Gene expression of ornithine decarboxylase in L1210 leukaemia cells exposed to <scp>dl</scp>-2-difluoromethylornithine in the presence of cadaverine

Alhonen‐Hongisto, Leena; Sinervirta, Riitta; Jänne, Olli A.; Jänne, Juhani

doi:10.1042/bj2320605

Cited by 21 publications

(7 citation statements)

References 18 publications

(8 reference statements)

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“…As to the specificity of polyamine requirements for cell growth, the present results, together with earlier experimental findings (Alhonen-Hongisto et al, 1982, 1985b suggest that, although the natural polyamines can be replaced to a large extent by cadaverine-based polyamines, in terms of supporting animal .cell growth a certain minimum production of natural polyamines appears to be required. This is exemplified by the fact that, although the total content of cadaverine was fully comparable with that of the natural polyamines in the parental cells (Table 2), these cells overproduced ODC.…”

Section: Discussionsupporting

confidence: 68%

“…In addition to the 'conventional' gene amplification in mouse (McConlogue et al, 1984;Kahana & Nathans, 1984;Alhonen-Hongisto et al, 1985a) and in human (Leinonen et al, 1987) tumour cells, an overproduction of ODC can occur in the absence of any increase in the gene copy number. We found (Alhonen-Hongisto et al, 1985b) that an exposure of mouse L1210 leukaemia cells to DFMO in the presence of micromolar concentrations of cadaverine led to a strikingly enhanced expression of ODC, with minimum changes in the gene dosage of the enzyme. Similarly, McConlogue et al (1986) showed that an overproduction of ODC can be based on an enhanced accumulation of mRNA in the absence of any gene amplification, or even on a more efficient translation of normal amounts of the message.…”

Section: Introductionmentioning

confidence: 79%

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Cadaverine supplementation during a chronic exposure to difluoromethylornithine allows an overexpression, but prevents gene amplification, of ornithine decarboxylase in L1210 mouse leukaemia cells

Alhonen‐Hongisto

Hirvonen

Sinervirta

et al. 1987

Biochemical Journal

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We recently selected a variant mouse L1210 leukaemia-cell line overproducing ornithine decarboxylase (ODC) (EC 4.1.1.17) as a result of chronic exposure to 2-difluoromethylornithine (DFMO) in the presence of micromolar concentrations of cadaverine. These cells, now grown for more than 2 years in the presence of DFMO and cadaverine, continued to accumulate ODC-specific mRNA in an amount 30-50 times higher than that in the parental cells, yet showing practically no changes in the gene dosage for the enzyme. However, analysis of the genomic DNA with the isoschizomeric restriction enzymes HpaII and MspI revealed that the ODC sequences in the overproducer cells were hypomethylated in comparison with the parental cells. The natural polyamines (putrescine, spermidine and spermine) were almost totally replaced by cadaverine and aminopropylcadaverine. Omission of cadaverine from the culture medium, but leaving 10 mM-DFMO, resulted in an about 10-fold ODC gene amplification within a few weeks. The accumulation of ODC mRNA was enhanced by the same factor. Concomitantly, the content of the natural polyamines was almost normalized, representing about 65% of that found in the parental cells. The present results suggest that, under a given selection pressure, an overproduction of the target gene product may be primarily based on an enhanced transcriptional activity, possibly associated with hypomethylation and, if not sufficient, a secondary amplification of the active gene occurs.

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Section: Discussionsupporting

confidence: 68%

Section: Introductionmentioning

confidence: 79%

Cadaverine supplementation during a chronic exposure to difluoromethylornithine allows an overexpression, but prevents gene amplification, of ornithine decarboxylase in L1210 mouse leukaemia cells

Alhonen‐Hongisto

Hirvonen

Sinervirta

et al. 1987

Biochemical Journal

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“…Post‐transcriptional stabilization of ODC has been suggested to be one mechanism by which ODC mRNA levels increase in response to DFMO treatment in COLO 320 cells [Celano et al, 1989]. Others have suggested that increases in ODC gene amplification can occur in response to DFMO treatment [Alhonen‐Hongisto et al, 1985], whether this occurs in C2 cells in response to DFMO treatment is not known. The increase in ODC mRNA levels in highly malignant C2 cells, in response to alterations in cellular polyamine levels, may represent a compensatory response on behalf of C2 cells to the stresses imposed by cellular polyamine depletion.…”

Section: Discussionmentioning

confidence: 99%

Transformation by H-ras can result in aberrant regulation of ornithine decarboxylase gene expression by transforming growth factor-?1

2001

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Inhibition of DNA synthesis and cell proliferation is frequently lost during malignant transformation and occasionally, tumour cell proliferation is actually stimulated by transforming growth factor beta(1) (TGF-beta(1)). The present study demonstrates a novel link between alterations in TGF-beta(1) regulation during cellular transformation and malignant conversion and the expression of ornithine decarboxylase (ODC) which is a key rate limiting activity in the biosynthesis of polyamines and which is an enzyme that plays an important role in cell growth and differentiation. H-ras transformed mouse 10T(1/2) cell lines of varying degrees of malignant potential were examined for possible TGF-beta(1)-mediated alterations in ODC expression. Selective induction of ODC gene expression occurred. This induction was dependent upon the cellular phenotype expressed and the mechanisms responsible for the regulation of the TGF-beta(1)-mediated alterations in ODC expression varied as a function of malignant potential. The TGF-beta(1)-mediated alterations in ODC gene expression involves de novo protein synthesis, transcriptional, and post-transcriptional events. Evidence is also presented to suggest a possible role for protein kinase C-mediated events, protein phosphatases, and G-protein-coupled events in the TGF-beta(1)-mediated regulation of ODC expression in H-ras transformed cells. Evidence is also presented to suggest a possible role for cellular polyamines in the TGF-beta(1)-mediated alterations in ODC expression in H-ras transformed cells. Additionally, alterations in cellular polyamines were shown to influence TGF-beta(1) gene expression in H-ras transformed cells and that these alterations occurred, in part, through post-transcriptional events. The TGF-beta(1)-mediated regulation of ODC expression in H-ras transformed cells of varying degrees of malignant potential appears to be complex, multifaceted, and interactive. This study illustrates the importance of TGF-beta(1)-mediated regulation of ODC expression as a result of H-ras mediated cellular transformation and malignant progression, and further suggests that this TGF-beta(1)-mediated regulation constitutes an integral part of an altered growth regulatory program.

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“…Some mechanism other than amplification, therefore, must exist through which a variant cell can increase its ODC mRNA pools, presumably by increased rate of transcription, increased efficiency of processing or transport, or increased stability. A variant of cultured L1210 leukemia cells that has an increased level of ODC mRNA without a concomitant increase in ODC gene copy number has recently been reported (2). Variant cells that overproduce mRNA for argininosuccinate synthetase (34) or for aryl hydrocarbon hydroxylase (12), without amplification of the corresponding gene, have also been described.…”

Section: Discussionmentioning

confidence: 99%

Multiple mechanisms are responsible for altered expression of ornithine decarboxylase in overproducing variant cells.

McConlogue¹,

Dana²,

Coffino³

1986

Mol. Cell. Biol.

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We selected and characterized a series of mouse S49 cell variants that overproduce ornithine decarboxylase (ODC). Previously, we described variants that have an amplified ODC gene and produce about 500-fold more ODC than the wild-type cells of origin (L. McConlogue and P. Coffino, J. Biol. Chem. 258:12083-12086, 1983). We examined a series of independent variants that overproduce ODC to a lesser degree and found that a number of mechanisms other than gene amplification are responsible for the increased ODC activity. Variants were selected for resistance to 0.1 mM difluoromethylornithine, an inhibitor of ODC, by either a single or a multistep process. AU showed increased ODC activity and increased ODC mRNA steady-state levels. The half-life of the enzyme was not increased in any of the variants. In one class of variant the increase of ODC mRNA was sufficient to account for ODC overproduction. In a second class, the rate of synthesis of ODC polypeptide per ODC mRNA was at least four-to eightfold higher than that in wild-type cells. Therefore, these variants were altered in the translatability of ODC mRNA. Southern analysis showed that gene amplification does not account for the increased ODC mRNA levels in any of the variants. In both variant and wild-type cells, ODC activity was responsive to changes in polyamine pools; activity was reduced following augmentation of pool size. This change in activity was associated with modification of the rate of synthesis and degradation of ODC but no change in the level of ODC mRNA.In animal cells ornithine decarboxylase (ODC) is an essential enzyme, the activity of which is regulated by a wide variety of hormonal, developmental, and cell growth-related stimuli (35). It is the initial and rate-limiting enzyme in the synthesis of polyamines; its product, putrescine, is the precursor of the polyamines spermidine and spermine. In general, cells that are proliferating exhibit more activity than their nonproliferating counterparts. There is evidence for multiple forms of regulation of ODC activity, including mRNA steady-state level, polypeptide synthesis, and polypeptide turnover.In S49 mouse T-lymphoma cells cyclic AMP modulates cell growth and ODC activity (7,10,22). ODC activity is also changed by treatments that modulate cellular polyamine levels, such as exposure to putrescine, the direct product of ODC. We generated ODC-overproducing variants of S49 cells by selecting for cells resistant to the ODC inhibitor difluoromethylornithine (DFMO) (19,20). These variants allowed us to identify the ODC polypeptide and to clone the ODC cDNA (21) DFMO. Clone Z.12 was selected from an unmutagenized wild-type (clone 24.3.2) S49 cell population by serially increasing the concentration of DFMO as described previously (20). The D2 clones were selected from a mutagenized population in a single step as follows. Wild-type cells were mutagenized with 2.5 ,g of N-methyl-N'-nitro-N-nitrosoguanidine per ml for 4.5 h. This treatment resulted in 30% cell survival and induction of resistance to 6-thio...

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Gene expression of ornithine decarboxylase in L1210 leukaemia cells exposed to dl-2-difluoromethylornithine in the presence of cadaverine

Cited by 21 publications

References 18 publications

Cadaverine supplementation during a chronic exposure to difluoromethylornithine allows an overexpression, but prevents gene amplification, of ornithine decarboxylase in L1210 mouse leukaemia cells

Cadaverine supplementation during a chronic exposure to difluoromethylornithine allows an overexpression, but prevents gene amplification, of ornithine decarboxylase in L1210 mouse leukaemia cells

Transformation by H-ras can result in aberrant regulation of ornithine decarboxylase gene expression by transforming growth factor-?1

Multiple mechanisms are responsible for altered expression of ornithine decarboxylase in overproducing variant cells.

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