Challakonda N. Murty

In vivo and in vitro studies on the effect of tryptophan on translocation of RNA from nuclei of rat liver.

Abstract The effect of a single administration of tryptophan on the transport of nuclear RNA to the cytoplasm of rat liver was investigated. Using a cell-free system containing nuclei (RNA labeled in vivo with [14C]orotate) and dialyzed cell sap of liver, the isolated hepatic nuclei of tryptophantreated rats demonstrated an increase in the release of labeled RNA into the medium in comparison to hepatic nuclei of control rats. Also, liver cell sap of tryptophan-treated rats with added tryptophan to the medium was found to stimulate the release of labeled RNA (poly(A)-mRNA) from isolated control nuclei. Pretreatment of rats with cycloheximide or puromycin before tryptophan administration prevented the liver cell sap effect. However, the hepatic nuclei of puromycin- and tryptophan-treated rats demonstrated an increase in the release of labeled RNA into the medium incubated with control liver sap, similar to that observed with hepatic nuclei of rats treated with tryptophan alone. These results suggest that the tryptophan-induced enhancement of nucleocytoplasmic translocation of RNA may be related to alterations in the controls within the nucleus and cytoplasm that act to regulate the intracellular transport of RNA.

Effect of tryptophan on nuclear envelope nucleoside triphosphatase activity in rat liver.

Summary The activity of nuclear envelope nucleoside triphosphatase, an enzyme implicated to be involved in nucleocytoplasmic translocation of mRNA, was investigated in the livers of rats that received a single tube-feeding of tryptophan. Tryptophan administered to fasted rats or to rats pretreated with puromycin or actinomycin D caused significant increases in the activity of Mg2+-dependent nucleoside triphosphatase in the hepatic nuclear envelopes over that in controls. Concomitant with this rapid (10 min) increase in the enzyme activity, there was more release of RNA from isolated nuclei of livers of tryptophan-treated rats than from those of control rats.

Effect of tryptophan on hepatoma and host liver of rats. Influence after treatment with hypertonic sodium chloride and carbon tetrachloride.

Abstract Female inbred Buffalo rats bearing intrahepatically transplanted hepatoma 5123 were subjected intraperitoneally to the acute administration of hypertonic NaCl or CCl 4 followed by a tube-feeding of l -tryptophan. The responses in terms of changes in polyribosomal aggregation and protein synthesis ( in vitro ) of host liver and hepatoma were evaluated. While treatment with hypertonic NaCl or CCl 4 caused disaggregation of polyribosomes and inhibition of protein synthesis in both host liver and hepatoma, the subsequent administration of tryptophan caused some improvement in both parameters in host liver but not in hepatoma. Administration of hypertonic NaCl alone caused a decrease in [ 14 C]orotate incorporation into poly(A)-mRNA of host liver and hepatoma, whereas administration of tryptophan after hypertonic NaCl caused a significant improvement in host liver alone. Following the tryptophan administration, the activities of nuclear DNA-dependent RNA polymerases I and II, and of nuclear-envelope nucleoside triphosphatase, as well as labeled nuclear RNA release in vitro were slightly elevated in host liver but not in hepatoma. Tryptophan-related compounds, 5-hydroxy- dl -tryptophan, 5-fluorotryptophan, indole, and 3-hydroxyanthranilic acid, when administered in place of l -tryptophan, did not appreciably affect polyribosomal aggregation or protein synthesis in vitro in host liver or hepatoma.

Effect of tryptophan on the inhibitory action of selected hepatotoxic agents on hepatic protein synthesis

Abstract Tryptophan has been demonstrated earlier to induce a rapid stimulation of polyribosomal aggregation and protein synthesis in the livers of rats and mice. Also, tryptophan has been reported to have a corrective effect upon hepatic polyribosomes and protein synthesis when administered in conjunction with some hepatotoxic agents which inhibit protein synthesis by a variety of different mechanisms. This investigation expands the information in regard to whether the inhibitory effects on protein synthesis by a variety of hepatotoxic agents may be ameliorated by tryptophan administration and determines whether tryptophan may act to prevent, to cure, or both in regard to the inhibitory effect of each of the toxic agents. Under the short-term experiments conducted in this study, tryptophan was found to act in a preventive and curative manner with α-amanitin, cordycepin, puromycin, and hypertonic NaCl; only preventive with CCl4 and NaF; only curative with actinomycin D and ethionine; and neither with sparsomycin. Experiments were conducted to determine whether hepatic mRNA synthesis and nucleocytoplasmic translocation of RNA were affected by tryptophan in rats after treatment with selected hepatotoxic agents and the results revealed, in general, that the effects due to tryptophan occurred only when tryptophan acted in a curative manner. In a few experiments, tryptophan-related compounds, 5-hydroxy- dl -tryptophan, 5-fluorotryptophan, 3-hydroxyanthranilic acid, and indole, were tested, using equimolar amounts to l -tryptophan, with some hepatotoxic agents and in general, they caused some improvement in hepatic polyribosomal aggregation and protein synthesis but usually much less than with l -tryptophan.

Tryptophan-induced stimulation of hepatic ornithine decarboxylase activity in the rat

The effect of a single tube feeding of L-tryptophan on hepatic ornithine decarboxylase (ODC) activity in rats was investigated. The levels of ODC activity in the livers of control and experimental rats were assayed in vitro by measuring the release of 14CO2 from DL-[1-14C]ornithine. Single tube feedings of varying levels of L-tryptophan (2.5-30 mg/100 g body wt) to overnight-fasted rats 1 hr before sacrifice exhibited increases in the hepatic ODC activities. L-Tryptophan (30 mg/100 g body wt) tube fed to overnight-fasted rats 1/6 to 12 hr before sacrifice induced hepatic ODC activities which were significantly elevated beginning at 1 hr and peaking at 2 hr (6.5-fold increase over controls). In vitro [14C]leucine incorporation into protein using hepatic microsomes of tryptophan-treated rats was significantly increased at 1 hr in comparison with that of controls. The tryptophan-induced stimulation of hepatic ODC activity was not affected by prior adrenalectomy but was abolished by pretreatment with cycloheximide. These studies demonstrate that a single feeding of L-tryptophan can significantly enhance in the rat the activity of ODC, a key enzyme in the biosynthesis of polyamines.

Effect of sparsomycin in vivo and in vitro on hepatic polyribosomes and protein synthesis

Abstract Sparsomycin (20 μg/20 g of body wt), when administered intraperitoneally to mice, induced marked disaggregation of hepatic polyribosomes and inhibited incorporation of [ 14 C]leucine into hepatic proteins by 90 per cent. However, addition of Sparsomycin in vitro to a cell-free amino acid incorporating system did not cause disaggregation of polyribosomes, yet inhibited the incorporation of [ 14 C]leucine into proteins by 80 per cent. In studies concerned with initiation of protein synthesis, it was observed that Sparsomycin inhibited the factor-dependent initiation of new polyphenylalanine chains, as well as factor-dependent binding of either phe- t RNA or met- t RNA to 40S ribosomal subunits. These findings are interpreted as suggesting that, in vivo , Sparsomycin disaggregates polyribosomes probably by interfering with the initiation process.

Effect of spermine on hepatic polyribosomes and protein synthesis in rats

Abstract The effect of a single administration of spermine (0.15 mmole/100 g body wt) on hepatic polyribosomes and protein synthesis of rats was investigated. The results revealed that there was a significant shift toward heavier aggregation of hepatic polyribosomes of the spermine-treated rats in comparison to control rats. In vitro [ 14 C]leucine incorporation into protein was significantly increased when using hepatic microsomes of experimental compared with control rats. The effects were present from 30 to 120 min after the administration of spermine. Doses of 0.0188 to 0.15 mmole spermine/100 g body wt tube-fed for 1 hr induced the effects. Only from 0.78 to 1.33% of the spermine administered by stomach tube was present within the liver after 1 hr. In assaying for nucleocytoplasmic translocation of RNA, nuclear envelope nucleoside triphosphatase activity became increased and the dialyzed cell sap stimulated in vitro release of labeled [ 14 C]orotate RNA from control nuclei when using livers of rats treated with spermine 10 min before killing compared to controls. The results of this study indicate that the oral administration of spermine has stimulatory effects on hepatic polyribosomes and protein synthesis.

Effect of Ethanol on Polyribosomes and Protein Synthesis of Transplantable Hepatomas and Host Livers of Rats

Summary The effect of ethanol administration on polyribosomes and in vitro protein synthesis of host liver and hepatoma was investigated. When Buffalo rats bearing hepatoma 5123 intrahepatically were tube-fed ethanol (0.75 g/100 g body wt as a 50% (v/v) solution in saline) 3 hr before killing, there was disaggregation of total polyribosomes and a decrease in in vitro protein synthesis ([14C]leucine incorporation into protein) in hepatoma but not in host liver in comparison with control groups. Evaluation of the disaggregation of the polyribosomes of the hepatoma of the ethanol-treated rats revealed that the membrane-bound fraction was affected more than the free fraction.

Effect of Hypertonic Sodium Chloride on Polyribosomes and Protein Synthesis of Kidneys of Rats

Abstract The effects of a single intraperitoneal administration of hypertonic NaCl (5.3 ml of 7.55% NaCl/100 g body wt) on polyribosomes and in vitro protein synthesis of kidneys of rats were investigated. The results revealed that there was marked disaggregation of kidney polyribosomes of rat (fasted or nonfasted) that received a single dose of hypertonic NaCl solution 30 min prior to sacrifice in comparison to control rats that received isotonic NaCl solution. In vitro [14C]leucine incorporation into protein was significantly decreased using either postmitochondrial supernatant or microsomes from kidneys of hypertonic NaCl-treated rats compared with control rats. Administration of a single dose of hypertonic NaCl to rats produced a significant increase in the activity of renal RNase in comparison to that of the control group. Assay of renal acid phosphatase in the particulate and soluble fractions revealed no changes in the activities in the control and experimental rats.