Ornithine Decarboxylase Pathway Research Articles

Identification and characterization of polyamine metabolism in citrus in response to 'Candidatus Liberibacter asiaticus' infection.

Citrus Huanglongbing, one of the most devastating citrus diseases, is caused by Candidatus Liberibacter asiaticus (CLas). Polyamines are aliphatic nitrogen-containing compounds that play important roles in disease resistance. However, the role of polyamine metabolism in the tolerance of citrus to infection with CLas have not been extensively studied. We used HPLC and UPLC-Q/TOF-MS to detect the contents of nine polyamine metabolism-related compounds (PMRCs) in six citrus cultivars with varying levels of tolerance to CLas. Moreover, we systematically detected the changes of PMRC and H2O2 contents, and compared the gene expression levels and activities of enzymes involved in the polyamine metabolic pathway among healthy, asymptomatic, and symptomatic leaves of Newhall navel oranges infected with CLas. The tolerant and moderately tolerant varieties showed higher PMRC levels than those of susceptible varieties. Compared with the healthy group, the symptomatic group showed significantly increased contents of arginine, ornithine, γ-aminobutyric acid, and putrescine by approximately 180, 19, 1.5, and 0.2 times, respectively, and up-regulated expression of biosynthetic genes. Arginase and ornithine decarboxylase enzyme activities were the highest in the symptomatic group, while arginine decarboxylase and agmatine deiminase enzyme activities were the highest in the asymptomatic group. The two polyamine biosynthetic pathways showed different trends with the increase of the titer of CLas, indicating that polyamines were mainly synthesized through the arginine decarboxylase pathway in the asymptomatic leaves, and were synthesized via the ornithine decarboxylase pathway in symptomatic leaves. These findings provide new insight into the changes of polyamine metabolism in citrus infected with CLas.

Genomic analysis of the polyamine biosynthesis pathway in duckweed Spirodela polyrhiza L.: presence of the arginine decarboxylase pathway, absence of the ornithine decarboxylase pathway, and response to abiotic stresses

Main conclusionIdentification of the polyamine biosynthetic pathway genes in duckweed S. polyrhiza reveals presence of prokaryotic as well as land plant-type ADC pathway but absence of ODC encoding genes. Their differential gene expression and transcript abundance is shown modulated by exogenous methyl jasmonate, salinity, and acidic pH.Genetic components encoding for polyamine (PA) biosynthetic pathway are known in several land plant species; however, little is known about them in aquatic plants. We utilized recently sequenced three duckweed (Spirodela polyrhiza) genome assemblies to map PA biosynthetic pathway genes in S. polyrhiza. PA biosynthesis in most higher plants except for Arabidopsis involves two pathways, via arginine decarboxylase (ADC) and ornithine decarboxylase (ODC). ADC-mediated PA biosynthetic pathway genes, namely, one arginase (SpARG1), two arginine decarboxylases (SpADC1, SpADC2), one agmatine iminohydrolase/deiminase (SpAIH), one N-carbamoyl putrescine amidase (SpCPA), three S-adenosylmethionine decarboxylases (SpSAMDc1, 2, 3), one spermidine synthase (SpSPDS1) and one spermine synthase (SpSPMS1) in S. polyrhiza genome were identified here. However, no locus was found for ODC pathway genes in this duckweed. Hidden Markov Model protein domain analysis established that SpADC1 is a prokaryotic/biodegradative type ADC and its molecular phylogenic classification fell in a separate prokaryotic origin ADC clade with SpADC2 as a biosynthetic type of arginine decarboxylase. However, thermospermine synthase (t-SPMS)/Aculis5 genes were not found present. Instead, one of the annotated SPDS may also function as SPMS, since it was found associated with the SPMS phylogenetic clade along with known SPMS genes. Moreover, we demonstrate that S. polyrhiza PA biosynthetic gene transcripts are differentially expressed in response to unfavorable conditions, such as exogenously added salt, methyl jasmonate, or acidic pH environment as well as in extreme temperature regimes. Thus, S. polyrhiza genome encodes for complete polyamine biosynthesis pathway and the genes are transcriptionally active in response to changing environmental conditions suggesting an important role of polyamines in this aquatic plant.

Metagenomic insights into the bacteria responsible for producing biogenic amines in sufu

Harmful levels of biogenic amines (BAs) are frequently identified in sufu. The microorganisms and mechanisms responsible for BA production in sufu, however, are not well documented. In this study, sufu samples were randomly obtained from various regions of China. Putrescine, tyramine, and histamine were quantitated as the most abundant BAs. According to the metagenome sequencing, the abundances and diversities of genes encoding the critical enzymes in BA production were acquired. The results showed that genes encoding arginine-, ornithine-, tryptophan-, and histidine decarboxylases were the predominant amino acid decarboxylase genes. Furthermore, 34 metagenome-assembled genomes (MAGs) were generated, of which 23 encoded at least one gene involved in BA production. Genetic analysis of MAGs indicated genera affiliated with Enterococcus, Lactobacillus-related, and Lactococcus were the major histamine-synthesizing bacteria, and tyrosine may be utilized by Bacillus, Chryseobacterium, Kurthia, Lysinibacillus, Macrococcus, and Streptococcus to product tyramine. The critical species involved in two putrescine-producing pathways were also explored. In the ornithine decarboxylase pathway, Lactobacillus-related and Veillonella were predicted to be the main performers, whereas Sphingobacterium and unclassified Flavobacteriaceae were the dominant executors in the agmatine deiminase pathway. The present study not only explained the BAs formation mechanism in sufu but also identified specific bacteria used to control BAs in fermented soybean products.

Polyamine biosynthetic pathways and their relation with the cold tolerance of maize (Zea mays L.) seedlings

ABSTRACT Background The present study was designed to investigate the inhibition role of two polyamine biosynthesis inhibitors, i.e., D-arginine (D-Arg) and DL-α-difluoromethylornithine (DFMO), in polyamine biosynthesis under chilling stress in different tissues of two maize inbred lines – Huang C (chilling-tolerance) and Mo17 (chilling-sensitive). Results The results showed that exposure to the lower concentration of polyamine biosynthesis inhibitors improved seedlings growth, such as the root length, root and shoot fresh weight, chlorophyll a (chl a). The effectiveness of 10 µM D-Arg treatments was more prominent than those of 10 µM DFMO. However, the higher concentration of inhibitors suppressed seedlings growth, and the exposure to 100 µM DFMO caused stronger decreases in the photosynthetic pigments, such as chlorophyll a (chl a), chlorophyll b (chl b), total chlorophyll and carotenoids, than the other treatments. Meanwhile, the inhibitor treatments caused the lower content of putrescine (Put) in roots, mesocotyls and coleoptiles in both maize inbred lines as compared with untreated plants. However, the lower concentration (10 µM) of polyamine biosynthetic inhibitors improved the Spd content, except 10 µM D-Arg in root of Huang C, and 10 µM DFMO in coleoptiles of both Mo17 and Huang C. The correlation analysis found that Spd was positively significantly correlated with root length and shoot fresh weight of seedling. Conclusion It was showed that the Spd played an important role in seedling growth improvement. At the same concentration of polyamine biosynthetic inhibitors, the Put contents in different tissues of the seedlings treated with DFMO were generally lower than those treated with D-Arg, except for Put contents in root of Mo17 with 10 µM treatment. Moreover, the treatments of 100 µM were more prominent than those of 10 µM treatments. Exposure to 100 µM D-Arg and 100 µM DFMO could each decrease the activities of Arginine decarboxylase (ADC), Ornithine decarboxylase (ODC) and S-adenosylmethionine decarboxylase (SAMDC) in all maize tissues. However, the decrease of the ADC activity was more prominent in 100 µM D-Arg-treated seedlings, while the decrease of SAMDC and ODC activities was prominent in 100 µM DFMO-treated seedlings. Genes involved in polyamine biosynthesis, such as ADC, ODC, SAMDC, and PAO, showed different expression patterns in response to chilling stress and polyamine biosynthesis inhibitors. This study suggested that Put was synthesized via both the ADC and ODC pathways after chilling stress, with the ODC pathway being the major one.

Effects of Amino Acid Decarboxylase Genes and pH on the Amine Formation of Enteric Bacteria From Chinese Traditional Fermented Fish (Suan Yu).

The formation of biogenic amines (BAs) is an important potential risk in Suan yu. This study investigated the amine production abilities of 97 strains of enteric bacteria screened from Suan yu. The genotypic diversity of amino acid decarboxylase and the effect of pH were explored on 27 strains of high-yield BAs. Results showed that high levels of putrescine, histamine, and cadaverine were produced by the 97 strains. In addition, 27 strains carried odc, speA, speB, adiA, and ldc genes. Thirteen carried hdc gene. Morganella morganii 42C2 produced the highest putrescine content of 880 mg/L via the ornithine decarboxylase pathway. The highest histamine content was produced by Klebsiella aerogenes 13C2 (1,869 mg/L). The highest cadaverine content was shown by Klebsiella pneumoniae 47C2 (1,821 mg/L). odc, adiA, speB, ldc, and hdc play important roles in the cellular acid stress response. Acid stress caused the growth delay but can increase the contents of putrescine, histamine, and cadaverine. Decarboxylase was strain specific rather than species specific. This study provides a reference for the genotypic diversity of decarboxylase and effect of pH on the types and abilities of BAs produced by enteric bacteria in Suan yu.

Polyamines are involved in chilling tolerance in tobacco (Nicotiana tabacum) seedlings

To better understand the role of polyamines (PAs) and their metabolism in responses to chilling stress in tobacco plants, this study investigated changes of endogenous PA contents, PA synthetase and oxidase activities, seedling growth and membrane permeability in tobacco (Nicotiana tabacum) cvs. MSk326 (MK, chilling-sensitive variety) and Honghuadajinyuan (HD, chilling-tolerant variety) under chilling stress. Chilling stress significantly reduced the shoot and root dry weights but improved PA contents, especially of putrescine, in the two tobacco cultivars, with higher contents in the tolerant variety. Moreover, stepwise regression analysis indicated that the relative growth rate of tobacco seedlings under chilling stress was positively correlated with endogenous PA contents, putrescine and spermidine, particularly. The above results suggested that the resistance of tobacco seedlings to chilling stress was promoted mainly by putrescine and spermidine. And further study specified that the higher PA contents in the chilling-tolerant tobacco variety were related to higher PA synthetase activities [arginine decarboxylase (ADC) and ornithine decarboxylase (ODC)] and lower oxidase activities (diamine oxidase and polyamine oxidase) under chilling stress. Again, the study demonstrated that PA biosynthesis in tobacco seedlings was mainly dependent on the ODC pathway under chilling stress, through ADC and ODC inhibitor application and recovery treatments. Compared with the ADC inhibitor (d-arginine), the ODC inhibitor (2-fluoromethyl ornithine) more significantly inhibited PA contents and increased membrane permeability in tobacco seedlings under chilling stress. In summary, the research findings suggested PAs play an important role in regulating the chilling tolerance of tobacco seedlings.

Lactobacillus rossiae strain isolated from sourdough produces putrescine from arginine

This work reports a Lactobacillus rossiae strain (L. rossiae D87) isolated from sourdough that synthesizes putrescine - a biogenic amine that raises food safety and spoilage concerns - from arginine via the ornithine decarboxylase (ODC) pathway. The odc and potE genes were identified and sequenced. These genes respectively encode ornithine decarboxylase (Odc), which participates in the decarboxylation of ornithine to putrescine, and the ornithine/putrescine exchanger (PotE), which exchanges ornithine for putrescine. Transcriptional analysis showed that odc and potE form an operon that is regulated transcriptionally by ornithine in a dose-dependent manner. To explore the possible role of the ODC pathway as an acid stress resistance mechanism for this bacterium, the effect of acidic pHs on its transcriptional regulation and on putrescine biosynthesis was analysed. Acidic pHs induced the transcription of the odc-potE genes and the production of putrescine over that seen at neutral pH. Further, putrescine production via the ODC system improved the survival of L. rossiae D87 by counteracting the acidification of the cytoplasm when the cells were subjected to acidic conditions. These results suggest the ODC pathway of L. rossiae D87 provides a biochemical defence mechanism against acidic environments.

Extracellular polyamines-induced proliferation and migration of cancer cells by ODC, SSAT, and Akt1-mediated pathway.

High levels of polyamines were observed and were related to a poor prognosis in cancer patients. However, the mechanism is not obvious. The aim of this study is to mimic the extracellular polyamines in a tumor microenviroment and to explore the role of extracellular polyamines in the proliferation and migration of cancer cells. Three different concentrations of polyamines composed of putrescine, spermidine, and spermine were used. Colony formation assay, wound healing assay, and transwell migration assay were performed. Akt1-overexpression cells were constructed. The related protein expression was examined using a western blot. In this study, polyamines promoted colony formation and cell migration in a concentration-dependent and time-dependent manner. Polyamines upregulated the expression of ornithine decarboxylase (ODC), SSAT, Akt1, Akt, hypoxia-inducible factors-1α, vascular endothelial growth factor, and matrix metalloproteinases, and downregulated p27 expression. The effects of combination of polyamines and Akt1 overexpression on colony formation and migration were more obvious than the effects of Akt1 overexpression alone. In Akt1-overexpression cells, polyamines also upregulated the expression of ODC, SSAT, hypoxia-inducible factors-1α, vascular endothelial growth factor, and matrix metalloproteinases and downregulated p27 expression. In conclusion, extracellular polyamines induced proliferation and cancer cell migration by inducing ODC and SSAT expression, and the Akt1-mediated pathway.

Gene polymorphisms in the ornithine decarboxylase-polyamine pathway modify gastric cancer risk by interaction with isoflavone concentrations.

The study aimed to examine the association between genes encoding molecules in the ornithine decarboxylase (ODC)-polyamine pathway (ODC1, AMD1, NQO1, NOS2A, and OAZ2) and gastric cancer risk and whether the gene-phytoestrogen interaction modifies gastric cancer risk. Among 76 gastric cancer cases and their 1:4 matched controls within the Korean Multi-center Cancer Cohort, a total of 30 SNPs in five genes involved in the ODC pathway were primarily analyzed. The second-stage genotyping in 388 matched case-control sets was conducted to reevaluate the significant SNPs interacting with phytoestrogens during the primary analysis. The summary odds ratios (ORs) [95% confidence intervals (CIs)] for gastric cancer were estimated. Interaction effects between the SNPs and plasma concentrations of phytoestrogens (genistein, daidzein, equol, and enterolactone) were evaluated. In the pooled analysis, NQO1 rs1800566 showed significant genetic effects on gastric cancer without heterogeneity [OR0.83 (95% CI0.70-0.995)] and a greater decreased risk at high genistein/daidzein levels [OR0.36 (95% CI0.15-0.90) and OR0.26 (95% CI0.10-0.64), respectively; p interaction<0.05]. Risk alleles of AMD1 rs1279599, AMD1 rs7768897, and OAZ2 rs7403751 had a significant gene-phytoestrogen (genistein and daidzein) interaction effect to modify the development of gastric cancer. They had an increased gastric cancer risk at low isoflavone levels, but a decreased risk at high isoflavone levels (p interaction<0.01). Our findings suggest that common variants in the genes involved in the ODC pathway may contribute to the risk of gastric cancer possibly by modulating ODC polyamine biosynthesis or by interaction between isoflavones and NQO1, OAZ2, and AMD1.

Whole-Genome Pathway Analysis on 132,497 Individuals Identifies Novel Gene-Sets Associated with Body Mass Index

Whole genome pathway analysis is a powerful tool for the exploration of the combined effects of gene-sets within biological pathways. This study applied Interval Based Enrichment Analysis (INRICH) to perform whole-genome pathway analysis of body-mass index (BMI). We used a discovery set composed of summary statistics from a meta-analysis of 123,865 subjects performed by the GIANT Consortium, and an independent sample of 8,632 subjects to assess replication of significant pathways. We examined SNPs within nominally significant pathways using linear mixed models to estimate their contribution to overall BMI heritability. Six pathways replicated as having significant enrichment for association after correcting for multiple testing, including the previously unknown relationships between BMI and the Reactome regulation of ornithine decarboxylase pathway, the KEGG lysosome pathway, and the Reactome stabilization of P53 pathway. Two non-overlapping sets of genes emerged from the six significant pathways. The clustering of shared genes based on previously identified protein-protein interactions listed in PubMed and OMIM supported the relatively independent biological effects of these two gene-sets. We estimate that the SNPs located in examined pathways explain ∼20% of the heritability for BMI that is tagged by common SNPs (3.35% of the 16.93% total).

Putrescine production via the ornithine decarboxylation pathway improves the acid stress survival of Lactobacillus brevis and is part of a horizontally transferred acid resistance locus

Decarboxylation pathways are widespread among lactic acid bacteria; their physiological role is related to acid resistance through the regulation of the intracellular pH and to the production of metabolic energy via the generation of a proton motive force and its conversion into ATP. These pathways include, among others, biogenic amine (BA) production pathways. BA accumulation in foodstuffs is a health risk; thus, the study of the factors involved in their production is of major concern. The analysis of several lactic acid bacterial strains isolated from different environments, including fermented foods and beverages, revealed that the genes encoding these pathways are clustered on the chromosome, which suggests that these genes are part of a genetic hotspot related to acid stress resistance. Further attention was devoted to the ornithine decarboxylase pathway, which affords putrescine from ornithine. Studies were performed on three lactic acid bacteria belonging to different species. The ODC pathway was always shown to be involved in cytosolic pH alkalinisation and acid shock survival, which were observed to occur with a concomitant increase in putrescine production.

Diabetes/high glucose induced arginase increases arterial smooth muscle cell proliferation and collagen synthesis/fibrosis through ornithine decarboxylase and ornithine aminotransferase pathways

Arginase can compete with nitric oxide (NO) synthase for their common substrate, L‐arginine, and its increased activity can decrease NO production. Arginase also provides ornithine for synthesis of polyamines via ornithine decarboxylase (ODC) and proline/collagen via ornithine aminotransferase (OAT). We hypothesize that diabetes/high glucose (HG) induce vascular wall thickening and fibrosis through increased level of arginase‐derived ornithine available for ODC and OAT. We exposed rat aortic smooth muscle cells to HG (25 mM), which resulted in increased arginase activity (57%), cell proliferation (22%), and ODC mRNA (25%). Treatment with arginase inhibitor BEC (S‐(2‐boronoethyl)‐l‐cysteine) prevented these changes. We observed increased perivascular fibrosis (68%) in coronary arteries of 8‐week wild‐type (WT) diabetic mice compared to arginase I partial knockout (AI+/−) diabetic mice. Aorta from diabetic WT mice also exhibited impaired endothelium‐dependent relaxation (EDR) compared to control (maximum EDR: 32.9% vs 61.8%). EDR was largely maintained in aorta from diabetic AI+/− mice. Furthermore, adenoviral overexpression of arginase I in isolated aorta concomitantly increased ODC expression, implying an effect of arginase on ODC. In conclusion, arginase appears to have important roles in HG/diabetes‐induced vascular dysfunction, proliferation and fibrosis through ODC and OAT pathway.

Hydroxydibenzoylmethane induces apoptosis through repressing ornithine decarboxylase in human promyelocytic leukemia HL-60 cells

Ornithine decarboxylase (ODC) is the rate-limiting enzyme in polyamine biosynthesis and a target for chemoprevention. Hydroxydibenzoylmethane (HDB), a derivative of dibenzoylmethane of licorice, is a promising chemopreventive agent. In this paper, we investigated whether HDB would inhibit the ODC pathway to enhance apoptosis in human promyelocytic leukemia HL-60 cells. We found ODC enzyme activity was reduced during HDB treatment. Overexpression of ODC in HL-60 parental cells could reduce HDB-induced apoptosis, which leads to loss of mitochondrial membrane potential (Δψ(m)), through lessening intracellular ROS. Furthermore, ODC overexpression protected cytochrome c release and the activation of caspase-3 following HDB treatment. The results demonstrated HDB-induced apoptosis was through a mechanism of down-regulation of ODC and occurred along a ROS-dependent mitochondria-mediated pathway.

Arginine metabolism in the sheep abomasal nematode parasites Haemonchus contortus and Teladorsagia circumcincta

The ornithine urea cycle, polyamine synthesis, nitric oxide synthesis and metabolism of arginine to putrescine have been investigated in L3 and adult Haemonchus contortus and Teladorsagia circumcincta. Neither parasite had a detectable arginine deiminase/dihydrolase pathway nor a functional ornithine urea cycle. Nitric oxide synthase was present in central and peripheral nerves, but was not detected in whole parasite homogenates. Both arginase (E.C. 3.5.3.1) and agmatinase (E.C. 3.5.3.11) activities were present in both species. Arginase did not require added Mn2+ and had an optimal pH of 8.5. Polyamine metabolism differed in the two species and from that in mammals. Ornithine decarboxylase (E.C. 4.1.1.17) was present in both parasites, but no arginine decarboxylase (E.C. 4.1.1.19) activity was detected in T. circumcincta. The flexibility of synthesis of putrescine in H. contortus may make this pathway less useful as a target for parasite control than in T. circumcincta, in which only the ornithine decarboxylase pathway was detected.

Putrescine production by engineered Corynebacterium glutamicum

Here, we report the engineering of the industrially relevant Corynebacterium glutamicum for putrescine production. C. glutamicum grew well in the presence of up to 500 mM of putrescine. A reduction of the growth rate by 34% and of biomass formation by 39% was observed at 750 mM of putrescine. C. glutamicum was enabled to produce putrescine by heterologous expression of genes encoding enzymes of the arginine- and ornithine decarboxylase pathways from Escherichia coli. The results showed that the putrescine yield by recombinant C. glutamicum strains provided with the arginine-decarboxylase pathway was 40 times lower than the yield by strains provided with the ornithine decarboxylase pathway. The highest production efficiency was reached by overexpression of speC, encoding the ornithine decarboxylase from E. coli, in combination with chromosomal deletion of genes encoding the arginine repressor ArgR and the ornithine carbamoyltransferase ArgF. In shake-flask batch cultures this strain produced putrescine up to 6 g/L with a space time yield of 0.1 g/L/h. The overall product yield was about 24 mol% (0.12 g/g of glucose).

Putrescine accumulation confers drought tolerance in transgenic Arabidopsis plants over-expressing the homologous Arginine decarboxylase 2 gene

In Arabidopsis, a model genus missing a functional ornithine decarboxylase pathway, most of the key genes involved in polyamine biosynthesis are duplicated. This gene redundancy has been related to the involvement of certain gene isoforms in the response to specific environmental stimuli. We have previously shown that drought stress induces Arginine decarboxlase 2 expression, while transcript levels for Arginine decarboxlase 1 remain constant. Accumulation of putrescine and increased arginine decarboxlase activity (EC 4.1.1.19) levels in response to different abiotic stresses have been reported in many different plant systems, but the biological meaning of this increase remains unclear. To get a new insight into these questions, we have studied the response to drought of transgenic Arabidopsis thaliana lines constitutively expressing the homologous Arginine decarboxlase 2 gene. These lines contain high levels of putrescine with no changes in spermidine and spermine content even under drought stress. Drought tolerance experiments indicate that the different degree of resistance to dehydration correlates with Put content. Although no significant differences were observed in the number of stomata between wild-type and transgenic plants, a reduction in transpiration rate and stomata conductance was observed in the ADC2 over-expressor lines. These results indicate that one of the mechanisms involved in the drought tolerance of transgenic plants over-producing Put is related to a reduction of water loss by transpiration.

Expression of arginine decarboxylase and ornithine decarboxylase genes in apple cells and stressed shoots

Arginine decarboxylase (ADC) and ornithine decarboxylase (ODC) are two important enzymes responsible for putrescine biosynthesis. In this study, a full-length ADC cDNA (MdADC) was isolated from apple [Malus sylvestris (L.) Mill. var. domestica (Borkh.) Mansf.]. Meanwhile, a partial ODC (pMdODC) could be amplified only by a second RCR from the RT-PCR products, whereas a full-length ODC could not be obtained by either cDNA library screening or 5'- and 3'-RACEs, suggesting quite low expression. Moreover, D-arginine, an ADC inhibitor, caused a decrease in ADC activity and severely inhibited the growth of apple callus, which could be partially resumed by exogenous addition of putrescine, whereas alpha-difluoromethylornithine (DFMO), an inhibitor for ODC, caused the incomplete repression of callus growth without changing ODC activity. RNA gel blot showed that the expression level of MdADC was high in young tissues/organs with rapid cell division and was positively induced by chilling, salt, and dehydration, implying its involvement in both cell growth and these stress responses. By contrast, the transcript of ODC could not be detected by RNA gel blot analysis. Based on the present study, it is possible to conclude that (i) the ODC pathway is active in apple, although the expression level of the pMdODC gene homologous with its counterparts found in other plant species is quite low; and (ii) MdADC expression correlates with cell growth and stress responses to chilling, salt, and dehydration, suggesting that ADC is a primary biosynthetic pathway for putrescine biosynthesis in apple.

Ornithine decarboxylase transgene in tobacco affects polyamines, in vitro-morphogenesis and response to salt stress

Summary Transgenic tobacco plants expressing the putrescine synthesis gene ornithine decarboxylase from mouse were raised to study the effects of up-regulation of a metabolic pathway as critical as the polyamine biosynthesis on the plant growth and development, in vitro -morphogenesis and their response to salt stress. Further, the response of the alternate pathway (arginine decarboxylase) for putrescine synthesis to the modulation of the ornithine decarboxylase pathway has also been investigated. The over-expression of the odc gene and increased levels of putrescine in tobacco led to a delay in plant regeneration on selection medium which could be overcome by the exogenous application of polyamine biosynthesis inhibitors and spermidine. Further, the lines generated had a variable in vitro morphogenic potential, which could be correlated to the shifts in their polyamine metabolism. These studies have brought forward the critical role played by polyamines in the normal development of plants and also their role in plant regeneration. Since polyamines are known to accumulate in cells under abiotic stress conditions, the tolerance of the transgenics to salt stress was also investigated and the transgenics with their polyamine metabolism up-graded showed increased tolerance to salt stress.

Stimulation of putrescine biosynthesis via the ornithine decarboxylase pathway by gibberellic acid in the in vitro rooting of globe artichoke ( Cynara scolymus )

Rooting and growth of globe artichoke (Cynarascolymus) cuttings were improved by supplementing the rooting mediumwith 10 μM gibberellic acid(GA3). Exogenous putrescine (Put) (0.5mM) raised the percentage of rooting to thesame level as GA3, but decreased the number of roots per plant.Addition of Put to GA3 produced the same effects on growthparametersand rooting percentage as Put alone. Changes in endogenous contents of freepolyamines (PAs) were determined in plants cultured 4 weeks in a rootingmedium.Whatever the treatments, rooting caused a large decrease in Put, the largelypredominant PA, in the leaves. GA3 increased the free Put content ofthe leaves and roots. DL-α-difluoromethylarginine (DFMA), which inhibitsarginine decarboxylase (ADC), did not affect rooting while Put content wasdecreased by 12–15%. DFMA changed neither GA3 effectsnor those from DFMO when used in combination. An inhibitor of ornithinedecarboxylase (ODC), DL-α-difluoromethylornithine (DFMO), inhibited leafgrowth as well as rooting and decreased the Put content; these effects werepartially reversed by addition of Put. GA3 combined with DFMO didnotalleviate the inhibitory effects of DFMO. The GA3 treatment caused aclear stimulation of ODC activity in the leaves; it also stimulated14C-putrescine uptake (2.8-fold) by leaves and its translocationtowards roots. These results indicate that the ODC pathway mediates thehormone-induced rooting response in globe artichoke. Moreover, a release of Putfrom its conjugates could contribute to the increase of free-Put levels in theroots under GA3 treatment. Thus, Put can be considered to be a goodmarker of rhizogenesis.

De novo synthesis of arginine and ornithine from citrulline in human colon carcinoma cells: metabolic fate of l-ornithine

In human colon carcinoma cells (HT-29 cells), l-arginine is the common precursor of l-ornithine which generates polyamines strictly necessary for cellular growth, and nitric oxide which has a strong antiproliferative activity. We show here that proliferative HT-29 cells possess the capacity for de novo synthesis of l-arginine from l-citrulline, but not from l-ornithine. l-Ornithine is apparently not an l-arginine precursor due to the absence of any detectable ornithine carbamoyltransferase activity. In contrast, the newly synthesized l-arginine was competent for urea and thus l-ornithine production in a context of a high putrescine production in the ornithine decarboxylase pathway and a low degradation of this polyamine in the diamine oxidase pathway. However, cells grown in an arginine-free culture medium containing added l-citrulline were unable to reach confluency. Furthermore, the low amount of nitric oxide produced from l-arginine by these cells was apparently not involved in the control of cell growth since inhibition of nitric oxide synthase activity was without effect. On the other hand, the capacity of more differentiated and less proliferative HT-29 cells for de novo l-arginine synthesis from l-citrulline was increased. It is concluded that l-citrulline is a precursor of l-arginine and l-ornithine in proliferative HT-29 cells and that the metabolic fate of l-ornithine in these cells is mainly devoted to polyamine synthesis. The similarity between differentiated HT-29 cells and the enterocytes of newborn animals in terms of l-arginine metabolism is finally discussed.