Increase in S-adenosylmethionine decarboxylase in SV-3T3 cells treated with S-methyl-5'-methylthioadenosine.
Ontology highlight
ABSTRACT:
SUBMITTER: Pegg AE
PROVIDER: S-EPMC1147951 | biostudies-other | 1987 May
REPOSITORIES: biostudies-other
Similar Datasets
Project description:The effects of alpha-difluoromethylornithine, a specific inhibitor of ornithine decarboxylase, on cell growth rate, polyamine content and the content of decarboxylated S-adenosylmethionine in SV-3T3 transformed mouse fibroblasts were studied. DL-alpha-Difluoromethylornithine at 1 mM or higher concentrations decreased the growth rate by over 90% after 2 or more days of exposure, but the cells remained viable, although quiescent for at least 9 days. Addition of 10 microM-spermidine or -spermine or 50 microM-putrescine at any time throughout this period completely reversed the inhibition of growth. Treatment with alpha-difluoromethylornithine decreased putrescine and spermidine contents by more than 98% and that of spermine by 60%, but cells exposed to exogenous polyamines did not require complete replenishment of the polyamine pools to resume growth. In fact, a virtually normal growth rate was obtained in cells lacking putrescine, having 2% of normal spermidine content and 156% of normal spermine. These results suggest that the well-known increase in putrescine and spermidine in cells stimulated for growth is not essential for this to occur and that mammalian cells can utilize spermine as their only polyamine. A substantial reversal of the growth-inhibitory effect of alpha-difluoromethylornithine was produced by a number of polyamines not normally found in mammalian cells, including the spermidine analogues aminopropylcadaverine and sym-homospermidine, which were partially converted into their respective spermine analogues by addition of an aminopropyl group within the cell. The spermine analogue sym-norspermine was also effective, but the maximal growth rate produced by these unphysiological polyamines was only 60-70% of that produced by the normal polyamines. These results indicate that spermidine and spermine have the optimal length for activation of the cellular processes critically dependent on polyamines and should help in identifying these processes. Exposure to alpha-difluoromethylornithine leads to an enormous rise in the concentration of decarboxylated S-adenosylmethionine, which reached a peak at 530-fold after 3 days of exposure and steadily declined to 140-fold after 11 days. This increase was abolished by addition of exogenous polyamines, which rapidly decreased the activity of S-adenosylmethionine decarboxylase. The increase in decarboxylated S-adenosylmethionine is unlikely to be solely responsible for the decrease to the same extent by spermine, sym-norspermidine and sym-homospermidine, which produce 97%, 16% and 60% of the control growth rate, respectively.(ABSTRACT TRUNCATED AT 400 WORDS)
| S-EPMC1144394 | biostudies-other
Project description:The contrasting control of lysosomal proteinases, protein turnover and proliferation was studied in 3T3 and SV-3T3 (SV-40-virus-transformed 3T3) cells. 1. In 3T3 cells, net protein accumulation proceeded from 5%/h (doubling time, T(d)=14h) in growing cells to 0%/h as cells became quiescent. SV-3T3 cells never ceased to gain protein, but rather decreased their protein accumulation rate from 6-7%/h (T(d)=10-12h) to 2%/h (T(d)=35-40h) just before culture death in unchanged medium. 2. In both cell types the rates of protein synthesis per unit of protein (a) were proportional to the initial serum concentration from 0 to 6%, and (b) declined under progressive depletion of undefined serum growth factors. In depleted growth medium, leucine incorporation per unit of protein in 3T3 and SV-3T3 cells declined to almost equal synthetic rates while the 3T3 cell existed in a steady state of zero net gain, and the SV-3T3 cell continued to gain protein at a rate of 2%/h. 3. Whereas a large fraction of the control of 3T3-cell net protein accumulation can be accounted for by an increase in degradation from 1%/h to 3%/h, the SV-3T3 cell did not exhibit a growth-related increase in degradation appreciably above 1%/h. 4. Thus, by using first-order kinetics, the continued net protein accumulation of the transformed cell can be accounted for by a failure to increase protein degradation, whereas fractional synthesis can be made to decline to a rate similar to that in the quiescent non-transformed cell. 5. Upon acute serum deprivation, both cell types similarly exhibited small rapid increases in proteolysis independent of cell growth state or lysosomal enzyme status. 6. The 3T3 cell increased its lysosomal proteinase activity in conjunction with increase in the growth-state-dependent proteolytic mechanism; however, the SV-3T3 cell failed to increase lysosomal proteinases or the growth-state-dependent proteolytic mechanism.
| S-EPMC1158580 | biostudies-other
Project description:The crenarchaeon Sulfolobus solfataricus uses arginine to produce putrescine for polyamine biosynthesis. However, genome sequences from S. solfataricus and most crenarchaea have no known homologs of the previously characterized pyridoxal 5'-phosphate or pyruvoyl-dependent arginine decarboxylases that catalyze the first step in this pathway. Instead they have two paralogs of the S-adenosylmethionine decarboxylase (AdoMetDC). The gene at locus SSO0585 produces an AdoMetDC enzyme, whereas the gene at locus SSO0536 produces a novel arginine decarboxylase (ArgDC). Both thermostable enzymes self-cleave at conserved serine residues to form amino-terminal beta-domains and carboxyl-terminal alpha-domains with reactive pyruvoyl cofactors. The ArgDC enzyme specifically catalyzed arginine decarboxylation more efficiently than previously studied pyruvoyl enzymes. alpha-Difluoromethylarginine significantly reduced the ArgDC activity of purified enzyme, and treating growing S. solfataricus cells with this inhibitor reduced the cells' ratio of spermidine to norspermine by decreasing the putrescine pool. The crenarchaeal ArgDC had no AdoMetDC activity, whereas its AdoMetDC paralog had no ArgDC activity. A chimeric protein containing the beta-subunit of SSO0536 and the alpha-subunit of SSO0585 had ArgDC activity, implicating residues responsible for substrate specificity in the amino-terminal domain. This crenarchaeal ArgDC is the first example of alternative substrate specificity in the AdoMetDC family. ArgDC activity has evolved through convergent evolution at least five times, demonstrating the utility of this enzyme and the plasticity of amino acid decarboxylases.
| S-EPMC2533785 | biostudies-literature
Project description:S-adenosylmethionine decarboxylase (AdoMetDC) is a critical enzyme in the polyamine biosynthetic pathway and a subject of many structural and biochemical investigations for anti-cancer and anti-parasitic therapy. The enzyme undergoes an internal serinolysis reaction as a post-translational modification to generate the active site pyruvoyl group for the decarboxylation process. The crystal structures of AdoMetDC from Homo sapiens, Solanum tuberosum, Thermotoga maritima, and Aquifex aeolicus have been determined. Numerous crystal structures of human AdoMetDC and mutants have provided insights into the mechanism of autoprocessing, putrescine activation, substrate specificity, and inhibitor design to the enzyme. The comparison of the human and potato enzyme with the T. maritima and A. aeolicus enzymes supports the hypothesis that the eukaryotic enzymes evolved by gene duplication and fusion. The residues implicated in processing and activity are structurally conserved in all forms of the enzyme, suggesting a divergent evolution of AdoMetDC.
| S-EPMC2847442 | biostudies-literature
Project description:BHMT (betaine-homocysteine methyltransferase) remethylates homocysteine to form methionine. SAM (S-adenosylmethionine) inhibits BHMT activity, but whether SAM modulates BHMT gene expression is unknown. Transcriptional regulation of the human BHMT is also unknown. The present study examined regulation of the human BHMT gene by SAM and its metabolite, MTA (5'-methylthioadenosine). To facilitate these studies, we cloned the 2.7 kb 5'-flanking region of the human BHMT gene (GenBank accession number AY325901). Both SAM and MTA treatment of HepG2 cells resulted in a dose- and time-dependent decrease in BHMT mRNA levels, which paralleled their effects on the BHMT promoter activity. Maximal suppression was observed with the BHMT promoter construct -347/+33, which contains a number of NF-kappaB (nuclear factor kappaB) binding sites. SAM and MTA treatment increased NF-kappaB nuclear binding and NF-kappaB-driven luciferase activities, and increased nuclear binding activity of multiple histone deacetylase co-repressors to the NF-kappaB sites. Overexpression of p50 and p65 decreased BHMT promoter activity, while blocking NF-kappaB activation increased BHMT expression and promoter activity, and prevented SAM but not MTA's ability to inhibit BHMT expression. The NF-kappaB binding site at -301 is responsible, at least in part, for this effect. Lower BHMT expression can impair homocysteine metabolism, which can induce ER (endoplasmic reticulum) stress. Indeed, MTA treatment resulted in increased expression ER stress markers. In conclusion, SAM and MTA down-regulate BHMT expression in HepG2 cells in part by inducing NF-kappaB, which acts as a repressor for the human BHMT gene. While SAM's mechanism is NF-kappaB-dependent, MTA has both NF-kappaB-dependent and -independent mechanisms.
| S-EPMC1698693 | biostudies-literature
Project description: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.
| S-EPMC1135037 | biostudies-other
Project description:Chronic inflammation is an underlying risk factor for colon cancer. Tumor necrosis factor alpha (TNF-?) plays a critical role in the development of inflammation-induced colon cancer in a mouse model. S-adenosylmethionine (SAMe) and its metabolite methylthioadenosine (MTA) can inhibit lipopolysaccharide-induced TNF-? expression in macrophages. The aim of this work was to examine whether SAMe and MTA are effective in preventing inflammation-induced colon cancer and if so identify signaling pathways affected. Balb/c mice were treated with azoxymethane (AOM) and dextran sulfate sodium to induce colon cancer. Two days after AOM treatment, mice were divided into three groups: vehicle control, SAMe or MTA. Tumor load, histology, immunohistochemistry, gene and protein expression were determined. SAMe and MTA treatment reduced tumor load by ?40%. Both treatments raised SAMe and MTA levels but MTA also raised S-adenosylhomocysteine levels. MTA treatment prevented the induction of many genes known to play pathogenetic roles in this model except for TNF-? and inducible nitric oxide synthase (iNOS). SAMe also had no effect on TNF-? or iNOS and was less inhibitory than MTA on the other genes. In vivo, both treatments induced apoptosis but inhibited proliferation, ?-catenin, nuclear factor kappa B activation and interleukin (IL) 6 signaling. Effect of SAMe and MTA on IL-6 signaling was examined using Colo 205 colon cancer cells. In these cells, SAMe and MTA inhibited IL-6-induced IL-10 expression. MTA also inhibited IL-10 transcription and signal transducer and activator of transcription 3 activation. In conclusion, SAMe and MTA reduced inflammation-induced colon cancer and inhibited several pathways important in colon carcinogenesis.
| S-EPMC3279046 | biostudies-literature
Project description:Neoplastic growth is associated with increased polyamine biosynthetic activity and content. Tumor promoter treatment induces the rate-limiting enzymes in polyamine biosynthesis, ornithine decarboxylase (ODC), and S-adenosylmethionine decarboxylase (AdoMetDC), and targeted ODC overexpression is sufficient for tumor promotion in initiated mouse skin. We generated a mouse model with doxycycline (Dox)-regulated AdoMetDC expression to determine the impact of this second rate-limiting enzyme on epithelial carcinogenesis. TetO-AdoMetDC (TAMD) transgenic founders were crossed with transgenic mice (K5-tTA) that express the tetracycline-regulated transcriptional activator within basal keratinocytes of the skin. Transgene expression in TAMD/K5-tTA mice was restricted to keratin 5 (K5) target tissues and silenced upon Dox treatment. AdoMetDC activity and its product, decarboxylated AdoMet, both increased approximately 8-fold in the skin. This enabled a redistribution of the polyamines that led to reduced putrescine, increased spermine, and an elevated spermine:spermidine ratio. Given the positive association between polyamine biosynthetic capacity and neoplastic growth, it was somewhat surprising to find that TAMD/K5-tTA mice developed significantly fewer tumors than controls in response to 7,12-dimethylbenz[a]anthracene/12-O-tetradecanoylphorbol-13-acetate chemical carcinogenesis. Importantly, tumor counts in TAMD/K5-tTA mice rebounded to nearly equal the levels in the control group upon Dox-mediated transgene silencing at a late stage of tumor promotion, which indicates that latent viable initiated cells remain in AdoMetDC-expressing skin. These results underscore the complexity of polyamine modulation of tumor development and emphasize the critical role of putrescine in tumor promotion. AdoMetDC-expressing mice will enable more refined spatial and temporal manipulation of polyamine biosynthesis during tumorigenesis and in other models of human disease.
| S-EPMC3499050 | biostudies-literature
Project description:The present study was designed to provide a better understanding of the role played by AdoMetDC (S-adenosylmethionine decarboxylase), the key rate-controlling enzyme in the synthesis of spermidine and spermine, in controlling polyamine levels and the importance of polyamines in cardiac physiology. The alphaMHC (alpha-myosin heavy chain) promoter was used to generate transgenic mice with cardiac-specific expression of AdoMetDC. A founder line (alphaMHC/AdoMetDC) was established with a >100-fold increase in AdoMetDC activity in the heart. Transgene expression was maximal by 1 week of age and remained constant into adulthood. However, the changes in polyamine levels were most pronounced during the first week of age, with a 2-fold decrease in putrescine and spermidine and a 2-fold increase in spermine. At later times, spermine returned to near control levels, whereas putrescine and spermidine levels remained lower, suggesting that compensatory mechanisms exist to limit spermine accumulation. The alphaMHC/AdoMetDC mice did not display an overt cardiac phenotype, but there was an increased cardiac hypertrophy after beta-adrenergic stimulation with isoprenaline ('isoproterenol'), as well as a small increase in spermine content. Crosses of the alphaMHC/AdoMetDC with alphaMHC/ornithine decarboxylase mice that have a >1000-fold increase in cardiac ornithine decarboxylase were lethal in utero, presumably due to increase in spermine to toxic levels. These findings suggest that cardiac spermine levels are highly regulated to avoid polyamine-induced toxicity and that homoeostatic mechanisms can maintain non-toxic levels even when one enzyme of the biosynthetic pathway is greatly elevated but are unable to do so when two biosynthetic enzymes are increased.
| S-EPMC1383688 | biostudies-literature
Project description:New therapeutic options are needed for treatment of human African trypanosomiasis (HAT) caused by protozoan parasite Trypanosoma brucei. S-Adenosylmethionine decarboxylase (AdoMetDC) is an essential enzyme in the polyamine pathway of T. brucei. Previous attempts to target this enzyme were thwarted by the lack of brain penetration of the most advanced series. Herein, we describe a T. brucei AdoMetDC inhibitor series based on a pyrimidineamine pharmacophore that we identified by target-based high-throughput screening. The pyrimidineamines showed selectivity for T. brucei AdoMetDC over the human enzyme, inhibited parasite growth in whole-cell assay, and had good predicted blood-brain barrier penetration. The medicinal chemistry program elucidated structure-activity relationships within the series. Features of the series that were required for binding were revealed by determining the X-ray crystal structure of TbAdoMetDC bound to one analog. The pyrimidineamine series provides a novel starting point for an anti-HAT lead optimization.
| S-EPMC5965259 | biostudies-literature