospholipase A by more than 95% has no effect on fusion. It is concluded that none of the events leading to cell fusion is accompanied by significant turnover of phospholipids or other membrane components. 2. Intracellular K(+) leaks out during virally mediated cell fusion; the loss is not as extensive as that of intracellularly accumulated choline or deoxyglucose. Movement of Ca(2+) into or out of cells could not be detected. 3. At concentrations of Lettrée cells insufficient to be agglutinated by virus, intracellularly accumulated choline and deoxyglucose leak out. Agglutination caused by concanavalin A does not result in leakage of intracellular metabolites. 4. P815Y cells, which agglutinate but do not fuse in the presence of virus, show leakage of intracellularly accumulated metabolites. The extent of leakage does not alter during the G(1) and S periods of the cell cycle. 5. Leakage is inhibited by Ca(2+), but is unaffected by EDTA. 6. It is concluded that the interaction of Sendai virus with mammalian cells causes a weakening of membrane integrity so that intracellular metabolites leak out. Such destabilization may facilitate viral entry and is therefore an interesting system for further biochemical studies.

Permeability of Dictyostelium discoideum towards amino acids and inulin: a possible relationship between initiation of differentiation and loss of "pool" metabolites.

SUMMARY: The permeability of Dictyostelium discoideum amoebae towards amino acids and inulin was studied under conditions suitable for development of aggregation-competence and in thick cell suspensions. During the pre-aggregation period, the amoebae released large amounts of amino acids and nucleotides into the suspending medium. The uptake of glutamate and lysine by amoebae was passive, but inulin was taken up actively, probably by pinocytosis. Hence the simplest explanation for leakage of metabolites is that the amoebae have no active mechanism for retaining them. A possible relationship between initiation of differentiation and loss of pool metabolites is discussed.

Leakage of metabolites from mycelium of a low-temperature basidiomycete at a supramaximal temperature

Colonies of a low-temperature basidiomycete (W1) growing on agar media at 15 °C ceased growth when transferred to 30 °C. Viability was retained during at least 7 days exposure to 30 °C since growth resumed after a lag when colonies were returned to 15 °C. Measurements were made of the leakage of metabolites at 15 °C and 30 °C from mycelium of young shake cultures during a 5-h incubation period. Leakage of ninhydrin-positive material, almost negligible at 15 °C, was rapid at 30 °C. Increases in leakage of carbohydrate and ultraviolet-absorbing material occurred during the first hour at 30 °C. Subsequent leakage of carbohydrate at 30 °C, but not of ultraviolet-absorbing material, was at a slightly higher rate than at 15 °C. The possibility that these results indicate loss of cell membrane function at the supramaximal temperature is discussed.

Mechanism of action of the antifungal antibiotic pyrrolnitrin.

Pyrrolnitrin at 10 mug/ml inhibited the growth of Saccharomyces cerevisiae, Penicillium atrovenetum, and P. oxalicum. The primary site of action of pyrrolnitrin on S. cerevisiae was the terminal electron transport system between succinate or reduced nicotinamide adenine dinucleotide (NADH) and coenzyme Q. At growth inhibitory concentrations, pyrrolnitrin inhibited endogenous and exogenous respiration immediately after its addition to the system. In mitochondrial preparations, the antibiotic inhibited succinate oxidase, NADH oxidase, succinate-cytochrome c reductase, NADH-cytochrome c reductase, and succinate-coenzyme Q(6) reductase. In addition, pyrrolnitrin inhibited the antimycin-insensitive reduction of dichlorophenolindophenol and of the tetrazolium dye 2,2'-di-p-nitrophenyl-(3,3'-dimethoxy-4,4'-bi-phenylene)5,5'-diphenylditetrazolium. The reduction of another tetrazolium dye, 2-p-iodophenyl-3-p-nitrophenyl-5-phenyltetrazolium chloride, that was antimycin-sensitive, was also inhibited by pyrrolnitrin. The antibiotic had no effect on the activity of cytochrome oxidase, and it did not appear to bind with flavine adenine dinucleotide, the coenzyme of succinic dehydrogenase. In whole cells of S. cerevisiae, pyrrolnitrin inhibited the incorporation of (14)C-glucose into nucleic acids and proteins. It also inhibited the incorporation of (14)C-uracil, (3)H-thymidine, and (14)C-amino acids into ribonucleic acid, deoxyribonucleic acid, and protein, respectively. The in vitro protein synthesis in Rhizoctonia solani and Escherichia coli was not affected by pyrrolnitrin. Pyrrolnitrin also inhibited the uptake of radioactive tracers, but there was no general damage to the cell membranes that would result in an increased leakage of cell metabolites. Apparently, pyrrolnitrin inhibits fungal growth by inhibiting the respiratory electron transport system.

Sugar Retention, Cofactor Levels, and Leakage of Metabolites in X-Irradiated, Starved Yeast Cells

Abstract : Irradiated cells retained a greater portion of (14C)sorbose, though rates of efflux were similar during a first phase of exit and rates of uptake were similar. The thiamine pyrophosphate content of irradiated cells was greater than that of unirradiated cells, whereas pyruvate decarboxylase activities and ATP and DPN contents were similar. Similar amounts of 260 millimicrons absorbing materials were lost by irradiated and unirradiated cells during starvation. Incubation with glucose or maltose increased the total loss, while incubation with severa less readily metabolized sugars or with hexitols increased the relative loss from irradiated cells. (Author)

Effect of vancomycin on Bacillus subtilis

Vancomycin inhibits growth and mucopeptide synthesis in Bacillus subtilis strain W23. These effects can be minimized by added magnesium. The metal was most effective in alleviating growth inhibition when added simultaneously with the antibiotic. Addition of magnesium at later time intervals showed less response. High concentrations of vancomycin (8.5 μg. per milligram dry wt.) caused a rapid lysis of this organism when incubated in a complete medium. Leakage of cell metabolites was observed with sublytic vancomycin concentrations (4.6 μg per milligram dry wt.). Magnesium was unable to prevent either lysis or leaching of cell metabolites. A mechanism of inhibition based on these findings is suggested.

A comparative study of respiratory metabolism in cultured mammalian cell strains

The respiration of three different cell types, fibroblastic (human skin fibroblast and Y5), epithelial (HeLa and HLM), and reticuloendothelial (L-5178Y) have been studied with particular reference to influences on cellular respiration of the following environmental factors: (1) the presence of Krebs cycle intermediates in the nutrient medium, (2) hydrogen ion concentration, (3) glucose concentration, (4) oxygen tension, and (5) carbon dioxide tension. Whereas the respiration of strain L, human fibroblast and L-5178Y cells declined rapidly after inoculation into new medium, this did not happen with HeLa, HLM and Y5 cells. The fall in respiration of the first group could be prevented by adding Krebs cycle acids to the medium. Keto-acids were found to leak more rapidly from strain L cells than from HeLa and the fall in respiration of the former is considered to be due to the rapid leakage of metabolites. Epithelial cells demonstrated a classical Crabtree effect but this was less pronounced in fibroblastic cells. In all cells respiration was less in cultures maintained at pH 7.8 than in cultures maintained at pH 7.4. In most cells it was also lower at pH 6.8 but the HLM cell was an exception. If maintained at oxygen tensions above or below that obtained in equilibrium with air, respiration was reduced. In all cells respiratory rate was higher in the presence of 1 per cent CO 2 than at lower concentrations. Higher concentrations were slightly inhibitory.