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IGFBP-3 mediates p53-induced apoptosis during serum starvation.

Insulin-like growth factor binding protein (IGFBP)-3, a p53-response gene, can induce apoptosis in an IGF-independent manner. Here we demonstrate that IGFBP-3 mediates p53-induced apoptosis during serum starvation using two foil neoplastic cell models: one which introduces p53 activity and one which eliminates it. We created a doxycycline-inducible p53 model from the p53-negative PC-3 prostate cancer cell line. Doxycycline treatment increased both p53 and IGFBP-3 levels. It also augmented apoptosis, but not during insulin-like growth factor-I co-treatment. In a second model, lung carcinoma H460 cells expressing fully functional p53 were stably transfected with E6, which targets p53 for degradation. H460-E6 cells contained less p53 and IGFBP-3 than control neo-transfected cells, and proteasome blockade restored both. In serum deprivation, H460-E6 cells had enhanced growth and less apoptosis than did H460-neo cells. Reductions in H460-neo apoptosis, comparable in magnitude to H460-E6, were achieved by adding anti-IGFBP-3-antibody or IGFBP-3 antisense oligomers, but not non-specific immunoglobulin or IGFBP-3 sense oligomers. In summary, turning p53 in two foil neoplastic cell models induced IGFBP-3 expression and increased apoptosis during serum starvation, an effect inhibited by insulin-like growth factor-I treatment and specific IGFBP-3 blockade. This is the first demonstration of inhibition of p53 action by antagonizing IGFBP-3.

Apoptosis↗

[Activation of the expression of the microcin C51 operon upon glucose starvation of cells at the exponential growth phase].

It was earlier shown that expression of the microcin C51 operon in Escherichia coli cells is activated upon decelerated growth of cells during their transition to the stationary growth phase and depends on the sigmaS subunit of RNA polymerase. Using a single-copy construct containing the cloned promoter region of the microcin C51 operon and a promoterless lac operon (P(mcc)-lac), it was shown that the promoter of the microcin operon was also induced by stress caused by the transition of cells at the exponential growth phase into the medium without glucose as a sole carbon source. Activation of P(mcc)-lac expression upon severe glucose starvation occurred in rpoS+ and rpoS- strains. In cells carrying the rpoD800 mutation that renders the sigma70 subunit of RNA polymerase temperature-sensitive, an activation of P(mcc)-lac expression was observed at nonpermissive temperature, in contrast to its complete inhibition in E. coli cells at the phase of delayed growth. Other stressors-nitrogen starvation, high temperatures, osmotic shock, tetracycline and chloramphenicol-did not activate P(mcc)-lac expression in cells at the exponential growth phase.

Bacteriocins↗

[Survival and antioxidant defence of the yeast Saccharomyces cerevisiae during starvation and oxidative stress].

The role of catalase in response of the yeast Saccharomyces cerevisiae to oxidative stress induced by hydrogen peroxide under starvation was investigated. It was shown that under conditions used in this study 0.5 mM H2O2 did not change the number of viable cells in the wild strain YPH250, but this parameter was decreased by 15% in the acatalsaemic strain YWT1. Cells treatment with 0.5 mM H2O2 for 30 min did not modify the levels of carbonyl proteins in the parental strain, but caused its 1.4-fold increase in the defective strain. The observed 1.5-fold activation of catalase in the wild strain cells in response to H2O2-stress suggests that under starvation conditions catalase can be involved in the yeast cell protection, particularly they can prevent oxidative modification of some antioxidant and associated enzymes.

Adaptation, Physiological↗

Low-energy protein diet and starvation diet in the obese--effect on energy metabolism.

38 obese patients with BMI in excess of 35 were monitored, the patients were divided into four groups. The first three were on a strict slimming diet which different as to the energy values (1.38 MJ--2.75 MJ) and the amount of protein (5.58 g N--17.91 g N) per 24 hours. The fourth group was on a starvation diet. Metabolic balance values and resting energy cut put were measured by indirect calorimetry during 16 days of monitoring. In all groups the body weight declined markedly by an average of 8.2-10.5 kg. Resting energy consumption dropped only during absolute fasting. The starvation diet patients exhibited a negative nitrogen balance. Positive nitrogen balance was found in groups on 11.2 g N and 17.91 g N/24 hours. Fat utilization increased and sugar utilization declined in all groups. Protein catabolism declined markedly only in the fasting patients. Summed up, the low-energy protein diet is--in comparison with absolute diet--a more physiological way to achieving weight reduction, in particular, because, despite the loss of weight, a positive weight balance is achieved, too.

Adult↗

[Decreased synthetic activity as a possible cause of the death of Escherichia coli bacteria during amino acid starvation].

The work is concerned with studying the breakdown of proteins and RNA when a polyauxotrophic Escherichia coli strain is incubated in a salt solution without amino acids, phosphorus, nitrogen and glucose at 43 degrees C as well as the ability of starving bacterial cells to recommence protein and RNA synthesis (also in the course of phage T4 infection) and to reproduce bacteriophages T4, lambda and MS2. Within the first two hours of the incubation, 12% of proteins and 40% of RNA break down to acid-soluble fragments. Then protein degradation stops while RNA decomposition goes on, but at a lower rate. Within 4-6 h of starvation, the rate of protein and RNA synthesis drops down 4-5 times and the survival rate equals 40-60% when the cells are transferred onto a complete medium. The quantitative characteristics of phages T4, lambda and MS2 reproduction fall down in prestarved cells. The authors speculate that E. coli cells die off in the course of starvation not because some unique structure is destroyed, but owing to the fact that the activity of enzymes and ribosomes gradually declines. As a result, the synthetic activity of the cell drops down abruptly and irreversibly because the enzymes are inactivated and RNA breaks down, which eventually causes cell death.

Amino Acids↗

[Physiologic significance of "stringent control" in Escherichia coli under extreme starvation].

The viability of three isogenic relA+/relA strain pairs of Escherichia coli (CP78/CP79; NF161/NF162; CP107/CP143) was studied during prolonged starvation for amino acids, glucose or phosphate. After amino acid limitation we found a prolonged viability of all relA+ strains which synthesized ppGpp. We suggest that some ppGpp-mediated pleiotropic effects of the stringent response (e.g. glykogen accumulation, enhanced protein turnover) might be involved in this prolongation of survival. After glucose or phosphate starvation there was no difference in the relA+/relA strains either in the ppGpp content or in the survival.

Escherichia coli↗

Effect of Schistosoma mansoni infection, starvation and molluscicides on acid phosphate, transaminases and total protein in tissues and hemolymph of Biomphalaria alexandrina.

The activities of GOT and GPT in the hemolymph of B. alexandrina were significantly decreased by S. mansoni infection. However, the total protein concentration and AcP activity were increased. Although the snail starvation decreased AcP activity in the ovotestis, it increased GOT activity in the other organs of the snails. On the other hand, the snail feeding after starvation increased significantly AcP activity in ovotestis. Natural and synthetic molluscicides inhibited the activities of GOT and GPT, however, they increased the total protein concentrations and AcP activities in the examined organs.

Acid Phosphatase↗

Growth factors and hyperthermia. II. Viability of Chinese hamster ovary HA-1 cells during serum starvation and hyperthermia.

We tested the possibility that hyperthermia kills HA-1 cells in a manner analogous to growth factor deprivation. HA-1 cells were inactivated by serum starvation when incubated in Eagle's MEM at a density of 40 cells/cm2 or less. Cells became resistant to the absence of serum when the cell density was greater than 400 cells/cm2 or when lethally irradiated HA-1 feeder cells were present. The feeder cells exerted their effect through a diffusible factor. In addition, a 1:1 mixture of Eagle's MEM and Ham's F-12 enabled HA-1 cells to remain viable without serum. Ten days growth in Eagle's MEM + Ham's F-12 without serum resulted in the formation of microcolonies of cells. This indicated that growth factor deprivation was not lethal to HA-1 cells, and it suggested that they may have been partially transformed. The presence of the growth factors insulin, transferrin, and fibroblast growth factor (FGF) reduced cell killing by a small amount during conditions of serum starvation. After hyperthermia, the presence of growth factors again diminished cell killing by a modest amount (approximately twofold). Feeder cells also improved cell survival after hyperthermia. The effect of feeder cells was greatest when cells were trypsinized immediately after hyperthermia. When cells were not trypsinized after heating, feeder cells increased survival less than twofold. In summary, the absence of growth factors was not lethal to HA-1 cells, and therefore the cytotoxic effects of hyperthermia could not be explained fully by the failure to bind growth factors. HA-1 feeder cells secreted undefined, growth-promoting substances, but feeder cells exerted only a small positive effect on cell survival after hyperthermia when cells were not trypsinized after heating.

Animals↗

Serum protein changes in cafeteria mice induced by starvation.

Serum protein changes in cafeteria and control mice induced by starvation have been studied. Animals were subjected to food deprivation at 0, 3, 6, 9, 12, 18, 24 or 36 hours. Results show a more stabilized situation in cafeteria mice than controls particularly in protein metabolism. Serum protein composition changed very little during starvation, suggests a lower protein and amino acid catabolism induced by the high adaptation to consume lipids.

Animals↗

Evidence for new factors in the coordinate regulation of energy metabolism in Escherichia coli. Effects of hypoxia, chloramphenicol succinate, and 2,4-dinitrophenol on glucose utilization, glycogen synthesis, adenylate energy charge, and hexose phosphates during the first two periods of nitrogen starvation.

We studied the effects of decreased aeration, chloramphenicol succinate, and 2,4-dinitrophenol on the cellular rates of glycogen synthesis and glucose utilization and on the cellular concentrations of adenine nucleotides, glucose 6-phosphate, fructose 1,6-diphosphate, and phosphoenolpyruvate during the first two periods of nitrogen starvation of Escherichia coli W4597(K). A quantitative relationship between the changes in the rates and the accompanying changes in the hexose phosphates is demonstrated. However, the relationship for glycogen synthesis is different in different sets of metabolic conditions. We suggest that this difference reflects a change in the steady state level of a previously unknown effector of ADP-glucose synthetase (glucose 1-phosphate adenylyltransferase, EC 2.7.7.27) the rate-limiting enzyme of bacterial glycogen synthesis. We show that the properties of the hypothetical in vivo effector are consistent with the inhibitory effects of ppGpp (guanosine 3'-diphosphate 5'-diphosphate) and pppGpp (guanosine 3'-diphosphate 5'-triphosphate) on this enzyme in vitro. In addition, tetracycline, an inhibitor of the synthesis of these nucleotides, apparently prevents the change in the quantitative relationship. The relationship between glucose utilization and the hexose phosphates is altered at the transition to Period II of nitrogen starvation. We propose that this change reflects the alteration of the cellular steady state level of an unknown effector of the glucose phosphotransferase system. In contrast to the ATP-hexose phosphate system of shared regulatory effects, the specific effects of the unknown effectors allow the rates of glucose utilization and glycogen synthesis to be altered independently of each other and independently of changes in the rate of glycolysis. This independence allows a greater latitude of response for the individual pathways in more severe metabolic stress or in accommodating the metabolic changes necessary for long term survival.

Adenine Nucleotides↗

Differential effects of ellipticine and aza-analogue derivatives on cell cycle progression and survival of BALB/c 3T3 cells released from serum starvation or thymidine double block.

10-[Diethylaminopropylamino]-6-methyl-5H-pyrido[3',4':4,5] pyrrolo[2,3-g]isoquinoline (BD-40) (NSC-327471D) is an aza-ellipticine derivative with a promising antitumor activity (M. Marty, C. Jasmin, P. Pouillard, C. Gisselbrecht, G. Gouvenia, and H. Magdalainat, 17th Annual Meeting of the American Society of Clinical Oncology, C-108, 1981) and less toxicity than ellipticine. We have compared the effects of ellipticine, several of its analogues, and two aza-analogue ellipticine derivatives (BD-40 and BR-1376) on cell cycle progression of BALB/c 3T3 mouse cells under different growth conditions. Both drug series were found to stop cell growth and block cells in G2 phase in exponentially growing cultures and cultures released from a thymidine double block. Long-term viability of these cells was completely suppressed after a short exposure to the drugs. In contrast, while ellipticine and its derivatives caused identical effects in cells recovering from serum starvation, BD-40 and BR-1376 did not block cells in G2 phase and did not prevent the completion of the first division round occurring after serum addition to quiescent cells. This transient refractory state was accompanied by a total conservation of long-term viability of these cells at least for the next 6 h following serum and drug addition. This lack of effect was not related to an impaired drug uptake by cells recovering from serum starvation or by a dramatic change in drug distribution inside the cells. These results indicate that the nitrogen substitution in the ellipticine heterocycle is an important if not unique feature for the particular effect of the aza-analogues of ellipticine. Furthermore, they suggest that, in contrast to ellipticine derivatives, these compounds require an activation step before exhibiting cytotoxicity.

Alkaloids↗

Influence of starvation on the intra-lysosomal proteolysis in rat liver.

Starvation induced changes in the intralysosomal proteolysis in rat liver were assessed in terms of the degradation of intravenously administered [131I]-human serum albumin 30 min after injection. Fasting for five days resulted in nearly 11% increase in the endocytic uptake of the labeled protein in lysosome rich fraction. However, the rate of degradation of internalized protein measured in terms of TCA soluble products showed a distinct decline in starved animals as compared to fed controls. The observed decrease in proteolysis was reversed completely by refeeding the starved rats for 10 days. The restoration of the degradation profiles in fasted animals was also accomplished by isolating lysosomes at a post injection period of 90 min. The results indicated that the starvation induced decrease in proteolysis was a consequence of delayed fusion of lysosomes and the phagosome containing the labeled protein.

Animals↗

Proteinase pattern in Trametes versicolor in response to carbon and nitrogen starvation.

In stationary cultures of Trametes versicolor seven proteinase bands were revealed by electrophoresis in mycelium and five in the medium. Under conditions of nitrogen starvation the number of bands in mycelium was unchanged whereas one extracellular proteinase was missing. In the case of carbon starvation one new intracellular proteinase activity appeared and one extracellular activity disappeared. Moreover, in all starved cultures distinct differences in the intensity of particular bands were observed.

Basidiomycota↗

Cysteine starvation, isoleucyl-tRNAIle, and the regulation of the ilvGEDA operon of Escherichia coli.

The involvement of undermodified tRNA in the regulation of the ilvGEDA operon has been investigated using Escherichia coli C6, a relA-, Cys-, Met- mutant. This strain accumulates thionucleotide-deficient or methyl-deficient tRNA when starved for cysteine or methionine, respectively. The levels of threonine deaminase, the ilvA gene product, and transaminase B, the ilvE gene product, were both lower in cysteine-starved cells, as compared with either growing or methionine-starved cultures. When cysteine was added to cysteine-starved cells, growth ensued promptly and both enzyme activities returned to control levels. Treatment of recovering cultures with valine limited growth by isoleucine limitation, but did not cause a derepression of the ilvGEDA operon. Valine treatment of nonstarved or methionine-starved cells led to the expected increase in threonine deaminase and transaminase B activities. Cysteine-starved cells slowly regained the ability to derepress the operon after 3 h of recovery in complete medium. In contrast, the induction of the lac operon was normal in cysteine-starved cultures, even in the presence of valine. The loss of derepressibility of the ilvGEDA operon was correlated with the presence of a kinetically and chromatographically altered tRNAIle in cysteine-starved cells. No changes in tRNAIle were observed after methionine starvation. Using the periodate method, we found that the charging of tRNAIle increased from the normal level of 60 to 80% or greater after starvation for cysteine. Under conditions where the ilvGEDA operon was fully derepressed in nonstarved cells, the charging of tRNAIle fell to 27%. Unexpectedly, nearly identical results were obtained with cysteine-starved cells after an identical derepression test. These results suggest that factors other than the aminoacylation state of tRNAIle may be important in the regulation of this operon. In particular, modifications to tRNA which involve cysteine may be necessary for controlling the expression of the ilvGEDA operon in E. coli.

Amino Acids↗

The effects of amino acid starvation on regulation of polypeptide chain initiation in Ehrlich ascites tumor cells.

The ratio of initiation of protein synthesis in Ehrlich ascites tumor cells in culture is reduced by over 60% in the absence of a single essential amino acid. Cell-free extracts prepared from control and amino acid-starved cells retain some of the translational characteristics of these cells and are able to form [40 S.Met-tRNAfMet] initiation complexes. Studies with inhibitors show that up to 63% of the translation directed by endogenous mRNAs in vitro depends on reinitiation of polypeptide chains. Amino acid starvation inhibits this activity, as well as protein synthesis due to in vitro polysomal run-off, by up to 75%. Analysis of [40 S.Met-tRNAfMet]initiation complexes formed in vitro on native 40 S subunits shows that amino acid starvation causes up to a 77% decrease in the concentration of these complexes relative to the corresponding fed controls. This difference is eliminated by the addition of highly purified eukaryotic initiation factor eIF-2. Factor eIF-3 also stimulates [40 S.Met-tRNAfMet] formation in the cell extracts but does not abolish the difference between fed and starved preparations. Mixing experiments have not so far revealed any inhibitor of initiation complex formation in the starved cell extracts.

Amino Acids↗

[Starvation due to child neglect. Case report and aspects of expert testimony].

Case report on 2,5 years old monozygotic twins who died of starvation due to negligence. Different gradings of protein-energy-malnutrition are discussed, among them the Waterlow-classification of PEM. The advantage of the Waterlow-classification is that stunting and wasting can be distinguished. The Waterlow-classification is suitable also for the classification of fatal cases of starvation in childhood. Practical experiences with own cases and cases from the literature are presented.

Adipose Tissue↗

Guanosine tetraphosphate inhibits protein synthesis in vivo. A possible protective mechanism for starvation stress in Escherichia coli.

Guanosine 3',5'-bispyrophosphate (ppGpp) accumulates in bacteria in response to either amino acid or energy source starvation. Here we demonstrate that levels of ppGpp similar to those induced by amino acid starvation inhibit the rate of protein synthesis by 84-91%. The intracellular concentration of ppGpp is manipulated in our studies by increasing the expression of a truncated relA gene encoding a smaller but catalytically active peptide with ppGpp synthetase activity. We find that the intracellular activity of the truncated RelA peptide is insensitive to chloramphenicol, unlike the product of the wild-type relA gene, ppGpp synthetase I. Previously, this same ppGpp expression system was used (Schreiber, G., Metzger, S., Aizenman, E., Roza, S., Cashel, M., and Glaser, G. (1991) J. Biol. Chem. 226, 3760-3767) to demonstrate that increasing the ppGpp concentration inhibits growth and ribosomal RNA transcription, and they found suggestive evidence for ppGpp inhibition of protein synthesis. We further investigated the effect of ppGpp on protein synthesis and find that ppGpp is a potent inhibitor of protein synthesis as well as glycerol accumulation but has no effect on transport of methionine, the amino acid used in measuring protein synthesis rates, or on uptake of alpha-methylglucoside, a non-metabolizable analogue of glucose.

Biological Transport↗

Signaling phosphate starvation.

Phosphate starvation induces the transcription of several genes involved in phosphate metabolism in the budding yeast Saccharomyces cerevisiae. The signal transduction pathway that mediates this response consists of components that resemble those used to regulate the eukaryotic cell cycle; these include a cyclin-dependent kinase or CDK (Pho85), a cyclin (Pho80) and a CDK inhibitor (Pho81). The possibility that this pathway mediates cell-cycle responses to phosphate starvation is discussed.

Cyclin-Dependent Kinases↗