Methotrexate Resistance in Datura innoxia (Uptake and Metabolism of Methotrexate in Wild-Type and Resistant Cell Lines)

Wu, Keqiang; Atkinson, Ian; Cossins, Edwin A.; King, John

doi:10.1104/pp.101.2.477

Cited by 15 publications

(8 citation statements)

References 21 publications

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“…This observation could be attributed mainly if not exclusively to MTX, which is known to act as an antiproliferative agent in plant cells (Wu et al, 1993). Indeed, exposure to MTX (100 mM) alone resulted in growth arrest of Arabidopsis cultures whereas cells treated with sulfanilamide (100 mM) alone grew similarly to control cells.…”

Section: Induction Of Folate Deficiency In Arabidopsis Cellsmentioning

confidence: 91%

“…Thus, Prabhu et al (1998) exposed Arabidopsis (Arabidopsis thaliana) plants to folate antagonists and determined that a continuous supply of folates was essential to maintain high rates of Ser synthesis. Also, plant cell cultures treated with the folate analog methotrexate (MTX) displayed an important inhibition of their growth (Wu et al, 1993), thus emphasizing the crucial role of folates in cell division and nucleotide synthesis. Apart from these data, the consequences of modification of folates pool size on the other pathways of C1 metabolism, particularly the synthesis of Met, are largely unknown.…”

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confidence: 99%

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Regulation of One-Carbon Metabolism in Arabidopsis: The N-Terminal Regulatory Domain of Cystathionine γ-Synthase Is Cleaved in Response to Folate Starvation

et al. 2007

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In all organisms, control of folate homeostasis is of vital importance to sustain the demand for one-carbon (C1) units that are essential in major metabolic pathways. In this study we induced folate deficiency in Arabidopsis (Arabidopsis thaliana) cells by using two antifolate inhibitors. This treatment triggered a rapid and important decrease in the pool of folates with significant modification in the distribution of C1-substituted folate coenzymes, suggesting an adaptive response to favor a preferential shuttling of the flux of C1 units to the synthesis of nucleotides over the synthesis of methionine (Met). Metabolic profiling of folate-deficient cells indicated important perturbation of the activated methyl cycle because of the impairment of Met synthases that are deprived of their substrate 5-methyl-tetrahydrofolate. Intriguingly, S-adenosyl-Met and Met pools declined during the initial period of folate starvation but were further restored to typical levels. Reestablishment of Met and S-adenosylMet homeostasis was concomitant with a previously unknown posttranslational modification that consists in the removal of 92 amino acids at the N terminus of cystathionine g-synthase (CGS), the first specific enzyme for Met synthesis. Rescue experiments and analysis of different stresses indicated that CGS processing is specifically associated with perturbation of the folates pool. Also, CGS processing involves chloroplastic serine-type proteases that are expressed in various plant species subjected to folate starvation. We suggest that a metabolic effector, to date unidentified, can modulate CGS activity in vivo through an interaction with the N-terminal domain of the enzyme and that removal of this domain can suppress this regulation.

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Section: Induction Of Folate Deficiency In Arabidopsis Cellsmentioning

confidence: 91%

mentioning

confidence: 99%

Regulation of One-Carbon Metabolism in Arabidopsis: The N-Terminal Regulatory Domain of Cystathionine γ-Synthase Is Cleaved in Response to Folate Starvation

et al. 2007

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“…MTX is an analog of dihydrofolate that primarily acts on dihydrofolate reductase (DHFR), an enzyme involved in the de novo synthesis of THF and in the regeneration of reduced folate coenzymes from oxidized dihydrofolate species. Previous studies have shown that MTX is useful to manipulate folate pools in plants (Wu et al, 1993;Prabhu et al, 1998;Loizeau et al, 2007). In our study, MTX was used at 100 mM to induce rapid perturbation of folate homeostasis in Arabidopsis cells, and the dynamics of global transcriptomic changes was followed 2, 6, and 24 h following drug addition to the culture medium.…”

Section: Resultsmentioning

confidence: 99%

“…By exposing Arabidopsis (Arabidopsis thaliana) plants to folate antagonists, Prabhu et al (1998) determined that a continuous supply of folates was essential to maintain high rates of Ser synthesis during the photorespiratory process. Also, plant cell cultures treated with the folate analog methotrexate (MTX) displayed an important inhibition of their growth (Wu et al, 1993), thus emphasizing the crucial role of folates in nucleotide synthesis and cell division. Last, folate limitation in Arabidopsis cells was associated with important perturbation of the activated methyl cycle, which corresponds to a recycling of Met and AdoMet following methylation reactions (Loizeau et al, 2007).…”

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confidence: 99%

A Genome-Wide and Metabolic Analysis Determined the Adaptive Response of Arabidopsis Cells to Folate Depletion Induced by Methotrexate

et al. 2008

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Control of folate homeostasis is essential to sustain the demand for one-carbon (C1) units that are necessary for major biological functions, including nucleotide synthesis and methylation reactions. In this study, we analyzed the genome-wide and metabolic adaptive response of Arabidopsis (Arabidopsis thaliana) cells to folate depletion induced by the antifolate methotrexate. Drug treatment induced a response typical to xenobiotic stress and important changes in folate content and composition. This resulted in a reduction of cell division and primary energy metabolism that was likely associated with perturbation of nucleotide homeostasis. Through a modification of serine metabolism, folate depletion also induced O-acetylserine accumulation and mimicked sulfur deficiency response. The major adaptive response to folate limitation concerned the composition of the folate pool rather than the intracellular level of cofactors. Thus, no significant change in the expression of genes involved in cofactor synthesis, degradation, or trafficking was observed. However, changes in the distribution of C1 derivative pools and increased expression levels for transcripts coding enzymes manipulating C1 moieties in plastids suggested a reorientation of C1 units toward the synthesis of purine and thymidylate. Also, no genomic or metabolic adaptation was built up to counterbalance the major impairment of the methyl index, which controls the efficiency of methylation reactions in the cell. Together, these data suggested that the metabolic priority of Arabidopsis cells in response to folate limitation was to shuttle the available folate derivatives to the synthesis of nucleotides at the expense of methylation reactions.

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“…1997), petunia (Barg et al. 1984) and Datura innoxia (Wu et al. 1993) cell cultures, in which DHFR gene amplification, over‐production and impairment of drug membrane transport were the main causes of MTX insensitivity, respectively.…”

Section: Introductionmentioning

confidence: 99%

Isolation and characterisation of an antifolate insensitive (afi1) mutant of Arabidopsis thaliana

et al. 2012

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Antifolates can impair the synthesis and/or function of folates in living organisms. Mechanisms of resistance or tolerance to antifolates have been mainly described in plants using the drug methotrexate. In this work, the antifolate trimethoprim (TMP) was used with the aim of revealing a novel mechanism of resistance. EMS mutagenised seeds from Arabidopsis were screened to isolate individuals insensitive to TMP. Genetic analysis revealed a homozygous recessive mutation that segregates with the phenotype of tolerance to 50 μm TMP. Mapping analysis localised the mutation at the end of the short arm of chromosome 3. Preliminary characterisation demonstrated up-regulation of several genes from the folate biosynthetic pathway in the TMP insensitive mutant, and a slight increase in total folate content in the mutant as compared with the Col-0 control. Moreover, sequence analysis of the DHFR (dihydrofolate reductase) genes, which encode a known target for resistance to antifolates, did not reveal any changes. This study is the first report of a stable mutant insensitive (afi1) to the antifolate trimethoprim in plants, and suggests the existence of a novel mechanism of resistance to antifolates.

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Methotrexate Resistance in Datura innoxia (Uptake and Metabolism of Methotrexate in Wild-Type and Resistant Cell Lines)

Cited by 15 publications

References 21 publications

Regulation of One-Carbon Metabolism in Arabidopsis: The N-Terminal Regulatory Domain of Cystathionine γ-Synthase Is Cleaved in Response to Folate Starvation

Regulation of One-Carbon Metabolism in Arabidopsis: The N-Terminal Regulatory Domain of Cystathionine γ-Synthase Is Cleaved in Response to Folate Starvation

A Genome-Wide and Metabolic Analysis Determined the Adaptive Response of Arabidopsis Cells to Folate Depletion Induced by Methotrexate

Isolation and characterisation of an antifolate insensitive (afi1) mutant of Arabidopsis thaliana

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