scholarly journals CGP 48664, a potent and specific S-adenosylmethionine decarboxylase inhibitor: effects on regulation and stability of the enzyme

1997 ◽  
Vol 322 (1) ◽  
pp. 297-302 ◽  
Author(s):  
Fredrik SVENSSON ◽  
Helmut METT ◽  
Lo PERSSON
Keyword(s):  
Potent Inhibitor ◽  
Therapeutic Agents ◽  
Cellular Systems ◽  
Polyamine Metabolism ◽  
Mrna Levels ◽  
Leukaemia Cell Line ◽  
Adenosylmethionine Decarboxylase ◽  
L1210 Cells ◽  
Polyamine Transport ◽  
Mouse Leukaemia

Mammalian S-adenosylmethionine decarboxylase (AdoMetDC) catalyses a regulatory important step in the biosynthesis of polyamines and is a potential target for therapeutic agents against various parasitic diseases and proliferative disorders. In the present study we examined the effects of a newly synthesized AdoMetDC inhibitor, 4-amidinoindan-1-one 2ƀ-amidinohydrazone (CGP 48664), on polyamine metabolism in the mouse leukaemia cell line L1210. Treatment of the cells with 2 ƁM CGP 48664 led to a depletion of cellular spermidine and spermine. The putrescine content, in contrast, was markedly increased. Cells seeded in the presence of the inhibitor showed a significant decrease in growth rate, which was fully reversed by the addition of 2 ƁM spermidine or 1 ƁM spermine. The syntheses of ornithine decarboxylase and AdoMetDC were greatly increased in cells treated with CGP 48664. These increases were not correlated with similar changes in the mRNA levels, indicating the involvement of a translational mechanism. CGP 48664 was demonstrated to be a very poor competitor of spermidine uptake in the L1210 cells. L1210 cells deficient in polyamine transport were as sensitive to the antiproliferative effect of the inhibitor as were the parental cells, indicating that CGP 48664 did not enter the cells by the polyamine transport system. In addition to inhibiting AdoMetDC, CGP 48664 stabilized the enzyme against degradation. In the present study we also demonstrated that aminoguanidine (AMG), which is frequently used in cellular systems to inhibit any action of serum polyamine oxidase, apparently inhibits AdoMetDC by an irreversible mechanism that markedly stabilizes the enzyme against proteolytic degradation. CGP 48664 and the parental compound methylglyoxal bis(guanylhydrazone), which is also a potent inhibitor of AdoMetDC, contain one or two AMG-like moieties; the importance of these residues in the inhibition of AdoMetDC is discussed.

scholarly journals Effect of 1-amino-oxy-3-aminopropane on polyamine metabolism and growth of L1210 cells

1989 ◽  
Vol 263 (1) ◽  
pp. 215-221 ◽  
Author(s):  
R Poulin ◽  
J A Secrist ◽  
A E Pegg
Keyword(s):  
Polyamine Metabolism ◽  
Reversible Inhibitor ◽  
Dependent Manner ◽  
Intact Cells ◽  
Adenosylmethionine Decarboxylase ◽  
L1210 Cells ◽  
Cell Extracts ◽  
Drug Concentrations ◽  
Dose Dependent

1-Amino-oxy-3-aminopropane (AOAP) was reported to inhibit several mammalian polyamine-biosynthetic enzymes in vitro, including ornithine decarboxylase (ODC) and S-adenosylmethionine decarboxylase (AdoMetDC) [Khomutov, Hyvönen, Karvonen, Kauppinen, Paalanen, Paulin, Eloranta, Pajula, Andersson & Pösö (1985) Biochem. Biophys. Res. Commun. 130, 596-602]. In order to clarify its mechanism of action in intact cells, the inhibitory properties of AOAP on the growth and polyamine metabolism of L1210 cells were compared with those seen in a variant subline (D-R cells) which overproduces ODC. As little as 20 microM-AOAP completely blocked proliferation of L1210 cells, and this effect was reversed by the concomitant addition of exogenous putrescine or spermidine. Growth of D-R cells was not affected by AOAP at concentrations up to 0.5 mM. There was no difference in the uptake of AOAP between the L1210 and the D-R cells. Exposure of L1210 or D-R cells to AOAP greatly decreased ODC activity in undialysed cell extracts, but did not decrease AdoMetDC. Activities of both enzymes were increased severalfold by AOAP treatment when activity was measured in dialysed extracts. Treatment with AOAP depleted intracellular putrescine and spermidine contents of L1210 cells, while inducing a massive accumulation of decarboxylated AdoMet. The 8-fold higher putrescine pool present in untreated D-R cells was depleted in a dose-dependent manner by AOAP, but a significant decrease in spermidine and accumulation of decarboxylated AdoMet required 10 times higher drug concentrations, and the changes were much less dramatic than in L1210 cells. These results indicate that in L1210 cells AOAP behaves primarily as a reversible inhibitor of ODC.

scholarly journals Overexpression of arginine decarboxylase in transgenic plants

1997 ◽  
Vol 325 (2) ◽  
pp. 331-337 ◽  
Author(s):  
Daniel BURTIN ◽  
Anthony J. MICHAEL
Keyword(s):  
Transgenic Plants ◽  
Arginine Decarboxylase ◽  
Polyamine Metabolism ◽  
Morphological Development ◽  
Polyamine Biosynthesis ◽  
Adenosylmethionine Decarboxylase ◽  
Two Generations ◽  
Key Enzyme ◽  
Polyamine Conjugate ◽  
And Control

The activity of arginine decarboxylase (ADC), a key enzyme in plant polyamine biosynthesis, was manipulated in two generations of transgenic tobacco plants. Second-generation transgenic plants overexpressing an oat ADC cDNA contained high levels of oat ADC transcript relative to tobacco ADC, possessed elevated ADC enzyme activity and accumulated 10–20-fold more agmatine, the direct product of ADC. In the presence of high levels of the precursor agmatine, no increase in the levels of the polyamines putrescine, spermidine and spermine was detected in the transgenic plants. Similarly, the activities of ornithine decarboxylase and S-adenosylmethionine decarboxylase were unchanged. No diversion of polyamine metabolism into the hydroxycinnamic acid–polyamine conjugate pool or into the tobacco alkaloid nicotine was detected. Activity of the catabolic enzyme diamine oxidase was the same in transgenic and control plants. The elevated ADC activity and agmatine production were subjected to a metabolic/physical block preventing increased, i.e. deregulated, polyamine accumulation. Overaccumulation of agmatine in the transgenic plants did not affect morphological development.

scholarly journals Inhibition of S-adenosylmethionine decarboxylase by inhibitor SAM486A connects polyamine metabolism with p53-Mdm2-Akt/protein kinase B regulation and apoptosis in neuroblastoma

2009 ◽  
Vol 8 (7) ◽  
pp. 2067-2075 ◽  
Author(s):  
Dana-Lynn T. Koomoa ◽  
Tamas Borsics ◽  
David J. Feith ◽  
Craig C. Coleman ◽  
Christopher J. Wallick ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Protein Kinase B ◽  
Polyamine Metabolism ◽  
Adenosylmethionine Decarboxylase

scholarly journals Role of the 5′-untranslated region of mRNA in the synthesis of S-adenosylmethionine decarboxylase and its regulation by spermine

1994 ◽  
Vol 302 (3) ◽  
pp. 765-772 ◽  
Author(s):  
L M Shantz ◽  
R Viswanath ◽  
A E Pegg
Keyword(s):  
Secondary Structure ◽  
Untranslated Region ◽  
Fold Increase ◽  
Negative Regulator ◽  
Mrna Levels ◽  
Coding Region ◽  
Polyamine Biosynthesis ◽  
Adenosylmethionine Decarboxylase ◽  
Spermine Synthase ◽  
Synthase Inhibitor

S-Adenosylmethionine decarboxylase (AdoMetDC), a rate-limiting enzyme in polyamine biosynthesis, is regulated by polyamines at the levels of both transcription and translation. Two unusual features of AdoMetDC mRNA are a long (320 nt) 5′-untranslated region (5′UTR), which is thought to contain extensive secondary structure, and a short (15 nt) open reading frame (ORF) within the 5′UTR. We have studied the effects of altering these elements on both the expression of AdoMetDC and its regulation by n-butyl-1,3-diaminopropane (BDAP), a spermine synthase inhibitor. Human AdoMetDC cDNAs containing alterations in the 5′UTR, as well as chimaeric constructs in which the AdoMetDC 5′UTR was inserted ahead of the luciferase-coding region, were transfected into COS-7 cells. Construct pSAM320, which contains all of the 5′UTR, the AdoMetDC protein-coding region and the 3′UTR, was expressed poorly (2-fold over the endogenous activity). Deletion of virtually the entire 5′UTR, leaving nt -12 to -1, increased expression 59-fold, suggesting that 5′UTR acts as a negative regulator. The same effect was seen when the 27 nt at the extreme 5′ end were removed (pSAM293, 47-fold increase), or when the internal ORF which is present in this region was destroyed by changing the ATG to CGA (pSAM320-ATG, 38-fold increase). The expression and regulation of pSAM44 (made by deleting nt -288 to -12), which has very little predicted secondary strucutre, was very similar to that of pSAM320 indicating that the terminal 27 nt including the internal ORF rather than extensive secondary structure may be responsible for the low basal levels of AdoMetDC expression. These results, confirmed using luciferase constructs, suggest that the negative effect on expression is predominantly due to the internal ORF. Depletion of spermine by BDAP increased the expression from pSAM320 more than 5-fold without affecting AdoMetDC mRNA levels. Expression from pSAM293 was unchanged by spermine depletion, whereas that from pSAM320-ATG was increased 2.5-fold. These results indicate the presence of a spermine response element in the first 27 nt of the 5′UTR that may include but is not entirely due to the internal ORF.

scholarly journals Role of diamine oxidase during the treatment of tumour-bearing mice with combinations of polyamine anti-metabolites

1983 ◽  
Vol 212 (3) ◽  
pp. 895-898 ◽  
Author(s):  
A Kallio ◽  
J Jänne
Keyword(s):  
Diamine Oxidase ◽  
Potent Inhibitor ◽  
Specific Inhibitor ◽  
Inhibitory Effect ◽  
Tumour Cells ◽  
Adenosylmethionine Decarboxylase ◽  
Growth Inhibitory ◽  
Sequential Combination ◽  
Marked Accumulation

Treatment of mice bearing L1210 leukaemia with 2-difluoromethylornithine, a specific inhibitor of ornithine decarboxylase (EC 4.1.1.17), produced a profound depletion of putrescine and spermidine in the tumour cells. Sequential combination of methylglyoxal bis(guanylhydrazone), an inhibitor of adenosylmethionine decarboxylase (EC 4.1.1.50), with difluoromethylornithine largely reversed the polyamine depletion and led to a marked accumulation of cadaverine in the tumour cells. Experiments carried out with the combination of difluoromethylornithine and aminoguanidine, a potent inhibitor of diamine oxidase (EC 1.4.3.6), indicated that the methylglyoxal bis(guanylhydrazone)-induced reversal of polyamine depletion was mediated by the known inhibition of diamine oxidase by the diguanidine. In spite of the normalization of the tumour cell polyamine pattern upon administration of methylglyoxal bis(guanylhydrazone) to difluoromethylornithine-treated animals, the combination of these two drugs produced a growth-inhibitory effect not achievable with either of the compounds alone.

scholarly journals Effect of inhibitors of S-adenosylmethionine decarboxylase on the contents of ornithine decarboxylase and S-adenosylmethionine decarboxylase in L1210 cells

1988 ◽  
Vol 254 (1) ◽  
pp. 45-50 ◽  
Author(s):  
R Madhubala ◽  
J A Secrist ◽  
A E Pegg
Keyword(s):  
Ornithine Decarboxylase ◽  
Protein A ◽  
Protein Translation ◽  
Adenosylmethionine Decarboxylase ◽  
Enzyme Content ◽  
L1210 Cells ◽  
Mrna Content ◽  
Important Regulator ◽  
Substantial Decline ◽  
Effect Of Inhibitors

Treatment of L1210 cells with either of two inhibitors of S-adenosylmethionine decarboxylase (AdoMetDC), namely 5′-deoxy-5′-[N-methyl-N-[2-(amino-oxy)ethyl])aminoadenosine or 5′-deoxy-5′-[N-methyl-N-(3-hydrazinopropyl)]aminoadenosine, produced a large increase in the amount of ornithine decarboxylase (ODC) protein. The increased enzyme content was due to a decreased rate of degradation of the protein and to an increased rate of synthesis, but there was no change in its mRNA content. The inhibitors led to a substantial decline in the amounts of intracellular spermidine and spermine, but to a big increase in the amount of putrescine. These results indicate that the content of ODC is negatively regulated by spermidine and spermine at the levels of protein translation and turnover, but that putrescine is much less effective in bringing about this repression. Addition of either spermidine or spermine to the cells treated with the AdoMetDC inhibitors led to a decrease in ODC activity, indicating that either polyamine can bring about this effect, but spermidine produced effects at concentrations similar to those found in the control cells and appears to be the physiologically important regulator. The content of AdoMetDC protein (measured by radioimmunoassay) was also increased by these inhibitors, and a small increase in its mRNA content was observed, but this was insufficient to account for the increase in protein. A substantial stabilization of AdoMetDC occurred in these cells, contributing to the increased enzyme content, but an increase in the rate of translation cannot be ruled out.

scholarly journals Enantiomeric selectivity of adenosine transport systems in mouse erythrocytes and L1210 cells

1989 ◽  
Vol 263 (3) ◽  
pp. 957-960 ◽  
Author(s):  
W P Gati ◽  
L Dagnino ◽  
A R P Paterson
Keyword(s):  
Cell Types ◽  
Mirror Image ◽  
Nucleoside Transport ◽  
Transport Systems ◽  
Simple Diffusion ◽  
L1210 Cells ◽  
Marked Preference ◽  
Adenosine Transport ◽  
Mouse Leukaemia ◽  
Enantiomeric Selectivity

In mediating the entry of adenosine into mouse erythrocytes and mouse leukaemia L1210 cells, nucleoside transport systems were stereoselective, showing a marked preference for the D-enantiomer of adenosine (D-Ado). Inward zero-trans fluxes of the mirror-image isomer, L-adenosine (L-Ado), in those cells were slow relative to those of D-Ado. Contributing to L-Ado fluxes in both cell types were (i) a transporter-mediated process of high nitrobenzylthioinosine-sensitivity and (ii) simple diffusion.

scholarly journals Stable intracellular acidification upon polyamine depletion induced by α-difluoromethylornithine or N1,N12-bis(ethyl)spermine in L1210 leukaemia cells

1995 ◽  
Vol 312 (3) ◽  
pp. 749-756 ◽  
Author(s):  
R Poulin ◽  
A E Pegg
Keyword(s):  
Steady State ◽  
Acid Stress ◽  
Weak Acid ◽  
Transport Processes ◽  
Set Point ◽  
L1210 Cells ◽  
Acid Load ◽  
Mouse Leukaemia ◽  
A Minor ◽  
Steady State Control

Polyamines play major roles in ionic and osmotic regulation, but their exact involvement in specific ion transport processes is poorly defined. Treatment of L1210 mouse leukaemia cells with either 5 mM alpha-difluoromethylornithine (DFMO), a suicide substrate of ornithine decarboxylase, or 25 microM N1,N12-bis(ethyl)spermine (BE-3-4-3), a dysfunctional polyamine analogue, caused a stable decreased in intracellular pH (pHi) by 0.1-0.4 unit from steady-state control values between 7.4 and 7.6, as measured either by partition of a weak acid or with a fluorescent pH-sensitive probe. This effect was not related to cell growth status or differences in metabolic acid generation, and was observed in either the presence or absence of HCO3-. Exogenous spermidine (10-25 microM) or putrescine (25-50 microM) fully reversed DFMO- or BE-3-4-3-induced acidification within 2 and 8 h respectively. Recovery of pHi in L1210 cells after a nigericin- or NH4(+)-mediated acid load in HCO3(-)-free buffers was mediated by Na+/H+ antiporter activity, in addition to a minor Na(+)-independent and amiloride-insensitive pathway. Decreased steady-state pHi was maintained in polyamine-depleted L1210 cells after recovery from acid stress. Moreover, the pHi-dependence of the rate of Na(+)-dependent H+ extrusion after an acid stress was altered by DFMO and BE-3-4-3, resulting in a set-point which was lower by 0.25-0.30 pH unit in polyamine-depleted cells. On the other hand, neither the rate nor the magnitude of Na+/H(+)-exchanger-mediated alkalinization induced by hypertonic shock was decreased by polyamine depletion. Thus polyamine depletion induces a persistent defect in pHi homeostasis which is due, at least in part, to a stable decrease in the pHi set-point of the Na+/H+ exchanger.

Polyamine homoeostasis

2009 ◽  
Vol 46 ◽  
pp. 11-24 ◽  
Author(s):  
Lo Persson
Keyword(s):  
Mammalian Cells ◽  
Cell Function ◽  
Protein S ◽  
26S Proteasome ◽  
Adenosylmethionine Decarboxylase ◽  
Specific Effects ◽  
Polyamine Transport ◽  
Polyamine Transporter ◽  
Reading Frames ◽  
Synthesis And Degradation

The polyamines are essential for a variety of functions in the mammalian cell. Although their specific effects have not been fully elucidated, it is clear that the cellular polyamines have to be kept within certain levels for normal cell function. Polyamine homoeostasis in mammalian cells is achieved by a complex network of regulatory mechanisms affecting synthesis and degradation, as well as membrane transport of polyamines. The two key enzymes in the polyamine biosynthetic pathway, ODC (ornithine decarboxylase) and AdoMetDC (S-adenosylmethionine decarboxylase), are strongly regulated by feedback mechanisms at several levels, including transcriptional, translational and post-translational. Some of these mechanisms have been shown to be truly unique and include upstream reading frames and ribosomal frameshifting, as well as ubiquitin-independent proteasomal degradation. SSAT (spermidine/spermine N1-acetyltransferase), which is a crucial enzyme for degradation and efflux of polyamines, is also highly regulated by polyamines. A cellular excess of polyamines rapidly induces SSAT, resulting in increased degradation/efflux of the polyamines. The polyamines appear to induce both transcription and translation of the SSAT mRNA. However, the major part of the polyamine-induced increase in SSAT is caused by a marked stabilization of the enzyme against degradation by the 26S proteasome. In addition, active transport of extracellular polyamines into the cell contributes to cellular polyamine homoeostasis. Depletion of cellular polyamines rapidly induces an increased uptake of exogenous polyamines, whereas an excess of polyamines down-regulates the polyamine transporter(s). However, the protein(s) involved in polyamine transport and the exact mechanisms by which the polyamines regulate the transporter(s) are not yet known.

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