Protein Synthesis In Mice Research Articles

Preparation of antibody (IgG)-ricin A-chain conjugate and its biologic activity.

As a model experiment for the preparation of cancer chemotherapeutic agents, a disulfide-linked conjugate of rabbit anti-mouse IgG (RaMIgG) and ricin A-cahin was synthesized. Activated sulfhydryl groups were first introduced into RaMIgG and then the modified RaMIgG was reacted with ricin A-chain. The fraction containing the conjugate was obtained from the reaction mixture by gel filtration through Sephadex G-150 and was characterized by sodium dodecyl sulfate-polyacrylamide disc gel electrophoresis and double immunodiffusion in agar. The inhibitory activity of A-chain against protein synthesis in a cell-free system and the binding activity of RaMIgG to mouse B lymphocytes were both fully retained. The purified conjugate selectively inhibited protein synthesis in mouse B lymphocytes possessing cell surface Ig, but not that in thymocytes. The cytotoxicity was inhibited by adding mouse IgG and an excess amount of RaMIgG to the cell culture. The effects of unlinked RaMIgG and A-chain on the cytotoxicity were negligible even when the two were used as a mixture.

Inhibition of protein synthesis in mouse L cells by poly-L-ornithine.

A snythetic polypeptide, poly-L-ornithine (pLo; average molecular weight, 13,000), inhibited protein synthesis in mouse L cells in suspension culture. The inhibition was dependent upon both the concentration of pLo and the cell density, and the dose for half-inhibition was correlated with the cell density. Cell viability remained high in the concentration range of PLo causing inhibition of protein synthesis. This inhibition was prevented by addiction of calf serum with pLo, but not by washing the treated cells with serum or a high concentration of salt. PLo had no effect on protein synthesis in a cell-free system prepared from L cells or rabbit reticulocytes. The polysome profiles of cells treated with pLo were similar to those of control cells. Experiments on the effect of interchange of the ribosomes and supernatants of control and pLo-treated cells showed that the ribosomes from pLo-treated cells were inactive. The inactivation of these ribosomes was partially prevented by the presence of 120 mM K+ in the medium during pLo treatment. The inhibition of protein synthesis by pLo, therefore, may result from the binding of pLo to the cell membranes, causing leakage of intracellular K+ and thus inactivating ribosomes.

Effect of reduced temperatures on protein synthesis in mouse L cells

The rate of incorporation of leucine into protein, the rate of polypeptide elongation and termination, and the relative quantity and size of polysomes were analyzed in mouse L cells grown in suspension culture at various temperatures between 0°C and 36°C. Between 10°C and 36°C protein synthesis exhibited two different apparent activation energies (39 kcal/mole, 10–25°C; 14 kcal/mole, 25–36°C), whereas elongation and termination had only one (16 kcal/mole). Below 36°C, the polysome level and size decreased, reaching a minimum of 30% of the control 36°C values at 10°C; below 10°C the level increased again back to control values at 0°C. The polysome decline was time dependent, requiring about 5 hr to reach the equilibrium value. This decline is completely reversible within 60 min, even in the presence of 4 μg/ml of actinomycin D, and even after 15 hr of incubation at the lower temperature. The results suggest that polypeptide initiation is rate limiting, particularly below 25°C; whereas above this temperature, elongation or perhaps some other process may be limiting. These results are quite different from those obtained for E. coli and rabbit reticulocyte protein synthesis.

Studies on the effect of diphtheria toxin on protein synthesis in mice.

SUMMARY Mice are several orders of magnitude more resistant to diphtheria toxin than are guinea-pigs and man. Resistance is, however, relative rather than absolute, because 2000 guinea-pig lethal doses cause death of mice within 36 hr. Mice subjected to this quantity of toxin subsequently received an injection of tritiated leucine during the latter stages of the toxaemia in order to assess de novo protein synthesis in vivo. All tissues from toxin-treated mice, with the exception of skeletal muscle and kidney, incorporated 3H-leucine to the same extent as did comparable tissues of normal animals, or to a greater extent. In view of the fact that the biochemical action of diphtheria toxin on more susceptible animals is known to result in a cessation of protein synthesis, the implications of the unexpected results are discussed in terms of species resistance to toxin and the pathophysiology of diphtheritic toxaemia.

Influence of inhibitors of protein synthesis on interferon formation in mice: II. Comparison of effects of glutarimide antibiotics and tenuazonic acid

Additional information has been obtained about the different patterns of interferon synthesis or release in mice injected with virus or bacterial endotoxin. Blockade of protein synthesis at different times after injection of Newcastle disease virus effectively halted further increase of circulating interferon. Data obtained by this method showed that interferon synthesis induced by virus is continuous for at least 10–12 hours. In the case of endotoxin, an enhanced and prolonged release of interferon was seen when blockade of protein synthesis was produced up to 30 min after the injection of endotoxin. By 1 or 2 hours after the injection of endotoxin, no significant enhancement of interferon release was effected by blockade of protein synthesis, indicating that the critical events involved in the release of preformed interferon by endotoxin occur within 30 min after the stimulus is injected. When a synthetic double-stranded polynucleotide, poly rI:rC, was used as the stimulus instead of endotoxin, the temporal relationship between blockade of protein synthesis and enhancement of interferon release in mice was similar. The effects of blockade of protein synthesis in mice described above were obtained using a series of glutarimide antibiotics (cycloheximide, acetoxycycloheximide, streptovitacin A, streptimidone) as well as tenuazonic acid, a chemically unrelated antibiotic.

Memory and cerebral protein synthesis in mice as affected by graded amounts of puromycin

The effects of intracerebral injections of graded amounts of puromycin on memory and on degree and duration of inhibition of cerebral protein synthesis have been studied in the mouse. As the concentration of the antibiotic was decreased it became progressively less effective in its behavioral and biochemical effects so that memory was retained in an increasing proportion of animals as the effect on protein synthesis diminished.

Organ Weight Changes and Protein Synthesis in Mice after Immunization to the Enzyme Triose Phosphate Dehydrogenase

Summary Mice immunized to the rabbit muscle enzyme, triose phosphate dehydrogenase, produced precipitating and neutralizing serum antibodies on and after the 9th day during the primary response. Administration of the antigen in adjuvant brought about approximately a 4-fold increase in the wet and dry weight of the spleen on the 7th and 8th days after the injection. After a decrease on the 9th day, spleen weight on the 11th day again reached a 4-fold enlargement. Most of this increase was caused by the mycobacterium component of the adjuvant. The relative rate of protein synthesis increased significantly in both the livers and spleens of animals that received antigen in adjuvant. C14-glycine incorporation into spleen protein for 90 min anticipated the succeeding 24-hr incremental increase in spleen weight. The possible contribution to spleen weight by migrating mononuclear cells from the granulomas is discussed.