Endogenous Polyamine Levels Research Articles

Effect of polyamines and polyamine precursors on hyperhydricity in micropropagated apple shoots

Hyperhydricity is a physiological disorder affecting tissue-culture-generated plants. It is associated with excessive hydration and poor lignification. The effects of exogenous polyamines (spermidine and putrescine) and polyamine precursors (arginine and ornithine) were studied on ‘Jonagold’ apple shoots subjected to hyperhydric conditions for one in vitro multiplication cycle (28 days) on a culture medium containing gelrite as gelling agent. Supplementation of the gelrite-containing medium with 10−5 M spermidine, ornithine, or arginine reduced the percentage of hyperhydricity by at least 50 %. Exogenous supplementation with spermidine or a polyamine precursor also caused total phenolics and the antioxidant capacity (measured by ORAC and DPPH assays) to decrease in shoots during the first 2 weeks. These results suggest an important role of spermidine and its precursor ornithine in countering hyperhydricity. Supplementation could help to maintain high endogenous levels of polyamines and phenolamides necessary for cell osmoregulation, antioxidant protection, cell wall cross-linking, and plant growth regulation.

Drought-inhibited ribulose-1,5-bisphosphate carboxylase activity is mediated through increased release of ethylene and changes in the ratio of polyamines in pakchoi

To study the mechanisms of drought inhibiting photosynthesis and the role of PAs and ethylene, the photosynthetic rate (Pn), the maximal photochemical efficiency of PSII (Fv/Fm), the intercellular CO2 concentration (Ci), photorespiratory rate (Pr), the amount of chlorophyll (chl), antioxidant enzyme activity, ethylene levels, RuBPC (ribulose-1,5-bisphosphate carboxylase) activity and endogenous polyamine levels of pakchoi were examined, and an inhibitor of S-adenosylmethionine decarboxylase (SAMDC) and an inhibitor of ethylene synthesis and spermidine (Spd) were used to induce the change of endogenous polyamine levels. The results show that drought induced a decrease in Pn and RuBPC activity, an increase in the intercellular CO2 concentration (Ci), but no change in the actual photochemical efficiency of PSII (ΦPSII), and chlorophyll content. In addition, drought caused an increase in the free putrescine (fPut), the ethylene levels, a decrease in the Spd and spermine (Spm) levels, and the PAs/fPut ratio in the leaves. The exogenous application of Spd and amino oxiacetic acid (AOAA, an inhibitor of ethylene synthesis) markedly reversed these drought-induced effects on polyamine, ethylene, Pn, the PAs/fPut ratio and RuBPCase activity in leaves. Methylglyoxal-bis(guanylhydrazone) (MGBG), an inhibitor of SAMDC resulting in the inability of activated cells to synthesize Spd and Spm, exacerbates the negative effects induced by drought. These results suggest that the decrease in Pn is at least partially attributed to the decrease of RuBPC activity under drought stress and that drought inhibits RuBPC activity by decreasing the ratio of PAs/fPut and increasing the release of ethylene.

Role of polyamines in regulating silymarin production in Silybum marianum (L.) Gaertn (Asteraceae) cell cultures under conditions of calcium deficiency

As part of our efforts to identify the possible role of polyamines (PAs) in silymarin (Sm) production, the effects of calcium deprivation on cell growth and on endogenous PAs levels and Sm production by milk thistle (Silybum marianum (L.) Gaertn) grown in cell cultures were examined. Young cultured cells of the H2 line of S. marianum were transferred to a medium without calcium and with ethylene glycol-bis-(β-aminoethyl) ether-N,N,N′,N′-tetraacetic acid present to chelate any free calcium in order to analyze the effects of this medium on the levels of PAs and Sm produced by the cells. During the 17 days of exposure to this calcium-free medium most of the cell populations were in the G0/G1 phase (from day 7 to day 14 of culture) while PA levels underwent a progressive decline up to day 17, after which they were no longer detectable. We observed that putrescine (Put) accumulation was always lower than that observed under normal conditions. The lack of calcium in the MS medium advances the onset of the stationary phase, whose beginning is marked by an increase in the Put/spermidine (Spd) index, raising the production of Sm; the suspensions were productive for a longer time and hence produced more of the substance. Our results indicate that under stress conditions the production of Sm in young-cell suspensions of S. marianum is not associated with high levels of PAs in the medium – contrary to what one would expect – allowing us to conclude that growth inhibition appears to be the factor responsible for the maximum Sm accumulation while PAs are not directly involved in the Sm synthesis pathway by milk thistle grown in culture.

Morphological and polyamine content changes in embryogenic and non-embryogenic callus of sugarcane

Differences in competence acquisition and subsequent embryo maturation in embryogenic and non-embryogenic callus of sugarcane var. SP79-1011 were evaluated using histomorphological analysis, growth curves, numbers of somatic embryos, and polyamine contents. Embryogenic callus was formed by cells with embryogenic characteristics such as a rounded shape, prominent nuclei, a high nucleus: cytoplasm ratio, small vacuoles and organized globular structures. However, non-embryogenic callus presented dispersed, elongated and vacuolated cells with a low nucleus: cytoplasm ratio; these characteristics did not allow for the development of somatic embryos even upon exposure to a maturation stimulus. These results suggest that non-embryogenic callus does not acquire embryogenic competence during induction and that maturation treatment is not sufficient to promote somatic embryo differentiation. The use of activated charcoal (AC; 1.5 g L−1) resulted in a higher somatic embryo maturation rate in embryogenic callus but did not yield success in non-embryogenic callus. Embryogenic callus incubated with control (10 μM 2,4-dichlorophenoxyacetic acid) and maturation (1.5 g L−1 AC) treatments for 28 days showed similar patterns of total free polyamines; these results differed from the results observed with non-embryogenic callus, suggesting that embryogenic callus already exhibits a characteristic pattern of endogenous polyamine levels. At 28 days of culture with maturation treatment, embryogenic callus exhibited significantly higher levels of free Spm than embryogenic callus incubated with control treatment and non-embryogenic callus incubated with both treatments. This result suggests that Spm could be important for the acquisition of embryogenic competence and somatic embryo maturation in sugarcane var. SP79-1011.

Mutational loss of Arabidopsis SLOW WALKER2 results in reduced endogenous spermine concomitant with increased aluminum sensitivity.

A previously-identified Arabidopsis mutant with hypersensitivity to aluminum, als7-1 was studied further to determine the nature of the mutation and subsequently establish the biochemical basis of the increase in Al sensitivity. Physiological analysis revealed that the Al hypersensitivity phenotype is correlated with increased Al uptake and Al-dependent gene expression, indicating that als7-1 has a defect in an Al-exclusion mechanism. Cloning of the als7-1 mutation showed that it negatively affects the gene encoding the putative nucleolar localised ribosomal biogenesis factor SLOW WALKER2, which is required for normal gametogenesis and mitotic progression. Molecular analysis indicated that Al hypersensitivity in als7-1 is correlated with loss of expression of a factor required for S-adenosylmethionine recycling and reduced levels of endogenous polyamines in the mutant. Further analysis shows that Al-dependent root growth inhibition is reversed by addition of exogenous spermine, which is correlated with a significant reduction in Al uptake by spermine treated roots. Endogenous spermine likely functions to compete with Al3+ for binding to extra- and intracellular anionic sites, which suggests that increased spermine levels may be an effective means to improve root growth in Al toxic acid soil environments.

Polyamines affect the cellular growth and structure of pro‐embryogenic masses in Araucaria angustifolia embryogenic cultures through the modulation of proton pump activities and endogenous levels of polyamines

Polyamines (PAs) are abundant polycationic compounds involved in many physiological processes in plants, including somatic embryogenesis. This study investigates the role of PAs on cellular growth and structure of pro-embryogenic masses (PEMs), endogenous PA and proton pump activities in embryogenic suspension cultures of Araucaria angustifolia. The embryogenic suspension cultures were incubated with putrescine (Put), spermidine (Spd), spermine (Spm) and the inhibitor methylglyoxal-bis(guanylhydrazone) (MGBG), respectively (1 mM). After 24 h and 21 days, the cellular growth and structure of PEMs, endogenous PA contents and proton pump activities were analyzed. The addition of Spm reduced the cellular growth and promoted the development of PEMs in embryogenic cultures, which could be associated with a reduction in the activities of proton pumps, such as H(+) -ATPase P- and V-types and H(+) -PPases, and alterations in the endogenous PA contents. Spm significantly affected the physiology of the A. angustifolia somatic embryogenesis suspension, as it potentially affects cellular growth and structure of PEMs through the modulation of proton pump activities. This work demonstrates the involvement of exogenous PAs in the modulation of cellular growth and structure of PEMs, endogenous PA levels and proton pump activities during somatic embryogenesis. To our knowledge, this study is the first to report a relationship between PAs and proton pump activities in these processes. The results obtained in this study offer new perspectives for studies addressing the role of PAs and proton pump on somatic embryogenesis in this species.

Modulation of polyamine levels in ginseng hairy root cultures subjected to salt stress

Modulation of differential gene expression and change of polyamine content by salt stress are analyzed for the first time in a well-known medicinal plant, Panax ginseng C.A. Meyer. Three ginseng genes (PgSPD, PgSAMDC, and PgADC) involved in polyamine biosynthesis showed differential up-regulation patterns after 1 and 7 days of salt treatments. The modulation of gene expression resulted in the elevation of total polyamine content with relatively high levels of spermidine and spermine, while putrescine level diminished depending on the salt concentration. Conversely, salt stress led to a significant increase in diamine oxidase and subsequent decline in polyamine oxidase. The proline content caused by salinity follows a similar pattern as the total polyamine content and exogenous spermidine also resulted in the alleviation of proline content under salinity. Further, polyamine biosynthesis inhibitors, such as cyclohexylamine and methylglyoxal bis-(guanylhydrazone) mediated down-regulation of PgSPD and PgSAMDC, and affected cellular polyamine levels. Thus, polyamines may enhance the ginseng plant tolerance in response to the salt stress by increasing the levels of endogenous polyamines.

THE INTERACTION BETWEEN ETHYLENE AND POLYAMINES DURING RIPENING OF OLIVE FRUIT

Ripening is a complex and highly regulated phenomenon involving many endogenous and environmental factors, all of which must be correctly balanced in order to achieve the best fruit quality. Ripening control in olive is an important issue because, owing to early harvest and long storage times, fruit quality (oil/canning) can deteriorate before reaching the consumer. Ethylene and polyamines are known to have opposite effects in relation to fruit ripening and senescence. In fact, polyamine and ethylene physiologies may be linked during fruit development. This work describes the changes in endogenous free polyamines levels and ethylene production during fruit development from two cultivars of Olea europaea L. (‘Arbequina’ and ‘Picual’). Ethylene production (the 1-aminocyclopropane-1-carboxylic acid (ACC) level) has been determined by gas chromatography and polyamine content (putrescine, spermidine and spermine) by dansylation and separation of dansyl derivatives by HPLC. To investigate the relationships between ethylene and free polyamines in olive fruit, the effect of exogenous polyamines (putrescine and spermidine) and aminooxy-acetic acid (AOA, inhibitor of ethylene biosynthesis) on polyamines and ACC levels was also examined in ripening olive fruit. Fruit treatments were performed in planta.

Effects of exogenous spermidine on photosynthesis, xanthophyll cycle and endogenous polyamines in cucumber seedlings exposed to salinity

The effects of exogenous spermidine (Spd, 1 mmol·L-1) on photosynthetic characteristics, xanthophylls cycle components and endogenous polyamines levels were investigated in cucumber seedlings subjected to salt stress (75 mmol·L-1 NaCl). Chlorophyll contents and net photosynthetic rate (PN) of cucumber seedlings showed a significant decrease under salinity but an increase with exogenous Spd application. Salt stress caused a remarkable decline in the maximum quantum efficiency (Fv/Fm) and the actual efficiency of photosystem II (ФPSⅡ), where an increase was observed in the constitutive loss processes (ΦNO). Application of exogenous Spd significantly decreased ФNO and enhanced regulated non-photochemical energy loss (ФNPQ) in the salt-stressed plants. Spd treatment caused an increase in the size of xanthophyll cycle pool (VAZ) and further enhanced de-epoxidation of the xanthophyll cycle (DEPS) under salt stress. These results suggest that exogenous Spd alleviated salt-mediated decline in photosynthetic efficiency through the enhanced involvement of the energy dissipation that is dependent on the xanthophyll cycle. In addition, foliar spray Spd significantly increased the free, bound and conjugated polyamines in the leaves of the salt stressed plants. Spd also increased the free putrescine (Put)/(Spd+Spm) ratio and decreased bound and conjugated Put/(Spd+Spm) under salinity. Thus, we conclude that Spd can alleviate salt-induced damage on cucumber seedlings by regulating the levels of endogenous polyamines, which was associated with an improvement in the photochemical efficiency of PSII of the salt stressed plants. Key words: Cucumber, endogenous polyamines, photosynthetic characteristics, salt stress, spermidine.

Selenium and spermine alleviate cadmium induced toxicity in the red seaweed Gracilaria dura by regulating antioxidants and DNA methylation

The protective role of exogenously supplied selenium (Se) and polyamines (PAs) such as putrescine (Put) and spermine (Spm) in detoxifying the cadmium (Cd) induced toxicity was studied in the marine red alga Gracilaria dura in laboratory conditions. The Cd exposure (0.4 mM) impede the growth of alga while triggering the reactive oxygen species (ROS viz. O(2)(•-) and H(2)O(2)) generation, inhibition of antioxidant system, and enhancing the lipoxygenase (LOX) activity, malondialdehyde (MDA) level and demethylation of DNA. Additions of Se (50 μM) and/or Spm (1 mM) to the culture medium in contrast to Put, efficiently ameliorated the Cd toxicity by decreasing the accumulation of ROS and MDA contents, while restoring or enhancing the level of enzymatic and nonenzymatic antioxidants and their redox ratio, phycobiliproteins and phytochelatins, over the controls. The isoforms of antioxidant enzymes namely superoxide dismutase (Mn-SOD, ~150 kDa; Fe-SOD ~120 kDa), glutathione peroxidase (GSH-Px, ~120 and 140 kDa), glutathione reductase (GR, ~110 kDa) regulated differentially to Se and/or Spm supplementation. Furthermore, it has also resulted in enhanced levels of endogenous PAs (specially free and bound insoluble Put and Spm) and n-6 PUFAs (C20-3, n-6 and C20-4, n-6). This is for the first time wherein Se and Spm were found to regulate the stabilization of DNA methylation by reducing the events of cytosine demethylation in a mechanism to alleviate the Cd stress in marine alga. The present findings reveal that both Se and Spm play a crucial role in controlling the Cd induced oxidative stress in G. dura.

Initial testing (stage 1) of the polyamine analog PG11047 by the pediatric preclinical testing program

PG11047 is a novel conformationally restricted analog of the natural polyamine, spermine that lowers cellular endogenous polyamine levels and competitively inhibits natural polyamine functions leading to cancer cell growth inhibition. The activity of PG11047 was evaluated against the PPTP's in vitro and in vivo panels. PG11047 was evaluated against the PPTP in vitro panel using 96 hr exposure at concentrations ranging from 10 nM to 100 µM. It was tested against the PPTP in vivo panels at a dose of 100 mg/kg administered by the intraperitoneal route weekly for 6 weeks. In vitro PG11047 demonstrated a concentration-response pattern consistent with cytostatic activity. The median EC(50) for PG11047 was 71 nM. Cell lines of the Ewing sarcoma panel had a lower median EC(50) value compared to the remaining cell lines in the panel, while cell lines of the neuroblastoma panel had a higher median EC(50) value. In vivo PG11047 induced significant differences in EFS distribution compared to control in 5 of 32 (15.6%) of the evaluable solid tumor xenografts and in 0 of 7 (0%) of the evaluable ALL xenografts. The single case of tumor regression occurred in an ependymoma xenograft. Further pediatric development of PG11047 will require better defining a target population and identifying combinations for which there is a tumor-selective cytotoxic effect. The regression observed for an ependymoma xenograft and the exquisite sensitivity of some Ewing sarcoma cell lines to the antiproliferative effects of PG11047 provide leads for further preclinical investigations.

Cloning, biochemical identification, and expression analysis of a gene encoding S-adenosylmethionine decarboxylase in navel orange (Citrus sinensis Osbeck)

SummaryPolyamines have been shown to be involved in a variety of plant physiological processes such as growth, development, and stress responses. As S-adenosylmethionine decarboxylase (SAMDC) is the rate-limiting enzyme for polyamine biosynthesis, efforts were made to isolate the gene encoding SAMDC from ‘Newhall’ navel orange (Citrus sinensis Osbeck). RT-PCR, in combination with a bioinformatics approach, produced a cDNA (CsSAMDC) containing an intact open reading frame (ORF) of 1,095 bp. The deduced amino acid sequence shared high homology (>75%) with four selected plant SAMDCs and possessed several highly-conserved amino acid residues or motifs. Expression of CsSAMDC in Escherichia coli gave rise to a protein of the expected size, which was shown to have biochemical activity via a SAMDC assay. The expression pattern of the CsSAMDC gene in ‘Newhall’ callus tissue broadly matched the growth of callus cells and the fluctuation in endogenous polyamine levels. Levels of transcription of CsSAMDC in ‘Newhall’ shoots were enhanced by treatment with 10% (w/v) polyethylene glycol. In addition, the steady state level of CsSAMDC mRNA varied in genotypes with varying levels of tolerance to salinity or to low temperature stress, as revealed by the earlier induction of CsSAMDC mRNA at the initial stages of either stress treatment in tolerant genotypes. These results suggest that CsSAMDC is a stress-responsive gene and that temporal changes in its expression profile may be part of the mechanism responsible for genotypic variations in stress tolerance.

Tuber formation and growth of Dioscorea cayenensis-D. rotundata complex: interactions between exogenous and endogenous jasmonic acid and polyamines

Tubers can be initiated and developed in vitro from nodal cuttings of yam (Dioscorea cayenensis-D. rotundata complex). The effect of exogenous jasmonic acid, alone or in combination with putrescine, on these processes was investigated in relationship to endogenous jasmonic acid and polyamine levels. Application of exogenous jasmonic acid at various concentrations positively affected microtuber formation and growth from yam nodal cuttings. In control conditions, 3 weeks were needed to obtain 100% of tuberisation. Jasmonic acid at low level (0.1 μM) accelerated tuber formation (46% after 1 week) as did putrescine (10 μM). But endogenous levels of jasmonic acid were not significantly affected by its exogenous presence in the medium. Jasmonic acid also interacted with other growth regulators as polyamines, but the decrease in time necessary to observe tuber formation could not be correlated with endogenous modifications of PUT content. The presence of jasmonic acid (0.1–1 μM) as PUT (1 μM) induced also an increase of tuber length and weight. The combination of jasmonic acid (0.1 μM) and putrescine (1 μM) had no positive effect on tuber formation (precocity) but had an additive effect on further growth (length and weight). In the future, these results could help the optimising in vitro conditions for mass production of larger yam microtubers.

Cinnamic acid-inhibited ribulose-1,5-bisphosphate carboxylase activity is mediated through decreased spermine and changes in the ratio of polyamines in cowpea

This study investigated the effects of cinnamic acid (CA) on ribulose-1,5-bisphosphate carboxylase (RuBPC) activity and the endogenous polyamine levels of cowpea leaves. The results show that 0.1 mM CA treatment decreased photosynthetic rate ( P n) and RuBPC activity, but it did not affect the maximal photochemical efficiency of PSII ( F v/ F m), the actual photochemical efficiency of PSII ( ΦPSII), intercellular CO 2 concentration ( C i), and relative chlorophyll content. These suggest that the decrease in P n is at least partially attributed to a lowered RuBPC activity. In addition, 0.1 mM CA treatment increased the putrescine (Put) level, but decreased spermidine (Spd) and spermine (Spm) levels, thereby reducing the (Spd+Spm)/Put (PAs) ratio in the leaves. The exogenous application of 1 mM Spd markedly reversed these CA-induced effects for polyamine and partially restored the PAs ratio and RuBPC activity in leaves. Methylglyoxal-bis (guanylhydrazone) (MGBG), which is an inhibitor of S-adenosylmethionine decarboxylase (SAMDC), results in the inability of activated cells to synthesize Spd and exacerbates the negative effects induced by CA. The exogenous application of 1 mM d-arginine ( d-Arg), which is an inhibitor of Put biosynthesis, decreased the levels of Put, but increased the PAs ratio and RuBPC activity in leaves. These results suggest that 0.1 mM CA inhibits RuBPC activity by decreasing the levels of endogenous free and perchloric acid soluble (PS) conjugated Spm, as well as the PAs ratio.

Conversion of protocorm-like bodies of Dendrobium huoshanense to shoots: The role of polyamines in relation to the ratio of total cytokinins and indole-3-acetic acidindole-3-acetic acid

In the present paper, a correlation between enhanced conversion of protocorm-like bodies (PLBs) of Dendrobium huoshanense to shoots by free polyamines (PAs) and changes in the levels of endogenous hormones is described. The endogenous levels of free spermidine (Spd) and putrescine (Put) increased during the conversion of PLBs to shoots. The exogenous addition of PAs, mainly Spd or Put at 2.0mM, not only elevated the endogenous levels of PAs but also promoted the frequency of conversion of PLBs to shoots. As compared with control, the enhanced conversion of PLBs to shoots by exogenous PAs was accompanied by an increase in ratio of total cytokinins (CTKs) to indole-3-acetic acid (IAA), which was due to decrease in the endogenous level of IAA and increase in the endogenous level of total CTKs, including the levels of isopentenyladenine+isopentenyladenine 9-riboside and zeatin+zeatin riboside. Analysis of enzyme activities showed that the increased endogenous level of total CTKs by PAs was related to the inhibition of CTK decomposition by CTK oxidase, while the decreased endogenous level of IAA was related to the promotion of IAA decomposition by IAA oxidase. Addition of PA biosynthetic inhibitors, involving alpha-difluoromethylornithine for Put and methylglyoxal(bis)-guanylhydrazone for Spd and Spm, decreased the conversion of PLBs to shoots, the ratio of total CTKs to IAA, and the levels of endogenous Put and Spd. This inhibition could be partly reversed by the application of exogenous Put or Spd.

Interaction Between Polyamine and Nitric Oxide Signaling in Adaptive Responses to Drought in Cucumber

The effect of polyamines (PAs) on nitric oxide (NO) generation was investigated in cucumber (Cucumis sativus cv. Dar) primary leaves using bio-imaging with an NO-selective fluorophor, DAF-2DA. Seedlings pretreated with PAs and subjected to water deficit showed early (after 5 h) and transient NO production. The amplitude of the response depended on the form of the applied polyamine. Spermine (1.0 mM) and spermidine (1.0 mM) induced higher NO-dependent fluorescence compared with putrescine (1.0 mM) and the control. The NO production was blocked by tungstate, an inhibitor of nitrate reductase, and partially by an inhibitor of nitric oxide synthase (NOS-like) enzymes. NO donor administration preceding drought had no effect on endogenous PA levels but was positively correlated with an alleviation of water deficit-induced membrane permeability and lipid peroxidation. Application of 2-(4-carboxyphenyl)-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide (cPTIO), a membrane-permeable NO scavenger, markedly reversed the NO donor effects. Similarly, pretreating seedlings with PAs resulted in lower ion leakage from the membrane and modified lipid peroxidation. The results indicate that NO may act downstream of PAs in cucumber seedlings under water stress.

Polyamine metabolism during the cell cycle of synchronized tobacco BY-2 cell line

The time courses of the contents of free, soluble and insoluble polyamine (PA) conjugates, PA biosynthetic and catabolic enzyme activities and mRNA levels of PA biosynthetic genes were monitored during the cell cycle of synchronized tobacco BY-2 cell line (Nicotiana tabacum L. cv. Bright Yellow 2). Progression through the cell cycle was characterized by specific biphasic changes of PA levels. The first, moderate peak in the amount of free PAs coincided with the S-phase. After a transient decline in G2 phase the contents of free PAs increased rapidly and peaked again during G2/M interface. Then sharply decreased with the minimum at the end of mitosis and during M/G1 transition and started to rise again with the next replication phase. Levels of PA soluble conjugates paralleled those of the free forms. Biosynthetic enzyme activities followed the biphasic manner analogous to the levels of free PAs and seemed to be regulated on both transcriptional and (post)translational level. PA cellular levels were further controlled by both catabolic degradation and conjugation of PAs. PA catabolism played an important role in the PA down-regulation during G2 phase and late mitosis, while the decline in free PAs in G2/M interface and during the whole mitosis resulted mainly from PA conjugation. This study's results demonstrate that during the cell cycle of tobacco BY-2 cells endogenous PA levels are intricately controlled, involving regulation of activities of biosynthetic, catabolic and conjugation enzymes.

Specific polyamine ratios as indicators of Pinus caribaea microshoot rooting phases

This study analysed the changes in the endogenous levels of total and individual polyamines (PAs) during the different rooting phases in Pinus caribaea microshoots: induction, induction-initiation and functional root system emergence. The analysis, performed using high-resolution liquid chromatography, demonstrated that during the inductive phase there is a drop in total PA content in all the fractions, with an increase in the degradation product diaminopropane. In the induction-initiation phase PAs are present in all the fractions. The microshoots with root nodules always presented higher Putrescine (Put) levels, whereas those that did not have them had similar spermidine and Put levels, while maintaining elevated spermine levels. The comparative analysis of microshoots with and without a rooting response indicates that high PA levels are maintained when root manifestation does not occur, coinciding with basal callus formation.

Niemann-Pick C1 Functions in Regulating Lysosomal Amine Content

Mutations in the late endosomal/lysosomal membrane protein Niemann-Pick C1 (NPC1) are known to cause a generalized block in retrograde vesicle-mediated transport, resulting in the hyper-accumulation of multiple lysosomal cargos. An important, yet often overlooked, category of lysosomal cargo includes the vast array of small molecular weight amine-containing molecules that are substrates for ion trapping in the highly acidic organelle lumen. We show here that the introduction of amine-containing molecules in lysosomes can significantly stimulate NPC1-mediated late endosome/lysosome fusion, and subsequently the secretion of lysosomal cargo. To illustrate the physiological importance of this NPC1-mediated transport pathway, we show that NPC1-deficient cells are more susceptible to the toxic effects of a lysosomotropic polyamine metabolite 3-aminopropanal. Moreover, NPC fibroblasts are shown to have higher levels of polyamine oxidase, an enzyme involved in the formation of 3-aminopropanal. Collectively, these findings provide strong support for a novel functional role for NPC1 and may also provide clues toward understanding NPC disease progression.

Putrescine Is Involved in Arabidopsis Freezing Tolerance and Cold Acclimation by Regulating Abscisic Acid Levels in Response to Low Temperature

The levels of endogenous polyamines have been shown to increase in plant cells challenged with low temperature; however, the functions of polyamines in the regulation of cold stress responses are unknown. Here, we show that the accumulation of putrescine under cold stress is essential for proper cold acclimation and survival at freezing temperatures because Arabidopsis (Arabidopsis thaliana) mutants defective in putrescine biosynthesis (adc1, adc2) display reduced freezing tolerance compared to wild-type plants. Genes ADC1 and ADC2 show different transcriptional profiles upon cold treatment; however, they show similar and redundant contributions to cold responses in terms of putrescine accumulation kinetics and freezing sensitivity. Our data also demonstrate that detrimental consequences of putrescine depletion during cold stress are due, at least in part, to alterations in the levels of abscisic acid (ABA). Reduced expression of NCED3, a key gene involved in ABA biosynthesis, and down-regulation of ABA-regulated genes are detected in both adc1 and adc2 mutant plants under cold stress. Complementation analysis of adc mutants with ABA and reciprocal complementation tests of the aba2-3 mutant with putrescine support the conclusion that putrescine controls the levels of ABA in response to low temperature by modulating ABA biosynthesis and gene expression.