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D. L. Kramer J. T. Miller R. J. Bergeron R. Khomutov A. Khomutov C. W. Porter 《Journal of cellular physiology》1993,155(2):399-407
Regulation of polyamine transport in murine L1210 leukemia cells was characterized in order to better understand its relationship to specific intracellular polyamines and their analogs and to quantitate the sensitivity by which it is controlled. Up-regulation of polyamine uptake was evaluated following a 48-hr treatment with a combination of biosynthetic enzyme inhibitors to deplete intracellular polyamine pools. The latter declined gradually over 48 hr and was accompanied by a steady increase in spermidine (SPD) and spermine (SPM) transport as indicated by rises in Vmax to levels ~4.5 times higher than control values. Restoration of individual polyamine pools during a 6-hr period following inhibitor treatment revealed that SPD and SPM uptake could not be selectively affected by specific pool changes. The effectiveness of individual polyamines in reversing inhibitor-induced stimulation of uptake was as follows: putrescine < SPD < SPM = the SPM analog, N1, N12-bis(ethyl)spermine (BESPM). In contrast to stimulation of transport, down-regulation by exogenous polyamines or analogs occurred rapidly and in response to subtle increases in intracellular pools. Following a 1-hr exposure to 10 μM BESPM, Vmax values for SPD and SPM fell by 70%, whereas the analog pool increased to only 400–500 pmol/106 cells—about 15–20% of the total polyamine pool (~2.8 nmol/106 cells). SPM produced nearly identical regulatory effects on transport kinetics. Both BESPM and SPM were even more effective at down-regulating transport that had been previously stimulated four to fivefold by polyamine depletion achieved with enzyme inhibitors. A dose response with BESPM at 48 hr revealed a biphasic effect on uptake whereby concentrations of analog < 3 μM produced an increase in SPD and SPM Vmax values, whereas concentrations 3 μM and higher produced a marked suppression of these values. Cells treated with 3 μM BESPM for 2 hr and placed in analog-free medium recovered transport capability in only 3 hr. Thus, whereas stimulation of polyamine transport is a relatively insensitive and slowly responsive process that tends to parallel polyamine depletion, down-regulation of polyamine transport by exogenous polyamines and analogs and its reversal are rapidly responsive events that correlate with relatively small (i.e., 15–20%) changes in intracellular polyamine pools. 相似文献
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The role of polyamines in animal cell physiology 总被引:3,自引:0,他引:3
M H Goyns 《Journal of theoretical biology》1982,97(4):577-589
The ubiquitous distribution of polyamines in nature suggests that they fulfil some fundamental role(s) in living organisms. In animal cells, polyamine content closely parallels changes in the rate of cell proliferation so that the highest content is always observed in rapidly growing cells. The activity of ornithine decarboxylase (which is the first enzyme in the polyamine biosynthetic pathway) has been found to increase significantly in many systems shortly after exposure to hormones. Also, addition of polyamines greatly stimulates cell-free macromolecular synthesis. Observations such as these have suggested that polyamine accumulation stimulates cell growth and is important in the regulation of macromolecular biosynthesis. However, it is also possible to interpret such data as evidence that polyamine accumulation is the result, not the cause, of increased cell growth. This review supports the latter concept and re-examines the significance of the early induction of ornithine decarboxylase activity and of the stimulatory effects of exogenous polyamine on macromolecular synthesis. It is proposed that the polyamines are important only in maintaining cell growth that has already been stimulated by other factors and that their biosynthesis is to a large extent determined by the accumulation of RNA in the cell. 相似文献
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Calixto M. Protacio Hector E. Flores 《In vitro cellular & developmental biology. Plant》1992,28(2):81-86
Summary The involvement of free and conjugated polyamines in tuber formation was studied in in vitro cultured node explants ofSolanum tuberosum cv. Superior. Tubers developed from the axillary buds in 100% of the explants cultured in MS medium containing high sucrose
levels and supplemented with kinetin (Kin) and chlorocholine chloride (CCC). The addition of growth regulators was not essential
for tuber formation, although smaller tubers were formed in the medium devoid of Kin and CCC. Tuber formation was inhibited
in about 75% of node explants treated with 0.5 mM difluoromethylornithine (DFMO), a specific and irreversible inhibitor of ornithine decarboxylase. The inhibitory effect of
DFMO was almost completely reversed by putrescine addition. Addition of difluoromethylarginine (DFMA), the analogous inhibitor
of arginine decarboxylase, had no effect on tuber formation. DFMO, but not DFMA, also inhibited the development of axillary
buds into shoots in light-grown node explants. Aminooxyphenylpropionic acid (0.1 to 0.25 mM), an inhibitor of phenylalanine ammonia lyase, caused a sharp reduction in cinnamoyl putrescines, but had no effect on tuber
formation. Our results suggest that hydroxycinnamic acids are not causal in tuber formation but may serve as polyamine storage
pools. Our findings support the hypothesis that polyamines derived via the ornithine decarboxylase-mediated pathway are necessary
for tuber formation in vitro, probably at the early phase of morphogenesis involving active cell division. 相似文献
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Changes in polyamines (PAs) in cells and cultivation media of alfalfa (Medicago sativa L.) and tobacco bright yellow 2 (BY-2) (Nicotiana tabacum L.) cell suspension cultures were studied over their growth cycles. The total content of PAs (both free and conjugated forms) was nearly 10 times higher in alfalfa, with high level of free putrescine (Put) (in exponential growth phase it represented about 65-73% of the intracellular Put pool). In contrast, the high content of soluble Put conjugates was found in tobacco cells (in exponential phase about 70% of the intracellular Put). Marked differences occurred in the amount of PAs excreted into the cultivation medium: alfalfa cells excreted at the first day after inoculation 2117.0, 230.5, 29.0 and 88.0 nmol g(-1) of cell fresh weight (FW) of Put, spermidine (Spd), spermine (Spm) and cadaverine (Cad), respectively, while at the same time tobacco cells excreted only small amount of Put and Spd (12.7 and 2.4 nmol g(-1) FW, respectively). On day 1 the amounts of Put, Spd, Spm and Cad excreted by alfalfa cells represented 21, 38, 12 and 15% of the total pool (intra- plus extra-cellular contents) of Put, Spd, Spm and Cad, respectively. In the course of lag-phase and the beginning of exponential phase the relative contents of extracellular PAs continually decreased (with the exception of Cad). On day 10, the extracellular Put, Spd, Spm and Cad still represented 11.3, 10.9, 2.1 and 27% of their total pools. The extracellular PAs in tobacco cells represented from day 3 only 0.1% from their total pools. The possible role of PA excretion into the cultivation medium in maintenance of intracellular PA contents in the cells of the two cell culture systems, differing markedly in growth rate and PA metabolism is discussed. 相似文献
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Two different fluorescence cytochemical methods, the formaldehyde-fluorescamine (FF) method and the orthophthalaldehyde (OPT) method as well as an immunocytochemical method have been developed for the localization of spermidine and spermine. Of these three methods, the FF-method is the most easy to perform. We have studied the relationship between fluorescence intensity induced by the FF-method and cellular polyamine levels measured by HPLC in MCF-7 cells and HeLa cells. The experiments were designed to obtain different cell concentrations of polyamines. Cells grown on microscope slides in Petri-dishes were partly depleted of spermidine by two days inhibition of their ornithine decarboxylase activity using alpha-difluoromethylornithine. One hr before harvest the cells were exposed to different concentrations (0-30 microM) of spermidine. Microfluorometric results and chemical determinations of spermidine and spermine were obtained from each separate slide. The cellular total polyamine (spermidine + spermine) concentration on the slides varied between 4 and 15 nmol per mg protein (MCF-7 cells) and 5 and 26 nmol per mg protein (HeLa cells) and the corresponding microfluorometric results between 60 and 115 arbitrary units (MCF-7 cells) and 80 and 160 arbitrary units (HeLa cells). Simple regression analysis showed a good linear relationship between cellular polyamine concentration and FF-fluorescence yield. The correlation coefficient for MCF-7 cells was 0.86 and for HeLa cells 0.82, significance of the correlations was p less than or equal to 0.0001. Our results add further credence to the specificity of the FF-method and indicate that the method may be useful for microfluorometric quantitation of polyamines in situ. 相似文献
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Silvia Cetrullo Benedetta Tantini Annalisa Facchini Carla Pignatti Claudio Stefanelli Claudio Marcello Caldarera Flavio Flamigni 《Amino acids》2011,40(4):1127-1137
Recent studies report that the primary transmitter of sympathetic nervous system norepinephrine (NE), which is actively produced
in failing human heart, is able to induce apoptosis of rat cardiomyocytes. Apoptotic cell death of cardiomyocytes is involved
in several cardiovascular diseases including ischemia, hypertrophy and heart failure, therefore representing a potential therapeutic
target. The natural occurring polyamines, putrescine, spermidine and spermine, are biogenic amines involved in many cellular
processes, including apoptosis. Thus, we have studied the involvement of polyamines in the apoptosis of cardiac cells induced
by the treatment with NE. The results indicate that NE caused an early induction of the activity of ornithine decarboxylase
(ODC), the first enzyme in polyamine biosynthesis, followed by a later increase of apoptotic cell death. This effect was prevented
in the presence of α-difluoromethylornithine, an irreversible inhibitor of ODC. Moreover, the study of some key signal transduction
pathways revealed an involvement of AMP-activated protein kinase, AKT and p38 mitogen-activated protein kinases, in the modulation
by polyamines of the response of cardiomyocytes to NE. In fact, polyamine-depleted cells showed an altered activation pattern
of these kinases that may contrast apoptosis and appeared to result from a differential effect on the specific phosphatases
that dephosphorylate and switch off these signaling proteins. In conclusion, these results indicate that in cardiac cells
polyamines are involved in the execution of the death program activated by NE, and suggest that their apoptosis facilitating
action is mediated by a network of specific phosphatases and kinases. 相似文献
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Coffino P 《Nature reviews. Molecular cell biology》2001,2(3):188-194
Proteins that are degraded by the proteasome are first modified by a set of enzymes that attach multiple copies of ubiquitin to substrate lysines, but a tiny minority, including the polyamine-synthesizing enzyme ornithine decarboxylase, is handled differently. This enzyme is targeted for destruction by another protein--antizyme. Why does ornithine decarboxylase have its own dedicated destruction mechanism, how does it work, and is it the only protein to be targeted to the proteasome in this way? 相似文献
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Rakesh Minocha Walter C. Shortle Stephanie L. Long Subhash C. Minocha 《Journal of Plant Growth Regulation》1994,13(4):187-193
A fast and reliable method for the extraction of cellular polyamines and major inorganic ions (Ca, Mg, Mn, K, and P) from several plant tissues is described. The method involves repeated freezing and thawing of samples instead of homogenization. The efficiency of extraction of both the polyamines and inorganic ions by these two methods was compared for 10 different tissues. In each case, the freeze-thaw procedure resulted in a precise and quantitatively equal, or greater, yield than homogenization. Freeze-thawing not only eliminates the need for various tissue homogenizers (such as polytrons, tissumizers, and mortars and pestles), but it is so simple that a large number of samples can be processed simultaneously. We routinely processed 50–80 samples for quantitation of polyamines and inorganic ions. Freeze-thawing was equally useful for the extraction of polyamines from liver, spleen, and kidney tissues of mice.New Hampshire Agricultural Experiment Station, scientific contribution no. 1845. 相似文献
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It has been shown that oxidation of polyamines by polyamine oxidases can produce toxic compounds (H2O2, aldehydes, ammonia) and that the polyamine oxidase-polyamine system is implicated, in vitro, in the death of several parasites. Using Amoeba proteus as an in vitro model, we studied the cytotoxicity to these cells of spermine, spermidine, their acetyl derivatives, and their hypothetical precursors. Spermine and N
1-acetylspermine were more toxic than emetine, an amoebicidal reference drug. Spermine presented a short-term toxicity, but a 48-h contact time was necessary for the high toxicity of spermidine. The uptake by Amoeba cells of the different polyamines tested was demonstrated. On the other hand, a high polyamine oxidase activity was identified in Amoeba proteus crude extract. Spermine (theoretical 100%) and N
1-acetylspermine (64%) were the best substrates at pH 9.5, while spermidine, its acetyl derivatives, and putrescine were very poorly oxidized by this enzyme (3–20%). Spermine oxidase activity was inhibited by phenylhydrazine (nil) and isoniazid ( 50%). Mepacrine did not inhibit the enzyme activity at pH 8. Neither monoamine nor diamine oxidase activity ( 10%) was found. It must be emphasized that spermine, the best enzyme substrate, is the most toxic polyamine. This finding suggests that knowledge of polyamine oxidase specificity can be used to modulate the cytotoxicity of polyamine derivatives. Amoeba proteus was revealed as a simple model for investigation of the connection between cytotoxicity and enzyme activity.Abbreviations DAO
diamine oxidase
- DFMO
DL--difluoromethylornithine
- DP
1-3-diaminopropane
- IC50
50% inhibition concentration
- MAO
monoamine oxidase
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N
1-ACSP;
N
1-acetylspermine
- N1-ACSPD
N
1-acetylspermidine
-
N
8-ACSPD
N
8-acetylspermidine
- ODC
ornithine decarboxylase
- PAO(s)
polyamine oxidase(s)
- PUT
putrescine
- SP
spermine
- SPD
spermidine 相似文献
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J C Allen C J Smith J I Hussain J M Thomas J M Gaugas 《European journal of biochemistry》1979,102(1):153-158
Spermine and spermidine in vitro are potent inhibitors of proliferation of phytohaemagglutinin-stimulated rat thymic lymphocytes, lymphoma cells and human lymphoblastic leukaemia cells, but only in media supplemented by foetal calf serum. This inhibition is shown to be due to a bovine plasma polyamine oxidase, with a high specificity for these polyamines. Spontaneously dividing lymphocytes are not subject to this inhibition. This, plus direct evidence from synchronous cultures of EB2 cells demonstrates that the inhibition is expressed in the late G1 or G1/S interface of the cell cycle. Putrescine was not an inhibitor in the presence of foetal calf serum but became so in the presence of human pregnancy serum, possibly due to the action of diamine oxidase. 相似文献
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Background
Chronic hypoxia is a major component of ischemic diseases such as stroke or myocardial infarction. Drosophila is more tolerant to hypoxia than most mammalian species. It is considered as a useful model organism to identify new mechanisms of hypoxic tolerance. The hypoxic tolerance of flies has previously been reported to be enhanced by low protein diets. This study analyses the mechanisms involved.Results
Feeding adult Drosophila on a yeast diet dramatically reduced their longevities under chronic hypoxic conditions (5% O2). Mean and maximum longevities became close to the values observed for starving flies. The action of dietary yeast was mimicked by a whole casein hydrolysate and by anyone of the 20 natural amino acids that compose proteins. It was mimicked by amino acid intermediates of the urea cycle such as L-citrulline and L-ornithine, and by polyamines (putrescine, spermidine and spermine). α-difluoromethylornithine, a specific inhibitor of ornithine decarboxylase, partially protected hypoxic flies from amino acid toxicity but not from polyamine toxicity. N1-guanyl-1,7 diaminoheptane, a specific inhibitor of eIF5A hypusination, partially relieved the toxicities of both amino acids and polyamines.Conclusion
Dietary amino acids reduced the longevity of chronically hypoxic flies fed on a sucrose diet. Pharmacological evidence suggests that the synthesis of polyamines and the hypusination of eIF5A contributed to the life-shortening effect of dietary amino acids. 相似文献20.
E. C. Koc S. Bagga D. D. Songstad S. R. Betz G. D. Kuehn G. C. Phillips 《In vitro cellular & developmental biology. Plant》1998,34(3):252-255
Summary The uncommon polyamines, norspermidine and norspermine, were detected in maizein vitro cultures of three different genotypes. The common polyamines, spermidine and spermine, along with the diamine, putrescine,
were also observed. The total amounts of the uncommon polyamines, norspermidine and norspermine, were comparable to the total
amounts of the common polyamines, spermidine and spermine, in the maize tissues. The titer for norspermidine was 6- to 15-fold
greater than that of its common counterpart (spermidine) in the three genotypes. Norspermidine was the predominant polyamine
among all triamines and tetramines detected in cell cultures of two of the three genotypes of maize examined and was predominant
along with spermine in the third genotype. Enzyme assays performed with extracts from callus of one of the genotypes suggested
a likely mechanism to account for the biosynthesis of the uncommon polyamines in cultured maize cells, through the actions
of putrescine aminopropyltransferase, polyamine oxidase, and Schiff-base reductase/decarboxylase enzyme activities. This is
the first report of the detection of uncommon polyamines in maize tissues, as well as the first report of these uncommon polyamines
in a monocotyledonous plant. 相似文献