Orthophosphate Turnover in the Extracellular and Intracellular Space of Mouse Liver

Till, U; Brox, D; Frunder, Horst

doi:10.1111/j.1432-1033.1969.tb00806.x

Cited by 9 publications

(3 citation statements)

References 10 publications

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“…Equilibration of intracellulrtr Pi with the y-P of ATP must be fast in the thyroid as it is in red cells [44] and liver cells [45]. It can be calculated that the y-P of ATP is renewed more than 6 times per minute in dog thyroid slices [46].…”

Section: Discussionmentioning

confidence: 99%

Action of Thyrotropin on Phosphate Incorporation into Thyroid Proteins in vitro

Lamy¹,

Dumont²

1974

European Journal of Biochemistry

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The kinetics of [32P]phosphate incorporation into dog thyroid slices, in vitro, has been studied.Phosphate penetration was saturable which suggests a mediated transport. This transport appeared to be the limiting step in the slow incorporation of [32P]phosphate in the trichloroacetic-acidsoluble fraction of the slices. The incorporation of [32P]phosphate in thyroid proteins was a fast process (half equilibrium with the trichloroacetic-acid-soluble fraction radioactivity in 30 min). About 50°/, of this incorporation took place in the nuclear proteins, 2501, of it in the histones (mostly in the arginine-rich histones). The turnover of protein P was fast in all classes of proteins.Thyrotropin, in vitro, often enhanced the incorporation of [32P]phosphate in the trichloroacetic-acid-soluble fraction and into total proteins of the slices. However, this effect was delayed and rather small. It is ascribed mainly to an enhancement of phosphate penetration into the cells.This suggests, that, in dog thyroid cells, most of protein phosphorylation is independent of thyrotropin and cyclic AMP. Thyrotropin markedly enhanced the phosphorylation of Fl (lysine-rich) histones. This effect was fast, specific and mimicked by dibutyryl cyclic AMP; it required higher concentrations of thyrotropin than the activation of iodide binding to proteins. The role pf this action of thyrotropin is discussed; the hypothesis is proposed that it is related to thyrotropin-induced growth of thyroid tissue. NaF (5 mM) enhanced total protein phosphorylation in the thyroid slices. This effect, which may be due to unspecific inhibition of protein phosphatases, may explain the fact that NaF mimics several effects of thyrotropin. Cyclic-AMP-activated protein kinases have been demonstrated in thyroid tissue [3,7-121. To investigate the role of cyclic-AMP-induced protein phosphorylation in the activation of thyroid by thyrotropin, two approaches may be considered: the study of the enzymes involved (kinases and phosphatases) and the study of the substrate. Whereas cyclic-AMP-activated protein kinases have Abbreviation. Cyclic AMP, adenosine 3': 5'-monophosphate. been much investigated, little is known about their natural substrates. I n intact rat liver cells, glucagon, dibutyryl cyclic AMP and cyclic AMP activate the phosphorypation of the lysine-rich histones (FI) [13,14] [23,24] are substrates in acellular systems. The purpose of this work is to define the nature of the phosphorylated proteins in the thyroid, the kinetics of these phosphorylations and their dependence on cyclic AMP and thyrotropin.Eur. J. Biochem. 45 (1974)

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Section: Discussionmentioning

confidence: 99%

Action of Thyrotropin on Phosphate Incorporation into Thyroid Proteins in vitro

Lamy¹,

Dumont²

1974

European Journal of Biochemistry

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“…The intracellular Pi level is closely related to uric acid production in liver by regulating purine nucleotide synthesis and degradation. The control step in purine nucleotide breakdown is catalyzed by adenosine monophosphate deaminase (35)(36)(37) and in purine synthesis by phosphoribosylpyrophosphate synthase (38). Both enzymes are inhibited by Pi at physiologic concentrations in vitro (36)(37)(38).…”

Section: Interpretation Of P-31 Spectra the Phosphomonoester Peakmentioning

confidence: 99%

Study of Liver Metabolism in Glucose-6-Phosphatase Deficiency (Glycogen Storage Disease Type 1A) by P-31 Magnetic Resonance Spectroscopy

et al. 1988

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ABSTRACT. Liver metabolism of two patients (aged 15 and 23 yr) was studied by P-31 magnetic resonance spectroscopy at 1.9 tesla. The P-31 spectra of liver showed the resonances of phosphomonoesters (including sugar phosphates), inorganic phosphate (Pi), phosphodiesters (e.g. glycerophosphorylcholine, glycerophosporylethanolamine), and ATP. These resonances were quantified by expressing their peak areas in mM (assuming that ATP concentrations in normal liver is 2.5 mM) or as a ratio relative to the area of the phosphodiester resonance. After an overnight fast liver phosphomonoesters in patients were 2.6 and 1.6 AU, respectively (controls 1.1 rt 0.5, mean f 2 SD, n = 17). At the same time liver Pi was decreased in patients to 1.3 and 1.0, respectively (controls 1.8 f 0.8). Based on chemical shift measurements the increase in phosphomnnoesters could be attributed to accumulation of sugar phosphates (mainly glycolytic intermediates). After 1 g/kg oral glucose, hepatic sugar phosphates decreased in patients by 64 and 40%, respectively, and reached normal levels (on the absolute intensity scale); whereas liver Pi increased by 130 and 40%, respectively. Liver Pi levels remained elevated in both patients 30 min after ingestion of glucose. Liver sugar phosphates and Pi did not change in control subjects (n = 4) after glucose. In contrast to some previous reports, we have found accumulation of glycolytic intermediates in the liver of glucose-6-phosphatase-deficient patients during fasting. In these patients high levels may enhance the activity of residual glucose-6-phosphatase thus increasing hepatic glucose production and reducing the degree of hypoglycemia during fasting. Hyperuricemia is another serious complication of glucose-6-phosphatase deficiency and was present in both patients. Levels of Pi are known to regulate synthesis and breakdown purines. The changes in liver Pi during fasting and refeeding in these patients may stimulate production of uric acid and contribute to the hyperuricemia. (Pediatr Res 23: 375-380,1988) Glucose-6-phosphatase is necessary for the release of free glucose from glucose-6-P derived from glycogenolysis or gluconeogenesis. The link between glucose-6-phosphatase deficiency and its metabolic manifestations is not clear (1). Fasting hypoglycemia, a typical feature of glucose-6-phosphatase deficiency, can be accounted for by the block in glucose release. However, the striking ability of glucose-6-phosphatase-deficient patients to maintain from 30-100% of the normal glucose output during fasting has been difficult to explain in view of the reduction in hepatic glucose-6-phosphatase activity by more than 95% (2, 3) (Collins JE, Leonard JV, unpublished observations). A second clinical problem in glucose-6-phosphatase-deficient patients is hyperuricemia and gout (4-9). It has been suggested that the block in hepatic glucose release leads to accumulation of glucose-6-P during fasting. Glucose-6-P would overflow into the pentose phosphate shunt, increasing the formation of ribose-5-P and ultima...

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“…Next, for experimental reasons, not all specific radioactivities are available. PPi is compartmentalized [l], and the specific radioactivity of the intracellular liver Pi cannot be determined directly within the first min after intravenous tracer injection (see [5]). I n conse-quence of this a simplified flux model (Fig.2) was constructed for the mononucleotide interconversions.…”

Section: Udp-j3-p 10 20mentioning

confidence: 99%

Enzymic Flux Rates within the Mononucleotides of the Mouse Liver

Zahn

Klinger

Frunder

1969

European Journal of Biochemistry

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Within the first 30 sec after intravenous injection of [32P]Pi the P-flux was measured within the mononueleotide pool of the normal mouse liver. It is only in this short time interval that the dynamic phase of tracer kinetics is met, permitting proper calculations of flux rates of the mononucleotides with high turnover.The simplified model applied for the calculation of the stated flux rates describes fairly adequately the main pathways within the complex network of possible mononucleotide interconversions.The P-flux from the ,8-P of ADP to the p-P of ATP, catalyzed preferentially by oxidative and substrate phosphorylation amounts to 33 pmoles per g fresh liver per min. 12, 0.7, 0.3 and 0.01 pmoles y-P of ATP are transfered per g per min to the ,8-P of ADP, UDP, GDP and CDP, respectively, via the group nucleoside monophosphate kinases. These flux rates are 1-2 orders of magnitude less than expected from the kinetics behaviour of the involved enzymes in vitro. The reverse reaction of the nucleoside monophosphate kinases is probably small. 9 , 4 and 0.5 pmoles of the y-P of ATP are transferred per g per min to the y-P of GTP, UTP and CTP via nucleoside diphosphate kinase.I n a preceding paper [l] the turnover of glycolytic metabolites and mononucleotides was examined in the normal mouse liver 30-60sec after intravenous injection of carrier-free [32P]Pi. After this labelling time, however, the specific radioactivities of different P-positions of the mononucleotides were partly in the prestationary phase of tracer kinetics [2], and thus did not permit proper calculations of all the corresponding flux rates [l]. Therefore, the specific radioactivities of the ,8-and y -P positions of ATP, ADP, GTP, CTP and UTP were determined 10-30 sec after tracer injection. Evaluation of these data provides quantitative information on the main energy generating and consuming reactions in the liver under conditions in vivo. MATERIALS AND METHODSThe methods used for the intravenous administration of carrier-free [32P]Pi, the preparation of the livers and the determination of the specific radioactivities of the mononueleotides were exactly as described previously [3] with one exception. The frozen liver was deproteinized with 10 Ol0 trichloraeetie acid [3]) of the first separation procedure containing mainly fructose 1,6-biphosphate, phosphoenolpyruvate, CTP, ATP and UDP were treated with charcoal according to [3]. The charcoal eluent containing the nucleotides was concentrated in vacuo.

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Orthophosphate Turnover in the Extracellular and Intracellular Space of Mouse Liver

Cited by 9 publications

References 10 publications

Action of Thyrotropin on Phosphate Incorporation into Thyroid Proteins in vitro

Action of Thyrotropin on Phosphate Incorporation into Thyroid Proteins in vitro

Study of Liver Metabolism in Glucose-6-Phosphatase Deficiency (Glycogen Storage Disease Type 1A) by P-31 Magnetic Resonance Spectroscopy

Enzymic Flux Rates within the Mononucleotides of the Mouse Liver

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