Polyamines as Biomarkers for Prostate Cancer

The observation that not only a number of enzymatic steps in polyamine synthesis, but also enzymes directing substrates toward polyamines, are overexpressed in ODC Polyamines in Different Phenotypes of Human Prostatic Cancer Cells Androgen AR Diff. ODC ODC activity Spd Spm Spd Spm Androgen AR Diff. ODC ODC activity Spd Spm Spd Spm

Liver

SSAT-overexpressing mice are extremely sensitive to the toxic effects exerted by polyamine analogs, such as DENSPM, in comparison with their wild-type littermates (14). Although the exact mechanisms of this toxicity are not known, treatment of trans-genic mice overexpressing SSAT under the control of metallothionein promoter with DENSPM caused a profound depletion of hepatic spermidine and spermine associated with swelling of hepatic mitochondria and marked mortality in comparison with their...

Proteins Involved in Apoptosis

The general steps involved in apoptosis are remarkably conserved during the evolutionary process, and are due to three types of proteins. These may be labeled as effectors, accessory molecules or adaptors, and regulators. A family of cysteine proteases called caspases are the effectors of apoptosis (6). Caspases exist in the cytoplasm as presynthesized proenzymes, and are activated in response to apoptotic stimuli. There are two basic types of caspases. Caspases 3, 6, and 7 are termed...

Translational Regulation

The translational efficiency of AdoMetDC mRNA is regulated by growth stimulation, changes in intracellular levels of spermidine and spermine, and according to cell type. In vitro experiments with a reticulocyte lysate system demonstrated that translation of AdoMetDC mRNA was maximal when polyamines had been removed from the lysate by gel filtration, and that adding back 80 iM spermine or 0.8 mM spermidine substantially reduced AdoMetDC synthesis, while having no effect on total protein...

Localization of Polyamine Related Enzymes in the Kidney

The basic pathways of polyamine biosynthesis and interconversion are shown in Fig. 1. Ornithine decarboxylase (ODC), the first rate-limiting enzyme of polyamine biosynthesis, is found throughout the kidney. The highest concentration is found in the proximal tubule, followed by the distal and then collecting tubules (2), with activity primarily localized to the proximal tubules (3-5). Expression of ODC, along with ornithine aminotransferase (OAT) is primarily in the outer (convoluted) proximal...

Effects of Elevated Intracellular Levels of Polyamines on Cultured Mammalian Cells

Experiments relying on exogenous addition of polyamines to assay buffers or on the cellular uptake of polyamines added to cell culture medium do not always mimic the normal compartmentalization of polyamines synthesized naturally in the cell, or necessarily lead to the same biological outcome (6,35). For this reason, we have used strategies relying on the manipulation of the natural polyamine biosynthetic pathway to examine the consequences of elevated intracellular polyamines on histone...

[Ca2cyt Through CCE and Transient Receptor Potential Channels

Polyamine-modulated Ca2+ driving force is critical for transmembrane influx of Ca2+ in intestinal epithelial cells, but specific Ca2+-permeable channels in this process are still unknown. In nonexcitable cells, passive Ca2+ leakage, receptor-operated Ca2+ channels, nonselective cation channels, and store-operated channels, all contribute to Ca2+ influx (13,14). It has been shown that intestinal epithelial cells lack VDCCs, but have developed the Ca2+ entry mechanism that is coupled with the...

Polyamine Depletion Stabilizes p53 mRNA

In response to polyamine depletion, cellular levels of p53 protein are greatly increased, and the ability of p53 to bind specific DNA sequences is significantly activated (16,17,25,26). Because expression of the p53 gene is rapid and transient, it acts as a mediator, linking short-term stress signals to growth arrest or apoptosis by regulating the activation of specific genes. p53 is an ephemeral protein, and its half-life is approx 25-40 min in variety of cell types (17). In addition to...

Mammary Carcinogenesis

Considerable evidence indicates that increased ODC activity and cellular polyamine levels are involved in carcinogenesis. Increased ODC activity is frequently detected in transformed cell lines (5), animal tumors (6), and some preneoplastic lesions (7). In addition, activation of the polyamine pathway has been found to be essential in several experimental systems of carcinogenesis (8,9). The observation that levels of polyamines and ODC activity are higher in human breast cancer specimens than...

Conclusions

Pp2a Stat3

If the scheme hypothesized in Fig. 7 proves correct, the obvious unanswered question are how does the removal of polyamines result in Src activation Could polyamines prevent the translocation of Src to the membrane Could they block its activation site Obviously, there are as many possibilities as there are steps in the activation process. The answers to these questions could prove interesting and important to our understanding not only of polyamine interactions, but also apoptosis itself. Fig....

M

Macrophage, Helicobacter pylori-induced apoptosis, arginase upregulation, 297 ornithine decarboxylase induction, 298, 307 polyamine oxidase role, 301-303, 305 polyamine regulation, 298, 305, 307 polyamine-synthesizing enzyme induction, 298, 299, 301 prospects for study, 307, 308 MAPK, see Mitogen-activated protein kinase Max, Myc dimers, 251 MCBG, see Methylglyoxal is(guanylhydrazone) MDL 27695, antitumor activity, 8 Methylglyoxal Ms(guanylhydrazone) (MCBG), polyamine synthesis inhibition, 6...

Introduction

The biosynthetic pathway of the polyamines is practically irreversible because it involves two decarboxylation reactions. The first evidence, however, indicating that spermidine can be converted to putrescine and spermine to spermidine emerged in the late 1960s when Siimes (1) found that radioactive spermidine and spermine yielded labeled putrescine and spermidine in rat liver in vivo. An additional 10 yr were required before the first enzyme, polyamine oxidase (PAO), of a separate polyamine...

Effect of Polyamine Biosynthesis on Apoptosis

Ornithine decarboxylase (ODC) is one of the rate-limiting enzymes of polyamine biosynthesis. Cells have developed a complex regulatory mechanism, which precisely controls intracellular polyamine levels by the combined effects of de novo synthesis, back conversion, export, and uptake of polyamines. The exact role of polyamines in apoptotic pathways is still unclear. An elegant series of studies by Packham and Cleveland (6) linked increases in ODC activity to apoptosis in murine myeloid cells....

Info

Bp 0.05 vs the combined DFMO treatment groups (10 and 14 wk). cp < 0.05 vs each of the remaining groups. (Reproduced with permission from ref. 44.) Fig. 3. Growth in nude mice of orthotopically implanted green fluorescent protein-tagged MDA-MB-435 breast cancer cells. A total of 1 x 106 cells in 0.1 mL Hank's balanced salt solution were injected in athymic mice in the absence and in the presence of DFMO treatment (2 in drinking H2O). Growth curve analysis using a linear model revealed no...

Nitric Oxide

In acute inflammatory models, (A) arginine to nitric oxide (NO) production is an early phase response. NO is an antiproliferative molecule resulting, to some degree, from nitrosylation and inactivation of ornithine decarboxylase (ODC). (B) The production of ornithine and subsequent metabolism to the pro-proliferative polyamines (ODC) and to proline (OAT) for extracellular matrix production is a later repair phase response. Agmatine induction of antizyme and SSAT can...

Polyamines and ECadherin Expression

A series of studies from our laboratory has demonstrated that polyamines are necessary for expression of E-cadherin in intestinal epithelial cells and that polyamine-modulated E-cadherin plays an important role in maintenance of normal epithelial barrier functions (24-26). To determine the involvement of polyamines in regulation of cell-cell adherens junctions and tight junctions, normal intestinal epithelial cells (IEC-6 line) were grown in the presence or absence of a-difluoromethylornithine...

Role of Arginine Metabolizing Enzymes

The majority of work completed to date on ODC and cardiac hypertrophy has measured the effects of altered ODC activity only on the polyamines, which are downstream products of the ODC reaction. However, the ODC protein is part of a more complex signaling cascade (Fig. 1). The ODC substrate ornithine is produced in all cells by the activity of arginase, which catalyzes the conversion of L-arginine to L-ornithine and urea (39). Arginase exists as two isoforms, with arginase I targeted to the...

Enzyme Activation

As well as stimulating the rate of proenzyme processing, putrescine stimulates catalytic activity of the processed enzyme by reducing the K for AdoMet (8). Hence supply of putrescine is linked to the production of dcAdoMet, the other precursor for spermidine production. Site-directed mutagenesis experiments demonstrated that residues Glu-178 and Glu-256 are critical for putrescine stimulation of catalytic activity and for proenzyme processing of mammalian AdoMetDC, but because the E11Q mutant...

References

J. (2003) Polyamine metabolism and cancer. J. Cell. Mol. Med. 7, 113-126. 2. Huang, Y., Pledgie, A., Casero, R. A. Jr., and Davidson, N. E. (2005) Molecular mechanisms of polyamine analogs in cancer cells. Anticancer Drugs 16, 229-241. 3. Igarashi, K. and Kashiwagi, K. (2000) Polyamines mysterious modulators of cellular functions. Biochem. Biophys. Res. Commun. 271, 559-564. 4. Childs, A. C., Mehta, D. J., and Gerner, E. W. (2003) Polyamine-dependent gene expression....

A

H. pylori induces mitochondrial membrane depolarization, translocation of cytochrome c from mitochondria to cytosol, and activation of caspase-3 that is inhibited by MDL 72527. RAW 264.7 cells were treated with HPL in the absence and presence of MDL 72527 (250 M). (A) Mitochondrial membrane potential was measured by flow cytometry using MitoCapture dye at the times indicated. Summary data of mean relative fluorescence, expressed as negative values to indicate...

Increased Expression of the TGFb Gene Contributes to Growth Inhibition in Polyamine Deficient Cells

Activation of TGF-b gene expression is involved in the process of growth inhibition after polyamine depletion (16). In normal IECs, increased synthesis of TGF-b in polyamine-deficient cells is associated with a decrease in cell proliferation. Exogenous spermidine given with DFMO prevents the increased expression of the TGF-b gene and returns cell growth to near normal. TGF -b added to the culture medium also decreases the rate of total cellular DNA synthesis and final cellular number in normal...

Polyamine Depletion Fails to Induce junD Gene Transcription but Stabilizes JunD mRNA

The increased JunD AP-1 activity after polyamine depletion is primarily the result of activation of junD gene expression in IECs (8). There are significant increases in JunD mRNA and protein in polyamine-deficient IEC-6 cells (9), although expression of the c-fos, c-jun, and junB genes are decreased (5,12). Increased JunD mRNA levels were approx 6.6 times the control levels on d 4 and approximately nine times on d 6 after treatment with DFMO. Increased levels of JunD mRNA were paralleled by a...

The Two Step Mechanism of Polyamine Internalization

Our investigations of the mechanism of Spd -BODIPY internalization have led us to define two successive steps in the transport process an initial influx across the plasma membrane into the cytosolic compartment via a classic permease or channel, immediately followed by its accumulation from the cytosol into PSVs via a H+-dependent vesicular antiporter (Fig. 3) (40). In this model, the equilibrium among the extracellular medium, cytosolic, and vesicular compartments strongly favors accumulation...

ODC Protein is Expressed in Neurons and Microglia Cells

To ensure the cellular localization of ODC in the untreated mouse brain, a multiple labeling approach was performed. Cytoplasmic ODC protein colocalized nicely with the specific marker of neuronal nuclei NeuN, which provided the evidence that neurons have the ability to produce polyamines (61). ODC-immunoreactive cells also colocalized with a marker of microglial cells, iba1. Moreover, ODC+ NeuN+ cells were more numerous and widely distributed than ODC7iba1+ cells across the cerebral tissue of...

Polyamines in Innate Immune Response In Vitro Modulation of Cytokine Release and Nitric Oxide Biosynthesis

The innate immune system is characterized by an unspecific and rapid response to microbial components as various as peptidoglycan, lipoproteins, lipoteichoic acid, bacterial CpG DNA, and the endotoxin lipopolysaccharide (LPS) ( 36,37). The latter is a major glycolipid constituting the outer membrane of Gram-negative bacteria. LPS is well known to activate macrophages and trigger the release of proinflammatory cytokines (Fig. 1) and therefore is widely used as a model of the innate immune system...

Polyamine Structure and Synthetic Analogs Patrick M Woster

The polyamines putrescine (1,4-diaminobutane), spermidine (1,8-diamino-4-aza-octane, 2), and spermine (1,12-diamino-4,9-diazadodecane, 3) (Fig. 1) are ubiquitous polycationic compounds that are found in significant amounts in nearly every prokary-otic and eukaryotic cell type. Spermidine and spermine primarily exist in aqueous solution at pH 7.4 as fully protonated polycations and possess the pKa values indicated in Fig. 1 (1). This high degree of positive charge is an important factor in the...

Voltage Dependence of Polyamine Block Interactions of Blocking and Permeant Cations

A coherent structural model of polyamine-induced rectification must also be able to explain the strong voltage dependence of polyamine blockade relative to other blockers, such as Mg2+ or quaternary ammonium compounds. This important property of rectification refers to the steepness of the conductance-voltage (g-V) relationship of strongly rectifying channels in the presence of various blocking agents. Although blockade by Mg2+ has a relatively shallow voltage dependence (an effective valence...

Spni

Proposed model for Helicobacter py lowi-induced transformation of gastric epithelial cells. H. pylori infection results in an increase in SMO PAOh1 mRNA, protein, and oxidase activity. The resulting H2O2 damages DNA, which, if the damage is sufficient, can cause apop-tosis or has the potential to lead to genetic mutation. If such mutations produce a growth or survival advantage, the potential for neoplastic transformation exists. Fig. 9. Proposed model for Helicobacter py lowi-induced...

[Ca2cyt and Polyamine Dependent Cell Migration

GI mucosal injury occurs commonly from mild physical trauma during digestion, critical and chronic illnesses, various surgical conditions, the ingestion of alcohol, aspirin, nonsteroidal anti-inflammatory compounds, or Helicobacter pylori infection. Restoration of normal intestinal mucosal integrity successful repair of wounds and ulcers requires epithelial cell decisions that regulate signaling networks controlling gene expression, survival, migration, and proliferation. In the acute response...

Increased p53 Causes Growth Inhibition in Polyamine Deficient Cells

The following three studies provide evidence showing that increased accumulation of p53 protein plays a critical role in the process of growth inhibition after polyamine depletion in normal IECs. In the first study, the time course of p53 gene expression and cell growth were examined after polyamine-deficient cells were exposed to exogenous spermidine (6,20). IEC-6 cells were initially treated with DFMO for 4 d and then exposed to spermidine. The changes in both p 5 3 gene expression and...

Increased JunDAP1 Inhibits Cell Proliferation Through p21

The following three logically linked studies show that increased JunD AP-1 inhibits IEC proliferation at least partially through the activation of the p21 promoter after polyamine depletion (8,3 2). The first study examined the association of increased JunD AP-1 activity with p21 gene expression after polyamine depletion. Consistent on its stimulatory effect on JunD AP-1 activity, depletion of cellular polyamines by DFMO also activated the p21 promoter, induced its mRNA expression, and...

Ill ilii

Effect of PD98059 Parke-Davis (PD) on DFMO-induced suppression of invasiveness in the presence (A) or in the absence (B) of hepatocyte growth factor (HGF) (40 ng mL) as a chemoattractant. MDA-MB-435 breast cancer cells were cultured in regular medium in the absence and in the presence of DFMO (1 mM) for 48 h before plating in the matrigel assay. PD treatment (50 M was added 1 h before plating in the matrigel assay (time found to be sufficient to block extracellular regulated kinase...

ODC Activity in the CNS of Vehicle and LPSTreated Mice

The ODC protein is highly regulated at the levels of transcription and translation, as well as at the posttranslational level. Furthermore, ODC enzyme has a very short halflife. To ascertain that LPS-induced increase in ODC gene expression was really associated with putrescine biosynthesis, ODC activity was measured in control mice and in mice that were treated with a suicide inhibitor of ODC, the difluoromethylornithine (DFMO). ODC activity was strongly induced by sixfold 3 h after a single...

Enzyme Inactivation and Degradation

AdoMetDC has a relatively short half-life (1-2 h), which varies with cell type, and is inversely related to cellular levels of spermidine and spermine (9). Many short-lived proteins in eukaryotic cells are degraded by the ubiquitin proteasome pathway (55,56). The adenosine triphosphate (ATP)-dependent ubiquitination process results in joining of a ubiquitin polypeptide to the protein substrate via an isopeptide bond between a lysine side chain of the substrate and the C terminus of ubiquitin....

Circulating Polyamines as Markers and Tracers for Prostatic Malignancies

Polyamines or their acetylated forms can also be secreted by cells and released into the circulation. Subsequently, circulating polyamines and their acetylated products can be taken up and reused by polyamine-demanding cells. Transport of polyamines from one cell to another via body fluids is believed to be carrier-mediated by either proteins (e.g., anti-polyamine antibodies, lipoproteins) or by cells (peripheral blood cells). Moulinoux et al. showed that more than 95 of circulating spermidine...

Effects of Increased Polyamine Biosynthesis in Mouse Skin

To examine the effects of elevated ODC and polyamines on histone acetylation in a more relevant physiological system in which epithelial cells can still both proliferate and differentiate, unlike the immortalized cell lines studied in the earlier experiments, primary keratinocytes were isolated and cultured from K6 ODC transgenic and normal littermate skin (34). After labeling the keratinocytes with 3H acetic acid in the presence of sodium butyrate, histones were isolated and examined by...

Polyamine biosynthesis pathway

S-adenosylmethionine Ornithine **- Arginine IS-adenosylmethionine j Ornithine C02 Decarboxylase Decarboxylase NH S dbS U,nU se N'-AcetyJspennidine Methylthioadenosine v X .- SSAT Fig. 1. Polyamine biosynthesis and catabolism. Spd-S, spermidine synthase, Sp-S, spermine synthase, PAO, polyamine oxidase, SSAT, spermidine spermine N1 -acetyltransferase. humans are constantly exposed to environmental carcinogens, and that the human diet contains nitrites and nitrates, which are converted in the gut...

DNA Nanoparticle Formation Aggregation and Resolubilization

DNA condensation to nanoparticles is a process in which a significant decrease in the total volume occupied by the DNA occurs, with an orderly collapse of DNA to compact particles of finite size and orderly morphology (13). Condensation is generally provoked under conditions of low concentrations (low micromolar) of DNA and multivalent cations, such as polyamines, inorganic cations (cobalt hexamine (Co(NH3)63+), and polymers, although the presence of dehydrating solvents can accelerate this...

Recent Advances in the Understanding of Mammalian Polyamine Catabolism

Polyamine Pathway

The Regulation and Potential Role of Polyamine Catabolism in Drug Response and Disease Processes Robert A. Casero, Jr., Alison V. Fraser, Tracy Murray-Stewart, Amy Hacker, Naveen Babbar, Jennifer Fleischer, and Yanlin Wang As more data emerge, the significance of polyamine catabolism in polyamine homeostasis, drug response, and disease etiology is expanding. Importantly, the regulation and function of the polyamine catabolic pathway has emerged as a rational target for drug intervention in both...

Polyamine Dependent Early Cellular Signals and Cell Proliferation

Activated Ras

When a resting cell is awakened to enter the cell cycle by mitogenic stimulation, a complex molecular machinery involving positive and negative regulation of cell-cycle progression is initiated. Part of the machinery is quite well-known. It involves receptors of various kinds, protein interactions, phosphorylation reactions, and molecular cascades carrying the signal to enter the cell cycle from the cell surface to the nucleus. In the nucleus, transcriptional activation results in an increased...

Acknowledgments

The work in the authors' laboratories has been supported by the Academy of Finland. References 1. Siimes, M. (1967) Studies on the metabolism of 1,4-14C-spermidine and 1, 4-14C -spermine in the rat. Acta Physiol. Scand. Suppl. 298, 1-66. 2. Holtta, E. (1997) Oxidation of spermidine and spermine in rat liver Purification and properties of polyamine oxidase. Biochemistry 16, 91-100. 3. Matsui, I., Wiegand, L., and Pegg, A. E. (1981) Properties of spermidine N-acetyltrans-ferase from livers of...

Intracellular Ca2 and Polyamine Dependent ECadherin Expression

Polyamines modulate Ca2+ cyt by governing the driving force for Ca2+ influx via controlling activity of Kv channels (5,11) and that Ca2+ mediates distinct actions of intestinal epithelial cells after increased polyamines (6). Because IEC-6 cells do not express VDCC, the inhibition of Kv channel expression and resultant membrane depolarization through polyamine depletion decreases the driving force for Ca2+ influx and reduces Ca2+ cyt concentration. To test the possibility that polyamines...

Pancreas

In terms of its polyamine content, the pancreas is a highly unique mammalian organ, being the richest source of spermidine and displaying the highest molar ratio (about 10) of spermidine to spermine (35,36). The physiological importance of the very high sper-midine content in the pancreas is not known, but it may be related to the intense protein synthesis going on in this organ or to maintenance of pancreatic structural integrity. Relatively little is known about the role of the polyamines in...

CMyc and Polyamine Induced Transcription of the ECadherin Gene

The c-Myc protein is a nuclear transcription factor, and its gene expression absolutely requires polyamines (41,42). Recently, E-cadherin was identified as a c-Myc target gene and activation of c-Myc enhances E-cadherin gene transcription in a cell type-dependent manner (43). Our recent study provides new evidence showing that polyamines regulate transcription of the E-cadherin gene through c-Myc in normal intestinal epithelial cells. Depletion of cellular polyamines dramatically inhibits c-myc...

S

Cancer Research Europe

Experiments are now in progress to clarify the properties of the transmembrane proteins of polyamine-uptake systems. The uptake of polyamines decreases following their accumulation in E. coli. One of the reasons for this is that spermidine inhibits the ATPase activity of PotA. Overexpression of PotD in E. coli also inhibited the uptake of spermidine and the synthesis of PotABCD messenger RNA (mRNA) (32). A 50 inhibition of the transcription was observed with a molar...

Effects of Polyamines on Chromatin Structure

Because of their cationic nature and strong affinity for DNA, it has long been thought that polyamines contribute to overall chromosome conformation, potentially influencing gene transcription. Indeed, biophysical and biochemical studies have shown that the highly positively charged polyamines bind strongly to DNA and affect its conformational state (10,11). Use of photoaffinity polyamine analogs has revealed that polyamines bind to specific locations and change the helical twist of DNA in...

Rationale for the Design of Polyamine Fluoroprobes and Initial Studies

At least four laboratories have independently developed fluorescent polyamine analogs as probes to visualize the internalization mechanism and to assess the subcellu-lar distribution of polyamines. Either the terminal (N1) amino group (29,39) or central (N4) amino groups (25,26,40) of a polyamine backbone has been used for conjugating the fluorophore. It is noteworthy that the size of the fluorophore in the conjugates was substantial relative to the polyamine backbone. Except for the...

Importance of Arginase

We have demonstrated that H. pylori upregulates expression of inducible nitric oxide (NO) synthase (iNOS) in gastritis tissues in vivo (39) and in macrophages in vitro (3), and iNOS-derived NO had been implicated as a causal factor in macrophage apoptosis in response to lipopolysaccharide plus interferon-y (40). When we identified that H. pylori also upregulated the arginase II isoform of arginase, which competes with iNOS for the same substrate, L-arginine, we determined the respective roles...

Structural Features of Inward Rectifier K Channels

All K+ channels are tetrameric arrangements of a core pore-forming domain, consisting of two transmembrane helices, a pore loop, and cytoplasmic N- and C-termini (20). Voltage-gated K+ (Kv) channels contain additional transmembrane helices with charged residues that sense membrane voltage, but Kir channel subunits contain only the core domain. The two transmembrane helices of the core domain (M1 and M2, corresponding to the pore-forming S5 and S6 segments of Kv channels) line the pathway of ion...

Effects of Elevated Levels of ODC and Polyamines on Chromatin Modifying Enzymes

Histones function as transcriptional modulators as a result of sophisticated molecular mechanisms that regulate their modification (1-4,22). The transcriptional status of a gene hinges largely on the acetylation status of the nucleosomal histones bound to the gene. Addition of acetyl groups to lysines in the N-terminal tail domains of the core histones, catalyzed by HAT enzymes, destabilizes nucleosomes, allowing regulatory factors access to recognition elements (27). Conversely, deacetylation...

P

P21, inhibition of cell proliferation, 63-65 p53, polyamine depletion effects, gene induction, 52-55 growth inhibition, 58-60 messenger RNA stabilization, 55, 57, 70 protein stabilization, 57, 58 Pancreatitis, transgenic rodents, 470, 471 PAO, see A Acetylpolyamine oxidase PAOhl, see Spermine oxidase PENSpm, antitumor activity, 14-16 structure, 15 structure activity relationships, 15-18 synthesis, 14 PET, see Positron emission tomography PKC, see Protein kinase C PMF-1, see Polyamine-modulated...

Parameters of Apoptotic Pathways Altered by Polyamine Depletion

Tnf Induced Apoptotic Pathway

Figure 5 indicates the apoptotic pathways delineated in IEC-6 cells and the references to the effects of polyamines on the specific components of those pathways. Yuan et al. (26) found that polyamine depletion prevented the release of cytochrome c from mitochondria after camptothecin. This was accompanied by increased Bcl-2 and Bcl-XL and decreased Bax and -Bid in the cells incubated with DFMO. In addition to inhibiting caspase 3 activation, DFMO significantly reduces the activation of caspase...

Proenzyme Processing

AdoMetDC belongs to a class of pyruvoyl-dependent enzymes that includes aspar-tate decarboxylase, phosphatidylserine decarboxylase, and histidine decarboxylase (41). The catalytic activities of these enzymes rely on a covalently bound pyruvoyl group rather than the pyridoxal-5'-phosphate cofactor more commonly employed by amino acid decarboxylases. Pyruvoyl enzymes are synthesized as an inactive proenzyme that undergoes an apparently autocatalytic internal cleavage reaction to yield two...

Dysfunction of Intestinal Epithelial Barrier After Polyamine Depletion

Polyamine depletion results in dysfunction of the epithelial barrier as indicated by a decrease in transepithelial electrical resistance (TEER) and increase in paracellular permeability (25,26). To study the possible role of decreased intercellular junctions in intestinal epithelial paracellular barrier function, TEER and paracellular flux of membrane impermeable tracers across the confluent monolayer were examined in the presence or absence of polyamines (25). IEC-6 cells were grown in control...

How Do Polyamines Modulate the Innate Immune Response in the CNS

The physiological relevance of such an interaction between polyamines and the innate immune system in the brain has yet to be clearly established. Our data that support polyamines acting as proinflammatory molecules are in disagreement with reports in which spermine in vitro was found to inhibit cytokine synthesis in macrophages in culture (49,51,71). However, this discrepancy can be explain by the fact that our experiments were performed in an in vivo model in the brain where microglial cells,...

Pathological Left Ventricular Hypertrophy

Cardiac hypertrophy and its transition to heart failure is a complex illness, and the most life-threatening disease in Western societies. Cardiac hypertrophy is defined as an increase in heart size mainly from an increased volume of muscle cells. Growth of the cardiac mass may be a physiological response, as observed in increased exercise, From Polyamine Cell Signaling Physiology Pharmacology and Cancer Research Edited by J.-Y. Wang and R. A. Casero, Jr. Humana Press Inc., Totowa, NJ pregnancy,...

Adaptive Immunity

The adaptive immune response is considered the predetermined response to a previously identified immunological stimulus. Thus, the response is specific to a particular pathogen and involves immunological memory. However, the lines between adaptive and innate immunity are frequently blurred by the close interactions between pathways, such that stimulation of antigen-presenting cells, such as macrophages and dendritic From Polyamine Cell Signaling Physiology Pharmacology and Cancer Research...

Polyamine Transport in E coli

Polyamine Binds Dna

E. coli contain putrescine and spermidine. In addition, cadaverine and aminopropyl-cadaverine can function similarly to putrescine and spermidine (11). Therefore, we first tried to identify the genes encoding the transport proteins for putrescine, spermi-dine, and cadaverine. To identify these genes, a mutant that is deficient in polyamine transport was first isolated. Genes for polyamine transport systems were then isolated by transforming the mutant with DNA fragments using pACYC184 as a...

Impact of Polyamines in Neurodegenerative Diseases

As seen in Subheading 3, polyamines play a major role in the control of the cerebral innate immune response during microbial challenges. These data may have a major clinical impact and suggest DFMO as a potential therapeutic drug to restrain neurodegeneration in brain disorders associated with inflammation. Indeed, polyamine levels in the blood are higher in patients suffering of Alzheimer's disease or ALS than healthy people of same age (73). Furthermore, the ALS-Therapy Development Foundation...

Polyamine Pools and Cancer Prevention Eugene W Gerner and David E Stringer

Polyamine contents increase during epithelial carcinogenesis in both rodents and humans. The activity of ornithine decarboxylase (ODC), the first enzyme in polyamine synthesis in mammals, is increased in epithelial cancers, and ODC has become a target for cancer chemoprevention. Difluoromethylornithine (DFMO) is a specific inhibitor of ODC and is under evaluation in clinical trials for prevention of epithelial cancers, including those of the colon, prostate, and skin. Tissue, serum, and other...

Myc Target Genes

Myc target genes can be generally classified in two ways those that are repressed or induced by Myc or those that are direct vs indirect transcription targets. Identifying Myc targets has become a prodigious task because Myc regulates the expression of so many genes indeed, a Web site has been established to keep track of all of these targets (http www.myccancergene.org) (84). Currently, 1697 Myc target genes have been identified using biased approaches and unbiased tools, such as Serial...

Posttranslational Regulation

Mammalian ODC has a very fast turnover (9,10). Although the half-life of ODC is usually something between 30 and 60 min, it may be as short as a few minutes. The turnover of mammalian ODC is affected by the cellular polyamine levels (9,10,97). In the presence of polyamine excess, there is a marked increase in the degradation rate of ODC, whereas when cells are depleted of their polyamines, the degradation of ODC is decreased. As with other cellular proteins with a fast turnover, ODC is degraded...

Polyamines and Cytoskeletal Regulation During Intestinal Epithelial Restitution

Johnson 1. Introduction Damage to gastrointestinal epithelia can result from infection (ulcer), chemical agents (alcohol, drugs), or mechanical forces (stretching), and immediate repair is required to restore the epithelial barrier against luminal antigens. The mucosa of the gastrointestinal tract has the unique ability to repair itself rapidly after damage. Mucosal repair consists of two phases. Early mucosal restitution is the rapid re-establishment of epithelial...

Polyamine Depletion Associated With Increased p53 Gene Expression

Considerable interest over the past several years has been devoted to the mechanisms of negative growth control. In the small intestinal mucosa, for example, mitotic activity is confined to the crypt region and must be finely tuned to the rapid rate of loss of mature enterocytes from the villous tip. The o reti c al ly, mucosal epithelial cell turnover is controlled by both positive and negative control mechanisms, with stimulation or suppression of cell renewal mediated through...

Expression of the TGFb Receptor Gene and Sensitivity to Exogenous TGFb

Polyamine-deficient cells are more sensitive to growth inhibition when they are exposed to exogenous TGF-b 16 . Because IECs produce both TGF-b receptors and ligands 49,56 , it is possible that the regulation of cellular responsiveness relies on the production of active TGF -b and its presentation to signaling receptors. Rao and colleagues 11 have further reported that depletion of cellular polyamines increases levels of the TGF-bRI mRNA and protein, but has no effect on the TGF-bRII receptor...

Spermine as a Biomarker for Malignant Behavior of Prostatic Cancer

Spermine is present in the prostate in high concentrations 10-20 mM . Several studies suggest that a reduction of spermine levels may result from shifts in cellular behavior of prostatic cancer cells 38-40 . Polyamine measurements in our panel of cultured cells or xenografts of human prostatic cell lines showed that the less differentiated cell lines e.g., PC-324, PC-339 contained clearly less spermine than the tumor cell lines with differentiation characteristics resembling androgen...

Nsaid

Ornithine decarboxylase is a potential surrogate endpoint biomarker for human cancer prevention trials. factors, AP-1 and nuclear factor NF -k B, linked to the expression of TNF-a-induced genes involved in immunity and inflammatory responses, and control of cellular proliferation, differentiation, and apoptosis 49,50 . The role of AP-1 and NF-kB activation in TNF -a signal transduction pathway to the development of SCC and PKCe transgenic mice is unknown and is important in view of the...

Polyamine Transport in Yeast

Polyamine uptake in yeast is energy-dependent and the Km value for putrescine, spermidine, and spermine are 770, 8.3, and 18 iM, respectively. A distinctive feature of the polyamine transport system in Saccharomyces cerevisiae is that it is strongly inhibited by Mg2 42 . Thus in a Mg2 -limited medium, polyamines, especially sper-mine, overaccumulate in cells and become toxic for growth. We isolated a mutant YTM22-8 whose growth was tolerant to spermine in a Mg2 -limited medium. This mutant was...

Mechanism of Action of Antipolyamine Therapy

Tumor Mechanism Study Presentation

Figure 4 illustrates our ongoing research on the mechanisms of antitumor action of DFMO in breast cancer cells. We have shown that, in MDA-MB-435 cells, DFMO activates the MAPK pathway, as evidenced by increased phosphorylation of extracellular regulated kinase ERK -1 and ERK-2 46 . A similar effect of DFMO has been reported by us in MCF-10A human breast epithelial cells 47 and by other investigators in intestinal epithelial and melanoma cells where this effect has been connected to induction...

Smads are the Downstream Intracellular Effectors of Activated TGFbTGFb Receptors After Polyamine Depletion

Polyamines Dna

The Smad proteins are a family of transcriptional activators that are critical for transmitting the TGF-b superfamily signals from the cell surface to the nucleus 51,52 . Based on distinct functions, Smads are grouped into three classes the receptor-regulated Smads R-Smads , Smad2, and Smad3 the common Smad co-Smad , Smad4 and the inhibitory Smads I-Smads , Smad6, and Smad7 53-55 . All TGF-b family members, including TGF - bs, activins, and bone morphogenetic proteins, use TGF-bRI and TGF-bRII...

Polyamine Uptake and Sequestering by Mammalian Cells A Two Step Two Compartment Process

Polyamine Cancer

In our initial studies with spermidine-C2-BODIPY Spd-C2-BODIPY Fig. 1 , the hypothesis that accumulation of the polyamine probe into PSVs proceeds via receptor-mediated endocytosis was initially evaluated by measuring the extent of its colocaliza-tion with fluorescent transferrin 26 . The extent of such colocalization was significant, but only partial, and limited to the largest PSVs. Thus PSVs did include vesicles converging with the recycling endocytosis pathway, but other structures were...

1

Mediators of innate immunity and polyamines. This figure depicts a central role of macrophages in the innate immune response and the potential involvement of polyamines in these events. Macrophages are first activated by pathogen components, called pathogen-associated molecular patterns. Then myeloid cells release proinflammatory cytokines TNF-a, interleukin 1, interleukin 12 , which can activate natural killer NK cells. NK cells release the cytokine interferon-g IFN-g , which further...

Intercellular Junctions and Epithelial Barrier

Integrity of the intestinal barrier depends on a complex of proteins composing different intercellular junctions, which include tight junctions, adherens junctions, and desmosomes. Over the past several years, our group and others have demonstrated that polyamines are necessary for expression and functions of intercellular junction proteins, especially adherens junctions, and play a critical role in maintenance of intestinal epithelial integrity 24-26 . The tight junction is the most apical of...

Functional Role of Polyamines in a Model of Inflammation Induced Neurotoxicity

To understand the functional role of polyamines in the CNS as being neurotoxic or neuroprotective molecules, we used a model of neurodegeneration caused by an exaggerated inflammatory response. Glucocorticoids are one of the most powerful endogenous suppressors for the innate immune response 68 and are essential molecules to avoid exaggerated responses during immunogenic challenges. As expected, the inflammatory response lasted longer in the brain of animals that received the glucocorticoid...

BDNA to ZDNA Transition and Stabilization

Molecular structure of left-handed Z-DNA was established 25 yr ago by single crystal X-ray crystallography of alternating purine pyrimidine sequences 16 . Z-DNA has a zig-zag arrangement of the sugar phosphate backbone with only deep minor grooves and no discernible major groove Fig. 4 . In contrast, the phosphate groups lie on a smooth helical line in B-DNA with minor and major grooves. Initially, Z-DNA was thought to be a nonphysiological structure, because 3 to 4 MNaCl was required to induce...

Contributors

Virtanen Institute for Molecular Sciences, University of Kuopio, Kuopio, Finland Allan A. Ancheta Department of Biochemistry, University of California, Riverside, CA Marie Audette Oncological and Molecular Endocrinology Research Center and Department of Medical Biology, Laval University, Quebec, Canada Naveen Babbar The Sidney Kimmel Comprehensive Cancer Center, Johns Hopkins University School of Medicine, Baltimore, MD Craig V. Byus Department of Biochemistry, University of...