Ivan Couée
email: ivan.couee@univ-rennes1.fr

Irène Hummel

Cécile Sulmon

Gwenola Gouesbet

Abdelhak El

Involvement of polyamines in root development

Keywords: adventitious root formation, agmatine, cell division and differentiation, lateral root formation, plant growth regulators, putrescine, root growth, spermidine, spermine ADC -arginine decarboxylase, DFMA -difluoromethylarginine, DFMO -difluoromethylornithine, DW -dry weight, ODC -ornithine decarboxylase

Root development is under the control of hormonal, metabolic, and environmental cues that can act on geneticallycontrolled developmental programmes and thus affect the plasticity of root architecture. These processes involve not only the five 'classical' plant hormones, but also other growth regulators, such as polyamines. The present review emphasises the importance of polyamines in the different aspects of root development: primary root growth and lateral and adventitious root formation. Free (agmatine, putrescine, spermidine, spermine), conjugated (such as hydroxycinnamate conjugates) and macromolecule-bound polyamines are reported to be present in root systems. Modifications of their endogenous levels by inhibitor treatment, by mutation, by gene manipulation, or by exogenous treatment can have drastic effects on root development and subsequent architecture. These effects may be related to the involvement of polyamines in the control of cell division and differentiation, which plays an important role in the root apex and during lateral and adventitious root formation. The exact mechanisms of action remain to be elucidated, but accumulating evidence in plant and animal cells supports the idea that, besides biophysical effects on membranes and nucleic acids, polyamines interact with protein kinases and transcription factors and are thus involved in signal transduction pathways. The high flexibility of polyamine metabolism in response to environmental stress and the metabolic link between polyamine and ethylene synthesis strongly suggest that polyamines may play a role in environmentally-induced plasticity of root development. Moreover, polyamines may be implicated in the establishment of biotic interactions between roots and rhizospheric micro-organisms.

Introduction

Root systems show considerable variation of architecture among plant species, among genotypes within the same species, and among the different regions of the same individual [START_REF] Lynch | Root architecture and plant productivity[END_REF]. This plasticity of root development probably represents an important adaptation of terrestrial plants in order to optimise utilisation of heterogeneous soil resources [START_REF] Bell | Dynamic phenotypic plasticity for root growth in polygonum: a comparative study[END_REF]. Development of root architecture mainly depends on -the growth of root individual axes, -the emergence of lateral roots from these root axes, and -the direction of elongation [START_REF] Lynch | Root architecture and plant productivity[END_REF].

Moreover, plants develop new roots from stems, petioles or leaves through adventitious processes.

The developmental stages of root growth and lateral and adventitious root formation result from the activation of various genetic programmes [START_REF] Rost | Root organization and gene expression patterns[END_REF]. A number of developmental genes involved in root apical meristem activity and in lateral and adventitious root formation have thus been characterised [START_REF] Smith | LRP1, a gene expressed in lateral and adventitious root primordia of Arabidopsis[END_REF][START_REF] Scheres | Roots redefined: anatomical and genetic analysis of root development[END_REF]. It is also well established that root development is controlled by hormonal signals, especially auxins [START_REF] Celenza | A pathway for lateral root formation in Arabidopsis thaliana[END_REF], ant that environmental cues can act on the genetic programmes of root development and on their hormonal and metabolic control [START_REF] Zhang | Dual pathways for regulation of root branching by nitrate[END_REF]. In this complex network of interactions resulting in root developmental plasticity, other growth regulators than the five 'classical' plant hormones (Kende and Zeevart, 1997) may be involved [START_REF] Mathesius | Auxin transport inhibition precedes root nodule formation in white clover roots and is regulated by flavonoids and derivatives of chitin oligosacharides[END_REF]. Thus, a large corpus of literature gives ample evidence that polyamines play an important role in primary, lateral and adventitious root development.

Polyamines are low molecular mass polycations that are found in all living organisms [START_REF]Metabolism and function of polyamines in plants: recent development (new approaches)[END_REF]. They are generally considered to be growth regulators, and, in higher plants, they have been implicated in a range of developmental processes [START_REF] Smith | Polyamines[END_REF][START_REF] Evans | Do polyamines have roles in plant development?[END_REF][START_REF] Galston | Polyamines and plant morphogenesis[END_REF][START_REF]Metabolism and function of polyamines in plants: recent development (new approaches)[END_REF]. Like in animals and in bacteria, polyamines could play a major role in proliferation and growth of plant cells, since application of exogenous polyamines stimulates development in several higher plants, suggesting that endogenous concentrations of these amines could be growth-limiting [START_REF] Galston | Polyamines and plant morphogenesis[END_REF]. As polyamine metabolism is responsive to external conditions, polyamines may play an important role in interactions between environment and development [START_REF] Galston | Plant polyamines in reproductive activity and response to abiotic stress[END_REF][START_REF] Bouchereau | Polyamines and environmental challenges: recent development[END_REF].

In plants, the diamine putrescine is generally synthesized by two pathways (Figure 1). One pathway starts with ornithine to give putrescine via ornithine decarboxylase (ODC), the other produces agmatine via arginine decarboxylase (ADC) followed by additional steps to produce putrescine [START_REF] Smith | Plant amines[END_REF]. A number of plant species have been shown to possess both enzyme systems. In other cases, such as that of Arabidopsis thaliana, ADC genes and enzymes have been clearly identified [START_REF] Galloway | Phylogenetic utility of the nuclear gene arginine decarboxylase: an example from Brassicaceae[END_REF], whereas evidence for ODC is lacking [START_REF] Hanfrey | Arabidopsis polyamine biosynthesis: absence of ornithine decarboxylase and the mechanism of arginine decarboxylase activity[END_REF]. Putrescine conversion to spermidine and then to spermine (Figure 1) occurs by successive transfers of aminopropyl groups from decarboxylated S-adenosylmethionine [START_REF] Martin-Tanguy | Polyamines in gravepine microcuttings cultivated in-vitro -effects of amines and inhibitors of polyamine biosynthesis on polyamine levels and microcutting growth and development[END_REF]. However, the dynamics of polyamine metabolism is complexified by the existence of degradation (Figure 1) and conjugation pathways and of transport and uptake mechanisms [START_REF]Metabolism and function of polyamines in plants: recent development (new approaches)[END_REF]. Besides biophysical effects through their positive charge at physiological pH [START_REF] Smith | Polyamines[END_REF], polyam-ines may be involved in signal transduction pathways, through effects on calcium fluxes [START_REF] Thomas | Inhibition of calcium signalling in murine splenocytes by polyamines: differential effects on CD4 and CD8 T-cells[END_REF], and interactions with transcription factors [START_REF] Wang | Cloning and characterization of polyamine-modulated factor-1, a transcriptional cofactor that regulates the transcription of the spermidine/spermine N1-acetyltransferase gene[END_REF], protein kinases [START_REF] Datta | Spermine stimulation of a nuclear NII kinase from pea plumules and its role in the phosphorylation of a nuclear polypeptide[END_REF], and 14-3-3 proteins [START_REF] Athwal | Divalent cations and polyamines bind to loop 8 of 14-3-3 proteins, modulating their interaction with phosphorylated nitrate reductase[END_REF]. Finally, polyamines have been shown to interact with phytohormones [START_REF] Altman | Polyamines and plant hormones[END_REF][START_REF] Alabadi | Arginase, arginine decarboxylase, ornithine decarboxylase, and polyamines in tomato ovaries[END_REF][START_REF] Tonon | Changes in polyamines, auxins and peroxidase activity during in vitro rooting of Fraxinus angustifolia shoots: an auxin-independent rooting model[END_REF].

The aim of this review is to assess the physiological importance of these correlations between polyamines and root development, and to present possible mechanisms of action of polyamines at the biochemical and molecular levels.

Identification and characterisation of polyamines in roots

Classical procedures for extracting and analysing polyamines [START_REF] Flores | Analysis of polyamines in higher plants by high performance liquid chromatography[END_REF][START_REF] Smith | Separation and quantification of polyamines in plant tissue by high performance liquid chromatography of their dansyl derivatives[END_REF] have identified in root tissues the presence of -free polyamines, such as agmatine, putrescine, spermidine, and spermine, -soluble conjugated polyamines, such as caffeoylputrescine and feruloylputrescine, and -insoluble bound polyamines [START_REF] Shen | Correlations between polyamine ratios and growth patterns in seedling roots[END_REF][START_REF] Martin-Tanguy | Inverse relationship between polyamine levels and the degree of phenotypic alteration induced by the rootinducing, left-hand transferred DNA from Agrobacterium rhizogenes[END_REF]Ben-Hayyim et al., 1994, 1996;[START_REF] Aribaud | Transglutaminaselike activity in Chrysanthemum leaf explants cultivated in vitro in relation to cell growth and hormone treatment[END_REF][START_REF] Watson | Arginine decarboxylase (polyamine synthesis) mutants of Arabidopsis thaliana exhibit altered root growth[END_REF][START_REF] Hennion | Amines of the subantarctic crucifer Pringlea antiscorbutica are responsive to temperature conditions[END_REF][START_REF] Tassoni | Polyamine content and metabolism in Arabidopsis thaliana and effect of spermidine on plant development[END_REF][START_REF] Hummel | Involvement of polyamines in root development at low temperature in the subantarctic cruciferous species Pringlea antiscorbutica[END_REF]. For instance, perchloric acid extraction from roots of Arabidopsis thaliana yields free polyamines and also conjugated or bound polyamines, in the perchloricacid-soluble and perchloric-acid-insoluble fractions [START_REF] Tassoni | Polyamine content and metabolism in Arabidopsis thaliana and effect of spermidine on plant development[END_REF]. Acetyl conjugates of polyamines have been reported in a very small number of higher plants. Thus, in the case of the subantarctic cruciferous species Pringlea antiscorbutica, Kerguelen cabbage, N1-acetylputrescine has been shown to exist in the roots [START_REF] Hennion | Amines of the subantarctic crucifer Pringlea antiscorbutica are responsive to temperature conditions[END_REF]. Besides, polyamines can be the precursors of several hundred alkaloids [START_REF]Metabolism and function of polyamines in plants: recent development (new approaches)[END_REF], some of which can be associated with root development. For instance, in species of the Asteraceae producing pyrrolizidine alkaloids, the synthesis is exclusively restricted to roots, and seems to be correlated with root growth, as it is reduced or ceases when root growth stops [START_REF] Ober | Homospermidine synthase, the first pathway-specific enzyme of pyrrolizidine alkaloid biosynthesis, evolved from deoxyhypusine synthase[END_REF]. Some plant cells seem to have the ability to synthesize uncommon long-chain polyamines previously repor- ted only in thermophilic bacteria [START_REF] Bouchereau | Polyamines and environmental challenges: recent development[END_REF]. Thus, [START_REF] Roy | Polyamines, both common and uncommon, under heat stress in rice (Oryza sativa) callus[END_REF] reported the production of long-chained polyamines in rice callus under heat stress. Such long-chained polyamines do not appear to have been reported yet in root tissues. However, other unusual polyamines, such as 4-aminobutylcadaverine, have been described in root nodules of bean plants [START_REF] Fujihara | A new polyamine 4aminobutylcadaverine, occurrence and its biosynthesis in root nodules of adzuki bean plant Vigna angularis[END_REF]. Finally, polyamine-binding transglutaminase-like activity and macromolecule-bound polyamines have been identified in roots initiated from leaf explants [START_REF] Aribaud | Transglutaminaselike activity in Chrysanthemum leaf explants cultivated in vitro in relation to cell growth and hormone treatment[END_REF].

Modifications of polyamine levels and root development

Studies in a number of plant species have shown that overall depletion of polyamine pools was linked with root growth inhibition. Indeed, decrease of polyamine level is often associated with ageing and senescence whereas accumulation, more precisely spermidine and spermine accumulation, is associated with growing tissue activity and organogenesis [START_REF] Perez-Amador | Expression of arginine decarboxylase is induced during early fruit development and in young tissues of Pisum sativum (L)[END_REF]. For example, in Phaseolus vulgaris, deple-tion of putrescine, spermidine, and spermine levels, resulting from decreased ADC activity, leads to decrease of root length [START_REF] Palavan-Unsal | Polyamine metabolism in the roots of Phaseolus vulgaris. Interaction of inhibitors of polyamine biosynthesis with putrescine in growth and polyamine biosynthesis[END_REF]. Polyamine accumulation has also been correlated to adventitious root formation [START_REF] Friedman | Polyamines and root formation in mung bean hypocotyl cuttings. I. Effect of exogenous compounds and changes in endogenous polyamines[END_REF][START_REF] Friedman | Polyamines and root formation in mung bean hypocotyl cuttings. II. Incorporation of precursors into polyamines[END_REF][START_REF] Jarvis | Involvement of polyamines with adventitious root development in stem cuttings of mung bean[END_REF][START_REF] Altamura | De novo root formation in tobacco thin layers affected by inhibition of polyamine biosynthesis[END_REF][START_REF] Altamura | Cytological events induced by the inhibition of polyamine biosynthesis in thin layers of tobacco[END_REF].

Difluoromethylarginine (DFMA), a specific inhibitor of ADC [START_REF] Burtin | Effects of the suicide inhibitors of arginine and ornithine decarboxylase activities on organogenesis, growth, free polyamine and hydroxycinnamoylputrescine levels in leaf explants of Nicotiana Xanthi n.c. cultivated in vitro in a medium producing callus formation[END_REF], has been shown to decrease root elongation in lateral roots in hairy root cultures [START_REF] Biondi | Hairy root cultures of Hyoscyamus-musticus -effect of polyamine biosynthesis inhibitors[END_REF]. Martin-Tanguy and Carré (1993) also showed that DFMA inhibited adventitious root growth from gravepine microcuttings. This probable involvement of the ADC pathway in root development was confirmed by the characterization of spe mutants of Arabidopsis, affected in ADC activity levels [START_REF] Watson | Arginine decarboxylase (polyamine synthesis) mutants of Arabidopsis thaliana exhibit altered root growth[END_REF]. Thus, spe double mutants exhibit highly kinked roots forming a tight cluster [START_REF] Watson | Arginine decarboxylase (polyamine synthesis) mutants of Arabidopsis thaliana exhibit altered root growth[END_REF]. High ADC activity has been found in roots of Arabidopsis [START_REF] Watson | Arginine decarboxylase (polyamine synthesis) mutants of Arabidopsis thaliana exhibit altered root growth[END_REF][START_REF] Hanfrey | Arabidopsis polyamine biosynthesis: absence of ornithine decarboxylase and the mechanism of arginine decarboxylase activity[END_REF] and high ADC mRNA levels are present in roots of soybean seedlings [START_REF] Nam | Differential expression of ADC mRNA during development and upon acid stress in soybean (Glycine max.) hypocotyls[END_REF]. In Arabidopsis, one of the two ADC genes is strongly expressed in roots [START_REF] Piotrowski | Plant C-N hydrolases: identification of a plant N-carbamoylputrescine amidohydrolase involved in polyamine biosynthesis[END_REF], and one of the two ADC promoters, which show distinct patterns of activity [START_REF] El Amrani | Genome-wide distribution and potential regulatory functions of AtATE, a novel family of miniature inverted-repeat transposable elements in Arabidopsis thaliana[END_REF], is highly active in roots (Hummel I, Bourdais G, Gouesbet G, Couée I, Malmberg RL, El Amrani A, submitted article). In Kerguelen cabbage, modifications with inhibitors revealed a positive correlation between increase of endogenous agmatine and stimulation of root growth [START_REF] Hummel | Involvement of polyamines in root development at low temperature in the subantarctic cruciferous species Pringlea antiscorbutica[END_REF]. Thus, it could be hypothesized that the size of the agmatine pool might be a direct link between ADC activity and root development. Such a link would imply the direct involvement of agmatine in mechanisms of plant development, which requires further investigation. At present, only studies on animal cells point out to the existence of protein targets of agmatine, such as receptors or enzymes [START_REF] Reis | Is agmatine a novel neurotransmitter in brain?[END_REF].

A positive correlation between spermidine or spermine contents and primary root growth is also generally found. Moreover, decrease of spermidine and spermine levels by cyclohexylammonium treatment, which inhibits spermidine synthase [START_REF] Martin-Tanguy | Polyamines in gravepine microcuttings cultivated in-vitro -effects of amines and inhibitors of polyamine biosynthesis on polyamine levels and microcutting growth and development[END_REF], has been shown to result in a decrease of lateral root development [START_REF] Hummel | Involvement of polyamines in root development at low temperature in the subantarctic cruciferous species Pringlea antiscorbutica[END_REF]. Since it is well established that primary root growth and lateral root formation involve intense mitotic activity and are dependent on an array of endogenous physiological factors [START_REF] Burtin | Polyamines, hydroxycinnamoylputrescines and root formation in leaf explants of tobacco cultivated in vitro -effects of the suicide inhibitors of putrescine synthesis[END_REF], the positive connection that is observed between spermidine and spermine contents and root development could be related to spermidine and spermine involvement in cell cycle activity, growth and differentiation [START_REF] Fuller | Polyamine biosynthesis and accumulation during the G1 to S phase transition[END_REF][START_REF] Rupniak | Inhibition of spermidine and spermine synthesis leads to growth arrest of rat embryo fibroblast in G1[END_REF]. Endogenous spermine content of less than 0.1 µmol g -1 DW in roots of untreated plantlets of Kerguelen cabbage was in the range of concentrations that gave large variations of root growth rate [START_REF] Hummel | Involvement of polyamines in root development at low temperature in the subantarctic cruciferous species Pringlea antiscorbutica[END_REF]. Finally, genes encoding spermine synthase and putative spermidine synthases have been shown to be strongly expressed in roots of Arabidopsis [START_REF] Hanzawa | Characterisation of the spermidine synthase-related gene family in Arabidopsis thaliana[END_REF]. Spermidine and spermine may thus be good candidates for regulation of root development.

In some plant species, treatments leading to putrescine accumulation in seedling root suggest a negative connection, within a certain concentration range, between putrescine level in root and growth rate of primary root. Thus, in roots excised from plants of Nicotiana tabacum, difluoromethylornithine (DFMO), the specific inhibitor of ODC [START_REF] Burtin | Effects of the suicide inhibitors of arginine and ornithine decarboxylase activities on organogenesis, growth, free polyamine and hydroxycinnamoylputrescine levels in leaf explants of Nicotiana Xanthi n.c. cultivated in vitro in a medium producing callus formation[END_REF], was found to cause an increase in root length in parallel with putrescine depletion, agmatine accumulation and no variation of spermidine pool [START_REF] Ben-Hayyim | Changing root and shoot architecture with the rolA gene from Agrobacterium rhizogenes: interactions with gibberellic acid and polyamine metabolism[END_REF]. In these excised roots, DFMO caused a fundamental change in root architecture with increase of primary root length. These changes were reversed when 1 mM putrescine was included in the DFMO treatment, thus indicating that effects of DFMO on development were indeed due to putrescine limitation. In this study, DFMO decreased free putrescine level in excised roots by a factor of 3 and exogenous putrescine prevented this effect whereas the spermidine titer remained unchanged [START_REF] Ben-Hayyim | Changing root system architecture through inhibition of putrescine and feruloyl putrescine accumulation[END_REF]. In other plant species, such as Kerguelen cabbage, large variations of the endogenous putrescine pool, from 2 to 12 µmol.g -1 DW, did not have any effect on root growth, but at concentrations lower than 2 µmol g -1 DW, decrease of the putrescine pool seemed to enhance primary root growth [START_REF] Hummel | Involvement of polyamines in root development at low temperature in the subantarctic cruciferous species Pringlea antiscorbutica[END_REF]. However, [START_REF] Lee | Polyamine regulation of growth and chilling tolerance of rice (Oryza sativa L.) roots cultured in vitro[END_REF] had shown that exogenously-added putrescine in concentrations varying from 0.01 to 1 mM enhanced elongation in excised rice roots under in vitro conditions at 25 • C. In the same study, DFMO was found to inhibit root elongation and polyamine levels and these effects were reversed by putrescine. In contrast, spermidine inhibited root elongation at concentrations greater than 0.1 mM and spermine, at all concentrations used, inhibited elongation of roots grown at 25 • C [START_REF] Lee | Polyamine regulation of growth and chilling tolerance of rice (Oryza sativa L.) roots cultured in vitro[END_REF]. Similarly, [START_REF] Tarenghi | Effects of inhibitors of polyamine biosynthesis and of polyamines on strawberry microcutting growth and development[END_REF] showed that, in strawberry microcuttings, 1 mM putrescine treatment led to increase of putrescine levels, greater number of adventitious roots and increase of root length. Taken together, all of these studies emphasize important inter-species differences, which might also depend on culture conditions. In spite of these discrepancies, a number of unrelated dicot species, such as Nicotiana tabacum, Phaseolus vulgaris, Vitis vinifera, Arabidopsis thaliana and Pringlea antiscorbutica seem to share common features of polyamine involvement in root development.

In contrast to the effects of endogenous increase [START_REF] Hummel | Involvement of polyamines in root development at low temperature in the subantarctic cruciferous species Pringlea antiscorbutica[END_REF], addition of 1 mM spermidine to culture media has been found to result in significant inhibition of root growth in Arabidopsis [START_REF] Tassoni | Polyamine content and metabolism in Arabidopsis thaliana and effect of spermidine on plant development[END_REF]. Similarly, we have found that exogenous spermine significantly inhibited root growth of Arabidopsis (our unpublished data). Exogenous treatments with spermidine and spermine result in major increases respectively of spermidine and putrescine [START_REF] Tassoni | Polyamine content and metabolism in Arabidopsis thaliana and effect of spermidine on plant development[END_REF], and of spermine (our unpublished data), in root tissues. Thus, exogenous treatments would seem to give significantly different results from those of inhibitor-based modifications of endogenous levels. This difference may be ascribed to the range of concentrations that were used. [START_REF] Locke | Contrasting effects of ethylene perception and biosynthesis inhibitors on germination and seedling growth of barley (Hordeum vulgare L.)[END_REF] reported enhanced root growth of barley seedlings treated with 1 µM putrescine, spermidine, or spermine. However, other mechanisms may be involved. Thus, uptake of exogenous polyamines and endogenous control of polyamine biosynthesis may not have the same developmental impact. Verification of such hypothesis would require further characterization of polyamine uptake systems in plant cells. Finally, the overall balance of the different free polyamines may be more important than a specific role of each polyamine. Thus, correlated changes of putrescine, spermine, spermidine are observed during seedling root development [START_REF] Shen | Correlations between polyamine ratios and growth patterns in seedling roots[END_REF], and regulations induced by inhibitor treatments or mutations result in modifications of more than one free polyamine [START_REF] Burtin | Effects of the suicide inhibitors of arginine and ornithine decarboxylase activities on organogenesis, growth, free polyamine and hydroxycinnamoylputrescine levels in leaf explants of Nicotiana Xanthi n.c. cultivated in vitro in a medium producing callus formation[END_REF][START_REF] Ben-Hayyim | Changing root and shoot architecture with the rolA gene from Agrobacterium rhizogenes: interactions with gibberellic acid and polyamine metabolism[END_REF][START_REF] Watson | Arginine decarboxylase (polyamine synthesis) mutants of Arabidopsis thaliana exhibit altered root growth[END_REF][START_REF] Hummel | Involvement of polyamines in root development at low temperature in the subantarctic cruciferous species Pringlea antiscorbutica[END_REF].

Besides direct effects, the relationship between polyamine metabolism, ethylene synthesis, and aminoacid metabolism may have an impact on root development. For instance, genetic modification of tryptophan decarboxylase and tyrosine decarboxylase pathways, which yield monoamines, can result in root-curling phenotypes, which have been ascribed to depletion of tryptophan pool [START_REF] Guillet | Expression of tryptophan decarboxylase and tyrosine decarboxylase genes in tobacco results in altered biochemical and physiological phenotypes[END_REF]. Ethylene is known to be involved in root development [START_REF] Ross | New interactions between classical plant hormones[END_REF], and treatment with methylglyoxal-bis-guanylhydrazone, which inhibits [START_REF] Bagni | Effect of various inhibitors of polyamine synthesis on the growth of Helianthus tuberosus[END_REF] the production of decarboxylated S-adenosylmethionine (Figure 1) and may thus provide more S-adenosylmethionine for ethylene production [START_REF] Locke | Contrasting effects of ethylene perception and biosynthesis inhibitors on germination and seedling growth of barley (Hordeum vulgare L.)[END_REF], has a strongly negative impact on root development in Kerguelen cabbage [START_REF] Hummel | Involvement of polyamines in root development at low temperature in the subantarctic cruciferous species Pringlea antiscorbutica[END_REF] and Arabidopsis [START_REF] Tassoni | Polyamine content and metabolism in Arabidopsis thaliana and effect of spermidine on plant development[END_REF]. In a similar way, [START_REF] Locke | Contrasting effects of ethylene perception and biosynthesis inhibitors on germination and seedling growth of barley (Hordeum vulgare L.)[END_REF] examined the effects of inhibitors of the ethylene pathway on germination in terms of modifications of polyamine levels, and came to the conclusion that during barley germination polyamines played a complementary, growth-promotive role to ethylene. The relationship between polyamine and gibberellin actions, which has been described in more detail for floral development [START_REF] Alabadi | Arginase, arginine decarboxylase, ornithine decarboxylase, and polyamines in tomato ovaries[END_REF][START_REF] Hanzawa | ACAULIS5, an Arabidopsis gene required for stem elongation, encodes a spermine synthase[END_REF], may also potentially be responsible for modifications of root development. Thus, [START_REF] Ben-Hayyim | Changing root and shoot architecture with the rolA gene from Agrobacterium rhizogenes: interactions with gibberellic acid and polyamine metabolism[END_REF] showed that treatment with gibberellic acid could reverse the effects of DFMO on tobacco roots. In the acaulis5 mutant of Arabidopsis, which is mutated in a gene encoding a spermine synthase, transcript levels of gibberellin-related genes are affected [START_REF] Hanzawa | ACAULIS5, an Arabidopsis gene required for stem elongation, encodes a spermine synthase[END_REF]. However, the acaulis5 mutant plants apparently show little morphological defects in other organs than stems and flowers [START_REF] Hanzawa | ACAULIS5, an Arabidopsis gene required for stem elongation, encodes a spermine synthase[END_REF], although the ACAULIS5 gene is strongly expressed in roots [START_REF] Hanzawa | Characterisation of the spermidine synthase-related gene family in Arabidopsis thaliana[END_REF].

The machinery for polyamine metabolism is generally present in the cells or tissues where their action takes place (Martin-Tanguy, 2001). However, despite scarcity of relevant studies, the possibility of longdistance transport, for instance in xylem, of polyamines must be considered [START_REF] Bagni | Uptake and transport of polyamines and inhibitors of polyamine metabolism in plants[END_REF]. Furthermore, numerous polyamine-derived alkaloids are synthesised in roots and transported to shoots by the xylem [START_REF]Metabolism and function of polyamines in plants: recent development (new approaches)[END_REF]. Further studies on such long-distance transport may open the possibility that polyamines or their derivatives play a role in the exchange of information between root and shoot development.

Control of cell division and differentiation in the root apex and during lateral and adventitious root formation

Longitudinal growth of root individual axes largely relies on the activity of root apical meristems, which involves active division and differentiation in meristem initials and active expansion and differentiation in the elongation zone [START_REF] Scheres | Roots redefined: anatomical and genetic analysis of root development[END_REF]. Lateral root formation requires a succession of steps with induction of primordia showing high meristematic activity and then organisation of lateral root meristems [START_REF] Malamy | Organization and cell differentiation in lateral roots of Arabidopsis thaliana[END_REF]. Adventitious root formation follows similar steps and share common mechanisms with lateral root formation [START_REF] Lund | Cuttings of a tobacco mutant, rac, undergo cell divisions but do not initiate adventitious roots in response to exogenous auxin[END_REF][START_REF] Ermel | Mechanisms of primordium formation during adventitious root development from walnut cotyledon explants[END_REF]. Such importance of meristematic activity for root growth has been shown to depend on gene expression control. For instance, control of cyclin gene expression and therefore abundance of cyclin may function to allow flexible root growth control and induction of lateral roots [START_REF] Doerner | Control of root growth and development by cyclin expression[END_REF]. Lateral root formation is initiated by auxin-mediated mitogenic activation, which lifts the G2 blockage of pericycle cells and launches a G2/M transition [START_REF] Clark | Root formation in ethylene-insensitive plants[END_REF]. The induction of lateral root formation is thus related to a specific step of the cell cycle.

Polyamines have been associated with cell proliferation in animal and plant cells [START_REF] Theiss | Regulation by polyamines of ornithine decarboxylase activity and cell division in the unicellular green alga Chlamydomonas reinhardtii[END_REF]. Increased levels of spermidine and spermine have often been found to occur during the transition from G1 to the S phase of cell cycle in animals and plants [START_REF] Fuller | Polyamine biosynthesis and accumulation during the G1 to S phase transition[END_REF], and conversely, inhibition of polyamine synthesis has been found to block the cell cycle [START_REF] Rupniak | Inhibition of spermidine and spermine synthesis leads to growth arrest of rat embryo fibroblast in G1[END_REF]. One can relate this involvement in G1/S transition with effects of polyamines on DNA configuration [START_REF] Bouchereau | Polyamines and environmental challenges: recent development[END_REF]. Other studies also point out to the involvement of polyamines in the control of G2/M transition [START_REF] Fowler | Polyamine metabolism and gene regulation during the transition of autonomous sugar beet cell in suspension culture from quiescence to division[END_REF].

It is also generally accepted that major changes of polyamine levels are associated with the control of the balance between division and differentiation [START_REF] Shen | Correlations between polyamine ratios and growth patterns in seedling roots[END_REF], and it has been suggested that the ODC pathway is active in dividing cells, whereas the ADC pathway predominates in mature tissue [START_REF] Flores | Changes in polyamine metabolism in response to abiotic stress[END_REF]. At any rate, there is clear evidence from in situ studies that increased polyamine synthesis is associated with lateral root formation and meristematic activity [START_REF] Schwartz | Localization of ornithine decarboxylase and changes in polyamine content in root meristems of Zea mays[END_REF]. However, complete experimental characterisation of this differential role for ADC-and ODC-mediated pathways remains to be carried out [START_REF] Granell | In situ hybridisation of ADC and ODC in different vegetative and reproductive tissues of the tomato[END_REF], and plants that appear to lack ODC, such as Arabidopsis, necessarily rely on considerably distinct mechanisms or on different roles for ADC [START_REF] Hanfrey | Arabidopsis polyamine biosynthesis: absence of ornithine decarboxylase and the mechanism of arginine decarboxylase activity[END_REF]. It must be pointed out however that DFMO treatment of Arabidopsis seedlings results in severe stunting of roots and of aerial organs [START_REF] Hanfrey | Arabidopsis polyamine biosynthesis: absence of ornithine decarboxylase and the mechanism of arginine decarboxylase activity[END_REF]our unpublished data).

Polyamine metabolism therefore shows plasticity in direct relationship with cellular processes of division and differentiation that are essential in root development. Indeed, in the roots of different species such as pea, tomato and maize, putrescine content increases as elongation progresses and is higher in differentiating zones, whereas spermine and spermidine are most abundant near apices [START_REF] Shen | Correlations between polyamine ratios and growth patterns in seedling roots[END_REF]. However, this would imply that other steps of polyamine metabolism than ADC and ODC must be differentially regulated during root growth and development. This remains to be verified experimentally. Moreover, involvement of polyamines in other essential aspects of root development such as columella or root hair development does not appear to have been studied yet.

Polyamines and environmental regulation of root development

Roots are often a primary target of environmental stresses, such as soil flooding or drought, which in turn have considerable impact on root development [START_REF] Vartanian | Drought rhizogenesis in Arabidopsis thaliana: differential response of hormonal mutants[END_REF]. A lot of these responses to stress involve changes in the balance of phytohormones which are known to have effects on root development [START_REF] Ross | New interactions between classical plant hormones[END_REF]. The relationship between ethylene and polyamines [START_REF] Locke | Contrasting effects of ethylene perception and biosynthesis inhibitors on germination and seedling growth of barley (Hordeum vulgare L.)[END_REF] has been discussed above, and [START_REF] Lee | Role of abscissic acid in chilling tolerance of rice (Oryza sativa L.) seedlings. 2. Modulation of free polyamine levels[END_REF] have reported the effect of abscisic acid on modulating the levels of polyamines.

On the other hand, independently of hormonemediated effects, environmental stress has been shown to induce considerable increase or decrease of polyamine levels, depending on type of stress, plant species and time of stress application [START_REF] Flores | Changes in polyamine metabolism in response to abiotic stress[END_REF][START_REF] Galston | Plant polyamines in reproductive activity and response to abiotic stress[END_REF][START_REF] Ali | Role of putrescine in salt tolerance of Atropa belladonna plant[END_REF][START_REF] Hennion | Amines of the subantarctic crucifer Pringlea antiscorbutica are responsive to temperature conditions[END_REF]. Moreover, in some cases, artificial modulations of polyamine contents have been shown to mimic external stimuli [START_REF] Hennion | Amines of the subantarctic crucifer Pringlea antiscorbutica are responsive to temperature conditions[END_REF]. In a number of stresses or of stress-related situations, such as salinity [START_REF] Chattopadhyay | Expression of arginine decarboxylase in seedlings of indica rice (Oryza sativa L.) cultivars as affected by salinity stress[END_REF][START_REF] Soyka | Arabidopsis knockout mutation of ADC2 gene reveals inducibility by osmotic stress[END_REF] or acid stress [START_REF] Nam | Differential expression of ADC mRNA during development and upon acid stress in soybean (Glycine max.) hypocotyls[END_REF], which are known to affect root development, the response to stress involves elevated levels of ADC or ODC activity or mRNA. This may have a direct impact on root development for the reasons presented above. However, the relationship between polyamines and response to stress is more complex than mere induction of polyamine biosynthetic pathways, with involvement of all the different classes of polyamines and of biosynthetic and degradation pathways [START_REF] Bouchereau | Polyamines and environmental challenges: recent development[END_REF]. Thus, increase of the activities of biosynthetic enzymes may finally result in increase of free polyamines or conjugated polyamines, which are likely to have different impacts on root growth.

Situations of acute stress are generally associated with reduction of root growth, or changes of root developmental programmes [START_REF] Vartanian | Drought rhizogenesis in Arabidopsis thaliana: differential response of hormonal mutants[END_REF]. However, root development under sub-optimal conditions, short of situations of acute stress, has received little attention. For instance, little is known about root development under chilling temperatures, although they may have a strong impact on root growth rate. When cultivated at 10 • C, soybean (Glycine max cv Aldana) primary root growth is slowed down to 1 mm.d -1 , which corresponds to a 10-fold decrease relative to growth at 25 • C [START_REF] Janas | Alterations in phenylpropanoid content in soybean roots during low temperature acclimation[END_REF]. In contrast, rate of root growth in Kerguelen cabbage showed a 2-fold decrease, relative to growth at 5-10 • C, when plants were cultivated at 25 • C [START_REF] Hennion | Amines of the subantarctic crucifer Pringlea antiscorbutica are responsive to temperature conditions[END_REF]. This species is characterised by high levels of polyamines, especially agmatine [START_REF] Hennion | Amines of the subantarctic crucifer Pringlea antiscorbutica are responsive to temperature conditions[END_REF], and agmatine levels show a drastic decrease upon heat treatment in parallel with termination of root growth [START_REF] Hennion | Amines of the subantarctic crucifer Pringlea antiscorbutica are responsive to temperature conditions[END_REF]. This result and the fact that, as discussed above, root growth rate seems to be positively related to agmatine content [START_REF] Hummel | Involvement of polyamines in root development at low temperature in the subantarctic cruciferous species Pringlea antiscorbutica[END_REF] strongly indicate that agmatine may be important not only in normal root growth, but also in sustaining root growth under chilling conditions.

Under such non-optimal or low-stress conditions where root growth is maintained, active changes of root architecture may participate in the response to environmental constraints. In heterogeneous soil, plasticity of root architecture could confer an advantage depending on resource distribution, including water, and edaphic constraints [START_REF] Lynch | Root architecture and plant productivity[END_REF]. [START_REF] Ben-Hayyim | Changing root system architecture through inhibition of putrescine and feruloyl putrescine accumulation[END_REF] showed that, in Nicotiana tabacum, modifying polyamine contents by DFMO treatment could convert a root system consisting of short roots of similar length to one having a dominant tap root from which numerous lateral roots were formed. This increase of overall root length and of lateral roots, with potentially deeper foraging of water resource, can be considered as a potentially adaptive response to drought stress [START_REF] Ben-Hayyim | Changing root system architecture through inhibition of putrescine and feruloyl putrescine accumulation[END_REF][START_REF] Tepfer | Control of root system architecture through chemical and genetic alterations of polyamine metabolism In: Davis TD & Haissig BE[END_REF].

In the soil, root development and acquisition of root architecture as well as root functions greatly rely on interactions of root tissues with the surrounding biotic environment [START_REF] O'connell | Engineering the rhizosphere: expressing a bias[END_REF], especially micro-organisms. El [START_REF] El Ghachtouli | First report of the inhibition of arbuscular mycorrhizal infection of Pisum sativum by specific and irreversible inhibition of polyamine biosynthesis or by gibberellic acid treatment[END_REF] showed that DFMO treatment, with reversion by exogenous putrescine, strongly inhibited root growth and arbuscular mycorrhizal infection of Pisum sativum. A possible role of polyamines in arbuscular mycorrhizal infection was therefore postulated. Similarly, ectomycorrhizal fungi were recently shown to promote root growth of scots pine through the potential involvement of the diamine cadaverine [START_REF] Niemi | Effects of exogenous diamines on the interaction between ectomycorrhizal fungi and adventitious root formation in Scots pine in vitro[END_REF].

Free polyamines, mainly putrescine, have been detected in root exudates of tomato [START_REF] Kuiper | Increased uptake of putrescine in the rhizosphere inhibits competitive root colonization by Pseudomonas fluorescens strain WCS365[END_REF] and Arabidopsis (our unpublished data). On the other hand, bacteria, including species that are known to be present in the soil, are known to possess a complex array of polyamine transport systems in the plasma membrane [START_REF] Kuiper | Increased uptake of putrescine in the rhizosphere inhibits competitive root colonization by Pseudomonas fluorescens strain WCS365[END_REF]. It is therefore reasonable to assume that polyamines may be part of the exchanges between root tissues and surrounding bacteria in the rhizosphere [START_REF] Fogel | Roots as primary producers in below-ground ecosystems[END_REF]. Thus, increased uptake of putrescine has been shown to have a bacteriostatic effect on Pseudomonas sp. [START_REF] Kuiper | Increased uptake of putrescine in the rhizosphere inhibits competitive root colonization by Pseudomonas fluorescens strain WCS365[END_REF]. At another level of involvement, inter-action between plants and soil-borne Agrobacterium rhizogenes results in possible introduction of a rootinducing, left-hand transferred DNA in plant tissues [START_REF] Tepfer | Transformation of several species of higher plants by Agrobacterium rhizogenes: sexual transmission of the transformed genotype and phenotype[END_REF], which carries genes that may profoundly modify root and shoot development. Polyamine metabolism in root cultures is altered by expression of rol genes from this left-hand transferred DNA [START_REF] Altabella | Effect of the rol genes from Agrobacterium rhizogenes on polyamine metabolism in tobacco roots[END_REF]. For instance, over-expression of the rolA gene under the control of the 35S promoter increases growth and ramification of excised roots, and results in depletion of putrescine and putrescine conjugates [START_REF] Ben-Hayyim | Changing root and shoot architecture with the rolA gene from Agrobacterium rhizogenes: interactions with gibberellic acid and polyamine metabolism[END_REF].

Conclusion and perspectives

All of these relationships between polyamines and the different aspects of root development, i.e. meristem activity, elongation, differentiation, lateral and adventitious root formation, may be extremely useful for biotechnological purposes. Various protocols for modifying polyamine levels thus appear to promote rooting in thin layer explants, leaf explants, microcuttings, and shoots [START_REF] Friedman | Polyamines and root formation in mung bean hypocotyl cuttings. I. Effect of exogenous compounds and changes in endogenous polyamines[END_REF][START_REF] Friedman | Polyamines and root formation in mung bean hypocotyl cuttings. II. Incorporation of precursors into polyamines[END_REF][START_REF] Jarvis | Involvement of polyamines with adventitious root development in stem cuttings of mung bean[END_REF][START_REF] Burtin | Polyamines, hydroxycinnamoylputrescines and root formation in leaf explants of tobacco cultivated in vitro -effects of the suicide inhibitors of putrescine synthesis[END_REF][START_REF] Altamura | De novo root formation in tobacco thin layers affected by inhibition of polyamine biosynthesis[END_REF][START_REF] Hausman | Involvement of putrescine in the inductive rooting phase of poplar shoots raised in vitro[END_REF][START_REF] Tonon | Changes in polyamines, auxins and peroxidase activity during in vitro rooting of Fraxinus angustifolia shoots: an auxin-independent rooting model[END_REF]. Such protocols may obviously be of great interest for applications to in vitro culture and micropropagation.

Roots are vital to plant growth and therefore crop productivity [START_REF] Lynch | Root architecture and plant productivity[END_REF]. Studies of the impact of polyamines on root architecture may suggest novel improvement strategies, through marker-assisted genetic breeding or genetic transformation, in order to obtain cultivars with larger root development. Polyamine metabolism and regulation are complex, and involve a great number of genes. However, it has been shown that single-gene genetic transformation was sufficient to obtain important modifications of polyamine balance [START_REF] Lepri | Over-expression of a cDNA for human ornithine decarboxylase in transgenic rice plants alters the polyamine pool in a tissue-specific manner[END_REF][START_REF] Bassie | Expression of heterologous Sadenosylmethionine decarboxylase cDNA in plants demonstrates that changes in S-adenosyl-L-methionine decarboxylase activity determine levels of the higher polyamines spermidine and spermine[END_REF]. Cultivars with wider or deeper root systems may be useful for cultivation on land with poor water or mineral resources [START_REF] Klepper | Root growth and water uptake[END_REF]. It may also be envisaged to utilise such cultivars for bioremediation of polluted land, if ever they are able not only to develop a wide root system under adverse conditions, but also to take up pollutants from surrounding soil. Polyamines have for instance been reported to enhance tolerance to xenobiotics such as herbicides [START_REF] Zheleva | Protective effects of exogenous polyamines against atrazine in pea plants[END_REF]. However, in the case of xenobiotics that use polyamine transport systems, as reported for paraquat in maize [START_REF] Hart | Transport interactions between paraquat and polyamines in roots of intact maize seedlings[END_REF], elevated polyamine levels may not be compatible with xenobiotic uptake.

The recent identification and characterisation of Arabidopsis mutants of polyamine biosynthesis pathways [START_REF] Watson | Arginine decarboxylase (polyamine synthesis) mutants of Arabidopsis thaliana exhibit altered root growth[END_REF][START_REF] Hanzawa | ACAULIS5, an Arabidopsis gene required for stem elongation, encodes a spermine synthase[END_REF] has confirmed the relationship between polyamine metabolism and developmental processes. However, further mutant screening and reverse genetics approaches will certainly yield in the near future more information on the involvement of genes related to polyamine synthesis and the roles of polyamines in root development. At present, only the two complementation groups of spe mutants [START_REF] Watson | Arginine decarboxylase (polyamine synthesis) mutants of Arabidopsis thaliana exhibit altered root growth[END_REF] provide a link between polyamines and root development. The existence of families of polyamine-related genes with tissue-specific expression and potentially different functions [START_REF] Galloway | Phylogenetic utility of the nuclear gene arginine decarboxylase: an example from Brassicaceae[END_REF][START_REF] Watson | Arginine decarboxylase (polyamine synthesis) mutants of Arabidopsis thaliana exhibit altered root growth[END_REF][START_REF] El Amrani | Genome-wide distribution and potential regulatory functions of AtATE, a novel family of miniature inverted-repeat transposable elements in Arabidopsis thaliana[END_REF][START_REF] Hanzawa | Characterisation of the spermidine synthase-related gene family in Arabidopsis thaliana[END_REF] raises the possibility that specific genes in these families are involved in root development. However, other aspects of polyamine metabolism, such as degradation and conjugation pathways, mechanisms of polyamine action, such as signal transduction pathways, and mechanisms of polyamine uptake remain poorly characterised in plants. Characterisation of these processes will be essential to explain the classical view of polyamine involvement in cell division and differentiation, or to elucidate the links that may exist between polyamines and regulation of genes controlling root development.

Figure 1 .

 1 Figure1. Pathways of polyamine metabolism and ethylene synthesis. Abbreviations: 1-aminocyclopropane-1-carboxylic acid (ACC), arginine decarboxylase (ADC), diamine oxidase (DAO), ornithine decarboxylase (ODC), polyamine oxidase (PAO), S-adenosylmethionine decarboxylase (SAMDC), spermidine synthase (SPDS), spermine synthase (SPMS).

Acknowledgement

The authors wish to thank Dr Josette Martin-Tanguy for helpful discussions.