Formation of a compensatory polyamine by Escherichia coli polyamine-requiring mutants during growth in the absence of polyamines

Igarashi, Kazuei; Kashiwagi, Keiko; Hamasaki, Hidehisa; Miura, Atsuko; Kakegawa, Tomohito; Hirose, Sachiko; Matsuzaki, Shinya

doi:10.1128/jb.166.1.128-134.1986

Cited by 178 publications

(100 citation statements)

References 28 publications

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“…Polyamine levels were measured by HPLC as described previously. 48 Cells treated with doxorubicin or cisplatin, or untreated controls (6 Â 10 5 cells) were harvested and polyamines were extracted from cell lysate with 5% trichloroacetic acid and the supernatant after centrifugation at 27 000 Â g for 15 min at 4 1C was used for polyamine measurement by HPLC. The precipitate was used for measurement of protein content.…”

Section: Discussionmentioning

confidence: 99%

Suppression of acetylpolyamine oxidase by selected AP-1 members regulates DNp73 abundance: mechanistic insights for overcoming DNp73-mediated resistance to chemotherapeutic drugs

et al. 2014

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Enhanced resistance to chemotherapy has been correlated with high levels of Delta-Np73 (DNp73), an anti-apoptotic protein of the p53 tumor-suppressor family which inhibits the pro-apoptotic members such as p53 and TAp73. Although genotoxic drugs have been shown to induce DNp73 degradation, lack of mechanistic understanding of this process precludes strategies to enhance the targeting of DNp73 and improve treatment outcomes. Antizyme (Az) is a mediator of ubiquitin-independent protein degradation regulated by the polyamine biosynthesis pathway. We show here that acetylpolyamine oxidase (PAOX), a catabolic enzyme of this pathway, upregulates DNp73 levels by suppressing its degradation via the Az pathway. Conversely, downregulation of PAOX activity by siRNA-mediated knockdown or chemical inhibition leads to DNp73 degradation in an Az-dependent manner. PAOX expression is suppressed by several genotoxic drugs, via selected members of the activator protein-1 (AP-1) transcription factors, namely c-Jun, JunB and FosB, which are required for stress-mediated DNp73 degradation. Finally, chemical-and siRNA-mediated inhibition of PAOX significantly reversed the resistant phenotype of DNp73-overexpressing cancer cells to genotoxic drugs. Together, these data define a critical mechanism for the regulation of DNp73 abundance, and reveal that inhibition of PAOX could widen the therapeutic index of cytotoxic drugs and overcome DNp73-mediated chemoresistance in tumors.

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Section: Discussionmentioning

confidence: 99%

Suppression of acetylpolyamine oxidase by selected AP-1 members regulates DNp73 abundance: mechanistic insights for overcoming DNp73-mediated resistance to chemotherapeutic drugs

et al. 2014

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“…The concentration and specific activities of substrates were as follows: 0.5 mM Measurement of Polyamine Contents in Whole Cells-YPH499, ⌬spe1, ⌬dur3⌬spe1, ⌬sam3⌬spe1, ⌬dur3⌬sam3⌬spe1, ⌬gap1⌬spe1, and ⌬agp2⌬spe1 mutants were grown in the CSD medium in the absence and presence of 0.05 mM putrescine and harvested at A 540 ϭ 0.25. Polyamines were extracted by treatment with 10% (w/v) trichloroacetic acid at 70°C for 1 h. Polyamine contents were determined by high pressure liquid chromatography as described previously (29). Protein was determined by the Bradford method (30).…”

Section: Methodsmentioning

confidence: 99%

Polyamine Uptake by DUR3 and SAM3 in Saccharomyces cerevisiae

Uemura

Kashiwagi

Igarashi

2007

Journal of Biological Chemistry

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It has been reported that GAP1 and AGP2 catalyze the uptake of polyamines together with amino acids in Saccharomyces cerevisiae. We have looked for polyamine-preferential uptake proteins in S. cerevisiae. DUR3 catalyzed the uptake of polyamines together with urea, and SAM3 was found to catalyze the uptake of polyamines together with S-adenosylmethionine, glutamic acid, and lysine. Polyamine uptake was greatly decreased in both DUR3-and SAM3-deficient cells. The K m values for putrescine and spermidine of DUR3 were 479 and 21.2 M, respectively, and those of SAM3 were 433 and 20.7 M, respectively. Polyamine stimulation of cell growth of a polyamine requiring mutant, which is deficient in ornithine decarboxylase, was not influenced by the disruption of GAP1 and AGP2, but it was diminished by the disruption of DUR3 and SAM3. Furthermore, the polyamine stimulation of cell growth of a polyamine-requiring mutant was completely inhibited by the disruption of both DUR3 and SAM3. The results indicate that DUR3 and SAM3 are major polyamine uptake proteins in yeast. We previously reported that polyamine transport protein kinase 2 regulates polyamine transport. It was found that DUR3 (but not SAM3) was activated by phosphorylation of Thr 250 , Ser 251 , and Thr 684 by polyamine transport protein kinase 2.Polyamines (putrescine, spermidine, and spermine) in cells, which are essential for cell growth, are regulated by biosynthesis, degradation, and transport (1-4). With regard to polyamine transport, the properties of four polyamine transport systems were characterized in Escherichia coli (5-8). They include spermidine-preferential and putrescine-specific uptake systems as well as PotE (involved in the excretion of putrescine by a putrescine-ornithine antiporter activity) and CadB (involved in the excretion of cadaverine by a cadaverine-lysine antiporter activity). The former two transport systems function at neutral pH (2), whereas the latter two transport systems at acidic pH (9). In Saccharomyces cerevisiae, we identified four genes that encode polyamine excretion proteins TPO1-TPO4, mainly located on the plasma membrane (10 -12). We also found that UGA4 (located on vacuoles) can catalyze the uptake of ␥-aminobutyric acid and putrescine (13), and TPO5 (located on Golgi or post-Golgi secretory vesicles) can catalyze the excretion of polyamines (14). Furthermore, we reported that GAP1, located on the plasma membrane, can catalyze the uptake of putrescine and spermidine together with the uptake of amino acids (15). Although it has been reported that AGP2 can selectively catalyze the uptake of spermidine (16), there is also a report that AGP2 functions as an amino acid permease (17). In this study, we looked for proteins that can preferentially catalyze the uptake of polyamines in S. cerevisiae. We found that DUR3 can catalyze the uptake of polyamines together with urea, and SAM3 (which belongs to the family of amino acid polyamine-organocation transporters (18)) can catalyze the uptake of putrescine and spermidine together wit...

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“…Cells were homogenized with a Teflon homogenizer, and polyamines (putrescine, spermidine and spermine) were extracted by the treatment with 0.3ml of 5% (w/v) trichloroacetic acid. Polyamine levels in the cellular extracts were measured by HPLC as previously reported (Igarashi et al 1986). …”

Section: Measurement Of Intracellular Polyamine Contents (Hplc: High-mentioning

confidence: 99%

Agmatine Suppresses Mesangial Cell Proliferation by Modulating Polyamine Metabolism

Eto

Isome

Sano

et al. 2006

Tohoku J. Exp. Med.

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Polyamines play an essential role in the growth and differentiation of mammalian cells. The depletion of intracellular polyamines results in the suppression of growth. Proliferation of glomerular mesangial cells (MC) is the most common pathologic change in many forms of glomerulonephritis. Agmatine is a metabolite of arginine via arginine decarboxylase (ADC), highly expressed in the kidney, and unique in its capacity to suppress intracellular polyamine levels required for proliferation. As agmatine enters mammalian cells via the polyamine transport system, its antiproliferative effects may preferentially target cells with increased proliferative kinetics. In the present study, we evaluated the antiproliferative effects of agmatine on human MC in vitro. MC proliferation was stimulated with 20% fetal bovine serum (FBS) or platelet-derived growth factor (PDGF-BB, 20 ng/ml). Cell proliferation was measured using the (4.3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl tetrazolium bromide) (MTT) proliferation assay. Intracellular polyamine levels were assayed by high performance liquid chromatography, and cell death was assessed by cellular DNA fragmentation enzyme-linked immunosorbent assay. The MTT proliferation assay showed that agmatine significantly suppressed proliferation of human MC treated with 20% FBS or 5% FBS + PDGF as compared to human MC treated with 5% FBS. Polyamine levels were markedly lower in cells treated with agmatine, and proliferation was rescued by administration of putrescine. The fragmented DNA was hardly detected in agmatine-treated human MC. In summary, human MC stimulated to increase their proliferative kinetics are significantly more sensitive to the antiproliferative effects of agmatine than normally cultured cells. Suppressed proliferation of the agmatine-treated human MC is not due to increased cell death. These results suggest that agmatine is a promising drug candidate for the treatment of human mesangial proliferative glomerulonephritis.

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Formation of a compensatory polyamine by Escherichia coli polyamine-requiring mutants during growth in the absence of polyamines

Cited by 178 publications

References 28 publications

Suppression of acetylpolyamine oxidase by selected AP-1 members regulates DNp73 abundance: mechanistic insights for overcoming DNp73-mediated resistance to chemotherapeutic drugs

Suppression of acetylpolyamine oxidase by selected AP-1 members regulates DNp73 abundance: mechanistic insights for overcoming DNp73-mediated resistance to chemotherapeutic drugs

Polyamine Uptake by DUR3 and SAM3 in Saccharomyces cerevisiae

Agmatine Suppresses Mesangial Cell Proliferation by Modulating Polyamine Metabolism

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