Cell cycle regulation of purine synthesis by phosphoribosyl pyrophosphate and inorganic phosphate

2013 ◽  
Vol 454 (1) ◽  
pp. 91-99 ◽  
Author(s):  
Alla Fridman ◽  
Arindam Saha ◽  
Adriano Chan ◽  
Darren E. Casteel ◽  
Renate B. Pilz ◽  
...  
Keyword(s):  
De Novo ◽  
Fold Increase ◽  
Pentose Phosphate ◽  
S Phase ◽  
G1 Phase ◽  
Synthetase Activity ◽  
Phosphoribosyl Pyrophosphate ◽  
Purine Synthesis ◽  
Oxidative Pathway ◽  
Intracellular Phosphate

Cells must increase synthesis of purine nucleotides/deoxynucleotides before or during S-phase. We found that rates of purine synthesis via the de novo and salvage pathways increased 5.0- and 3.3-fold respectively, as cells progressed from mid-G1-phase to early S-phase. The increased purine synthesis could be attributed to a 3.2-fold increase in intracellular PRPP (5-phosphoribosyl-α-1-pyrophosphate), a rate-limiting substrate for de novo and salvage purine synthesis. PRPP can be produced by the oxidative and non-oxidative pentose phosphate pathways, and we found a 3.1-fold increase in flow through the non-oxidative pathway, with no change in oxidative pathway activity. Non-oxidative pentose phosphate pathway enzymes showed no change in activity, but PRPP synthetase is regulated by phosphate, and we found that phosphate uptake and total intracellular phosphate concentration increased significantly between mid-G1-phase and early S-phase. Over the same time period, PRPP synthetase activity increased 2.5-fold when assayed in the absence of added phosphate, making enzyme activity dependent on cellular phosphate at the time of extraction. We conclude that purine synthesis increases as cells progress from G1- to S-phase, and that the increase is from heightened PRPP synthetase activity due to increased intracellular phosphate.

scholarly journals Phosphoribosyl pyrophosphate and phosphoribosyl pyrophosphate synthetase in rat mammary gland. Changes in the lactation cycle and effects of diabetes, insulin and phenazine methosulphate

1986 ◽  
Vol 238 (2) ◽  
pp. 553-559 ◽  
Author(s):  
S Kunjara ◽  
M Sochor ◽  
N Salih ◽  
P McLean ◽  
A L Greenbaum
Keyword(s):  
Mammary Gland ◽  
Fold Increase ◽  
Pentose Phosphate ◽  
Diabetic Rat ◽  
Synthetase Activity ◽  
Phosphoribosyl Pyrophosphate ◽  
Rat Mammary Gland ◽  
Lactation Cycle ◽  
Phenazine Methosulphate

Changes in the tissue content of phosphoribosyl pyrophosphate (PPRibP), glucose 6-phosphate, ribose 5-phosphate (Rib5P), RNA and DNA, of the activity of PPRibP synthetase (EC 2.7.6.1) and the conversion of [1-14C]- and [6-14C]-glucose into 14CO2 were measured at mid-lactation in the normal and diabetic rat and in pregnancy, lactation and mammary involution in the normal rat. The PPRibP, glucose 6-phosphate and Rib5P contents increase during pregnancy and early lactation to reach a plateau value at mid-lactation, before falling sharply during weaning. The PPRibP content, PPRibP synthetase activity and flux of glucose through the oxidative pentose phosphate pathway (PPP) all change in parallel during the lactation cycle. Similarly, after 3 and 5 days duration of streptozotocin-induced diabetes, ending on day 10 of lactation, there were parallel declines in PPRibP content, PPRibP synthetase and PPP activity. The effect of streptozotocin was prevented by pretreatment with nicotinamide and partially reversed by insulin administration. Addition of insulin to lactating rat mammary-gland slices incubated in vitro significantly raised the PPRibP content (+47%) and the activity of the PPP (+40%); phenazine methosulphate, which gives a 2-fold increase in PPP activity, raised the PPRibP content of lactating mammary gland slices by approx. 3-fold. It is concluded that Rib5P, generated in the oxidative segment of the PPP, is an important determinant of PPRibP synthesis in the lactating rat mammary gland and that insulin plays a central role in the regulation of the bioavailability of this precursor of nucleotide and nucleic acid synthesis.

scholarly journals Metabolic interaction between amino acid deprivation and cisplatin synergistically reduces phosphoribosyl-pyrophosphate and augments cisplatin cytotoxicity

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Nisreen Wahwah ◽  
Debanjan Dhar ◽  
Hui Chen ◽  
Shunhui Zhuang ◽  
Adriano Chan ◽  
...  
Keyword(s):  
Amino Acid ◽  
Essential Amino Acid ◽  
De Novo ◽  
Synthetase Activity ◽  
Amino Acid Deprivation ◽  
Phosphoribosyl Pyrophosphate ◽  
Nucleotide Synthesis ◽  
Intracellular Phosphate ◽  
Dna Strand

AbstractCisplatin is a mainstay of cancer chemotherapy. It forms DNA adducts, thereby activating poly(ADP-ribose) polymerases (PARPs) to initiate DNA repair. The PARP substrate NAD+ is synthesized from 5-phosphoribose-1-pyrophosphate (PRPP), and we found that treating cells for 6 h with cisplatin reduced intracellular PRPP availability. The decrease in PRPP was likely from (1) increased PRPP consumption, because cisplatin increased protein PARylation and PARP1 shRNA knock-down returned PRPP towards normal, and (2) decreased intracellular phosphate, which down-regulated PRPP synthetase activity. Depriving cells of a single essential amino acid decreased PRPP synthetase activity with a half-life of ~ 8 h, and combining cisplatin and amino acid deprivation synergistically reduced intracellular PRPP. PRPP is a rate-limiting substrate for purine nucleotide synthesis, and cisplatin inhibited de novo purine synthesis and DNA synthesis, with amino acid deprivation augmenting cisplatin’s effects. Amino acid deprivation enhanced cisplatin’s cytotoxicity, increasing cellular apoptosis and DNA strand breaks in vitro, and intermittent deprivation of lysine combined with a sub-therapeutic dose of cisplatin inhibited growth of ectopic hepatomas in mice. Augmentation of cisplatin’s biochemical and cytotoxic effects by amino acid deprivation suggest that intermittent deprivation of an essential amino acid could allow dose reduction of cisplatin; this could reduce the drug’s side effects, and allow its use in cisplatin-resistant tumors.

scholarly journals Scc2-mediated loading of cohesin onto chromosomes in G1 yeast cells is insufficient to build cohesion during S phase

2017 ◽  
Author(s):  
Kim A Nasmyth
Keyword(s):  
Replication Fork ◽  
De Novo ◽  
S Phase ◽  
Yeast Cells ◽  
G1 Phase ◽  
Cohesin Complex ◽  
Previous Conclusion ◽  
Sister Chromatids ◽  
Chromatid Cohesion ◽  
Cohesin Subunit

SummarySister chromatids are held together from their replication until mitosis. Sister chromatid cohesion is mediated by the ring-shaped cohesin complex and it is thought that cohesin holds sister chromatids together by entrapping sister DNAs within the cohesin ring (Haering et al., 2008). However, how this occurs is not well understood. Because cohesin binds to DNA prior to replication it is possible that the replication fork passes through the lumen of the ring thereby placing replicated sisters inside cohesin rings. If this is the case, loading of cohesin in the G1 phase may be sufficient to build cohesion.We show here that Scc2, a cohesin subunit required for loading cohesin onto chromosomes de novo, is necessary for establishment of cohesion even after Scc2-mediated loading has already taken place during late G1 or early S phase. Our results challenge a previous conclusion based on related experiments whereby Scc2 was found not to be required for cohesion establishment during S phase (Lengronne et al., 2006).

scholarly journals Stimulation of pentose phosphate pathway dehydrogenase enzyme activities in ethionine-treated mice

1968 ◽  
Vol 106 (4) ◽  
pp. 769-776 ◽  
Author(s):  
Hsien-Gieh Sie ◽  
William H. Fishman
Keyword(s):  
Dehydrogenase Activity ◽  
Fold Increase ◽  
Pentose Phosphate ◽  
Hepatic Glycogen ◽  
Synthetase Activity ◽  
Phosphogluconate Dehydrogenase ◽  
Glycogen Synthetase ◽  
Period 2 ◽  
Marked Preference ◽  
Glucose 6 Phosphate Dehydrogenase

1. Mice treated with ethionine (intraperitoneally, 5mg./day for 4 days or 10mg./day for 3 days) showed a profound loss of hepatic glycogen, a decrease of glycogen synthetase activity, a development of hypoglycaemia, a two- to five-fold increase in the activity of glucose 6-phosphate dehydrogenase but no change in 6-phosphogluconate dehydrogenase and an earlier manifestation of the solubilization of phosphorylase as compared with glycogen synthetase. The administration of ATP did not prevent these effects. 2. During the early post-injection period (2–3 days) there was a further enhancement of the activity of glucose 6-phosphate dehydrogenase (tenfold) in the liver and a clear elevation of 6-phosphogluconate dehydrogenase activity (twofold). Subsequently, the glycogen concentration was restored, followed by an earlier reassociation of glycogen particle with phosphorylase than with glycogen synthetase, along with a disappearance of ethionine effect at about the eighteenth day. 3. Glucose 6-phosphate dehydrogenase from both control and ethionine-treated animals showed a marked preference for glucose 6-phosphate as substrate rather than for galactose 6-phosphate, whose rate of oxidation was only 10% of that of the glucose 6-phosphate. 4. Since actinomycin D, puromycin, 5-fluorouracil and dl-p-fluorophenylalanine failed to block the ethionine-enhanced glucose 6-phosphate dehydrogenase activity, the possibility that new enzyme protein synthesis is responsible for the effect is doubtful.

scholarly journals Improving the Production of Riboflavin by Introducing a Mutant Ribulose 5-Phosphate 3-Epimerase Gene in Bacillus Subtilis

2021 ◽  
Author(s):  
Bin Yang ◽  
Yiwen Sun ◽  
Shouying Fu ◽  
Miaomiao Xia ◽  
Chuan Liu ◽  
...  
Keyword(s):  
Bacillus Subtilis ◽  
Pentose Phosphate Pathway ◽  
De Novo ◽  
Fold Increase ◽  
Fermentation Medium ◽  
Pentose Phosphate ◽  
Oxidative Pentose Phosphate Pathway ◽  
Growth Defect ◽  
Riboflavin Production ◽  
Expression Of Genes

Abstract Background: Ribulose 5-phosphate (Ru5P) and guanosine 5'-triphosphate (GTP) are two key precursors of riboflavin, whereby Ru5P is also a precursor of GTP (purine de novo synthesis pathway). As a product of the oxidative pentose phosphate pathway (OPPP), Ru5P can flow back into the Embden-Meyerhof-Parnas pathway (EMP) through the non-oxidative pentose phosphate pathway (NOPPP). Major enzymes of the NOPPP include transaldolase (Tal), transketolase (Tkt), and ribulose 5-phosphate 3-epimerase (Rpe), which catalyze the conversion of Ru5P into xylulose 5-phosphate (X5P). Thus, inactivation of Rpe can reduce the consumption of Ru5P, enhancing the carbon flux toward riboflavin biosynthesis. However, there are no studies on the mutation or deletion of Rpe for improving riboflavin production. Results: We mutated the genomic copy of rpe, in the riboflavin-producing Bacillus subtilis BSLY. The resulting strain BSR produced 320.50±31.28 mg/L of riboflavin in a fermentation medium containing 40 g/L sucrose as the carbon source, representing a 5-fold increase over the parental strain. We also constructed an rpe nonsense mutation strain BSRN, which produced 365.69±27.53 mg/L riboflavin in fermentation medium. However, BSRN had a growth defect compared with BSR and BSLY, which was also present in media containing glucose, maltose, and fructose. The growth of BSRN was restored when mutant or wild-type rpe was overexpressed. Notably, BSR exhibited respective increases of the inosine and guanine titers by 163% and 40%. Furthermore, a 35.96-fold increase of inosine 5’-monophosphate (IMP) titer was detected in BSRN. The transcription levels of most OPPP, purine, and GTP synthesis genes were unchanged in BSR, except for the levels of zwf and ndk, which were respectively 49% lower and 310% higher than in BSLY. The production of riboflavin was increased to 479.90±33.21 mg/L when ribA was expressed in BSR. The expression of zwf, gntZ, prs, and purF, respectively enhanced the riboflavin production by 48%, 31%, 26%, and 35%. Finally, overexpression of the rib operon by the pMX45 plasmid and mutant gnd by pHP03 plasmid in BSR led to a 3.05-fold increase of the riboflavin production (977.29±63.44 mg/L).Conclusions: This paper describes a mutation of the ribulose 5-phosphate 3-epimerase and its influence on riboflavin production. The results of qRT-PCR and HPLC analyses indicated that the rpe mutant showed a different pattern of purine metabolism, while the cells maintained generally normal levels of the transcription of genes related to PPP and purine de novel synthesis pathways. With the increased expression of genes in the OPPP, purine, and riboflavin synthesis pathways, the production of riboflavin was effectively enhanced, showing the potential for further engineering of this strain.

scholarly journals Metabolic supervision by PPIP5K, an inositol pyrophosphate kinase/phosphatase, controls proliferation of the HCT116 tumor cell line

2021 ◽  
Vol 118 (10) ◽  
pp. e2020187118
Author(s):  
Chunfang Gu ◽  
Juan Liu ◽  
Xiaojing Liu ◽  
Haibo Zhang ◽  
Ji Luo ◽  
...  
Keyword(s):  
Tumor Cell ◽  
Cell Line ◽  
De Novo ◽  
Adenine Nucleotide ◽  
Hct116 Cell ◽  
Tumor Cell Line ◽  
Pentose Phosphate ◽  
Metabolic Reprogramming ◽  
Phosphoribosyl Pyrophosphate ◽  
Inositol Pyrophosphate

Identification of common patterns of cancer metabolic reprogramming could assist the development of new therapeutic strategies. Recent attention in this field has focused on identifying and targeting signal transduction pathways that interface directly with major metabolic control processes. In the current study we demonstrate the importance of signaling by the diphosphoinositol pentakisphosphate kinases (PPIP5Ks) to the metabolism and proliferation of the HCT116 colonic tumor cell line. We observed reciprocal cross talk between PPIP5K catalytic activity and glucose metabolism, and we show that CRISPR-mediated PPIP5K deletion suppresses HCT116 cell proliferation in glucose-limited culture conditions that mimic the tumor cell microenvironment. We conducted detailed, global metabolomic analyses of wild-type and PPIP5K knockout (KO) cells by measuring both steady-state metabolite levels and by performing isotope tracing experiments. We attribute the growth-impaired phenotype to a specific reduction in the supply of precursor material for de novo nucleotide biosynthesis from the one carbon serine/glycine pathway and the pentose phosphate pathway. We identify two enzymatic control points that are inhibited in the PPIP5K KO cells: serine hydroxymethyltransferase and phosphoribosyl pyrophosphate synthetase, a known downstream target of AMP-regulated protein kinase, which we show is noncanonically activated independently of adenine nucleotide status. Finally, we show the proliferative defect in PPIP5K KO cells can be significantly rescued either by addition of inosine monophosphate or a nucleoside mixture or by stable expression of PPIP5K activity. Overall, our data describe multiple, far-reaching metabolic consequences for metabolic supervision by PPIP5Ks in a tumor cell line.

scholarly journals Enzymes of the pathway of purine synthesis in the rat mammary gland. Changes in the lactation cycle and the effects of diabetes

1988 ◽  
Vol 250 (2) ◽  
pp. 395-399 ◽  
Author(s):  
S Beardsley ◽  
S Kunjara ◽  
A L Greenbaum
Keyword(s):  
Mammary Gland ◽  
De Novo ◽  
Salvage Pathway ◽  
Adenine Phosphoribosyltransferase ◽  
Phosphoribosyl Pyrophosphate ◽  
Purine Synthesis ◽  
Enzyme Change ◽  
Rat Mammary Gland ◽  
Salvage Pathways ◽  
Lactation Cycle

Measurements were made of the activities of the enzymes of the ‘de novo’ and salvage pathways of purine synthesis [phosphoribosyl pyrophosphate amidotransferase (EC 2.4.2.14), adenine phosphoribosyltransferase (EC 2.4.2.7) and hypoxanthine phosphoribosyltranferase (EC 2.4.2.8)] at different stages of the lactation cycle, and the effects of diabetes on the activity of these enzymes in lactation were studied. A distinctive pattern of enzyme change was observed, in which the ‘de novo’ pathway enzyme phosphoribosyl pyrophosphate amidotransferase increased sharply between days 10 and 14 of pregnancy, and then remained sensibly constant until the height of lactation, whereas the enzymes of the salvage pathway increased later in pregnancy and continued to rise during lactation. Diabetes severely depressed the activity of the enzymes of the salvage pathway, but appeared to be without effect on the ‘de novo’ pathway enzyme. These results are discussed in relation to the provision of purine precursors from tissues outside the mammary gland.

scholarly journals Concentration of phosphoribosyl pyrophosphate in the kidney during development and in experimental diabetic hypertrophy

1986 ◽  
Vol 234 (3) ◽  
pp. 579-585 ◽  
Author(s):  
S Kunjara ◽  
M Sochor ◽  
A Adeoya ◽  
P McLean ◽  
A L Greenbaum
Keyword(s):  
De Novo ◽  
Pentose Phosphate ◽  
Short Term ◽  
Phosphoribosyl Pyrophosphate ◽  
Nucleotide Synthesis ◽  
Immature Rat ◽  
Adult Rat ◽  
Developmental Growth ◽  
Kidney Hypertrophy ◽  
Salvage Pathways

The effect of developmental growth on the kidney content of phosphoribosyl pyrophosphate PPRibP was studied in rats at ages between the foetal animal and up to 100 days of age. In addition, the effect of short-term diabetes (up to 14 days) on the renal content of PPRibP was studied in immature rats and in adults aged approx. 60 days. The developmental pattern of PPRibP is such that the PPRibP content is lowest in the young rat and increases as the rate of kidney growth slows. In the adult rat, the early kidney hypertrophy of diabetes is accompanied by a fall in PPRibP content and, again, the PPRibP content returns to normal as the rate of kidney hypertrophy diminishes. Induction of diabetes in the immature rat causes a lesser degree of kidney hypertrophy and also a smaller depression of renal PPRibP content. The activity of PPRibP synthetase (EC 2.7.6.1) is not significantly affected by age or diabetes. The changes in PPRibP content are discussed in relation to the generation of ribose 5-phosphate by the pentose phosphate pathway and the utilization of PPRibP for nucleotide synthesis via the ‘de novo’ and salvage pathways.

Purine synthesis de novo and its regulation in rat hepatocytes

1980 ◽  
Vol 58 (8) ◽  
pp. 599-606 ◽  
Author(s):  
Christine Des Rosiers ◽  
Marcel Lalanne ◽  
Joan Willemot
Keyword(s):  
Pentose Phosphate Pathway ◽  
De Novo ◽  
Rat Hepatocytes ◽  
Isolated Hepatocytes ◽  
Pentose Phosphate ◽  
Male Rats ◽  
P Availability ◽  
Prolonged Incubation ◽  
Purine Synthesis ◽  
Stimulation Of

Purine synthesis de novo and its regulation were studied in freshly isolated hepatocytes from fed adult male rats. These cells incorporated [14C]formate mainly into purine ribonucleotides. The immediate effect of increasing the concentration of inorganic phosphate in the incubation medium was an increase in 5-phosphoribosyl 1-pyrophosphate (PP-ribose-P) availability and a stimulation of purine synthesis de novo. However, prolonged incubation of cells in 25 mM phosphate resulted in a decreased PP-ribose-P availability and purine synthesis de novo. Methylene blue and phenazine methosulfate decreased PP-ribose-P availability and purine synthesis de novo although they stimulated considerably the pentose phosphate pathway. In contrast, epinephrine and glucagon increased significantly PP-ribose-P availability and purine synthesis de novo, but they did not change the activity of the pentose phosphate pathway. These results show a relationship between PP-ribose-P availability and purine synthesis de novo in rat hepatocytes. They emphasize the complexity of the regulation of PP-ribose-P availability.

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