Josette Martin-Tanguy

The occurrence and possible function of hydroxycinnamoyl acid amides in plants

In recent years polyamines have been shown to participate in various aspects of plant growth processes [1, 2, 4, 5, 14, 28, 67, 68, 69, 71]. In general, free putrescine, spermidine and spermine were the only amines measured, and only little attention has been given to bound or conjugated polyamines. Although the sporadic identification of the conjugates formed between amines and cinnamic acids in various plants has been reported over many years, it is only recently that the widespread nature and potential significance of these amides has been recognised.

The distribution of hydroxycinnamic acid amides in flowering plants

Abstract Hydroxycinnamic acid amides have been identified as the main phenolic constituents in the reproductive organs of a range of flowering plants.

Hydroxycinnamic acid amides in fertile and cytoplasmic male sterile lines of maize

Abstract Hydroxycinnamic acid (HCA) amides in fertile and cytoplasmic male sterile lines of maize were determined in reproductive organs, developing grains and cobs. HCA amides occurred in large amounts in the anthers of fertile plants (line F7N) and were absent from the anthers of cytoplasmic male sterile lines (lines F7T and F7C). Restoration of fertility was associated with the production of these compounds (line FC31). Considerable variations were observed in the concentrations of HCA amides at different stages of growth and grain maturation. Changes of HCA amides in the grains which were to produce sterile plants followed a pattern similar to that obtained with the grains which were to produce fertile plants. Accumulation of HCA amides was substantially higher in fertile lines whatever their genotype (F7N, FC31 and F7T x FC31) than in sterile lines. Marked changes occurred in the HCA amide content of embryo and endosperm during grain development. Many changes in HCA amides were observed in cobs during development and maturation, but no substantial differences could be observed between fertile and sterile lines.

Polyamines, floral induction and floral development of strawberry (Fragaria ananassa Duch.)

In the short-day plant, strawberry (Fragaria ananassa Duch.), polyamines (putrescine, spermidine and spermine), conjugated spermidine (water-insoluble compounds) and bound amines (putrescine, spermidine, phenylethylamine, 3-hydroxy, 4-methoxyphenylethylamine) accumulated in the shoot tips during floral induction and before floral emergence. Different associations of free amines and conjugated amines were observed during floral induction, as compared with the reproductive phase. During the whole period of floral development, phenylethylamine (an aromatic amine) was the predominant amine, representing 80 to 90% of the total free amine pool. Phenylethylamine conjugates (water-insoluble compounds) were the predominant amides observed prior to fertilization. These substances decreased drastically after fertilization. In vegetative shoot tips from plants grown continously under long days, free polyamines (putrescine, spermidine) and bound polyamines (putrescine, spermidine) were low and no change was observed. Free amines (spermine and phenylethylamine), bound aromatic amines (phenylethylamine, 3-hydroxy, 4-methoxyphenylethylamine), conjugated spermidine and phenylethylamine did not appear. Male-sterile flowers were distinguished by their lack of conjugated spermidine and phenylethyalamine and by a decrease in free phenylethylamine. In normal and sterile strawberry plants α-DL-difluoromethylornithine (DFMO), a specific irreversible inhibitor of ornithine decarboxylase (ODC), caused inhibition of flowering and free and polyamine conjugates. When putrescine was added, polyamine titers and flowering were restored. A similar treatment with α-DL-difluoromethylarginine (DFMA), a specific, irreversible inhibitor of arginine decarboxylase (ADC), did not affect flowering and polyamine titers. These results suggest that ornithine decarboxylase (ODC) and polyamines are involved in regulating floral initiation in strawberry. The relationship between polyamines, aromatic amines, conjugates, floral initiation and male sterility is discussed.

Hydroxycinnamic acid amides (HCA) in zea mays: Distribution and changes with cytoplasmic male sterility

Earlier findings, obtained in our laboratory, indicated the presence of large amounts of hydroxycinnamic acid amides (HCA) in a number of species of flowering plants, representing 13 different families [l]. HCAare hydroxycinnamic acid amine conjugates. The link between these molecules is an amide bound. They occur as basic (water-soluble) or as neutral (water-insoluble) forms. In the basic forms, only one amine group of an aliphatic amine (diamine or polyamine) is linked with a phenolic acid. Neutral forms can be divided into two classes. In class 1, each terminal amine group of an aliphatic amine is neutralized by a phenolic acid. In class 2, the amine group of an aromatic amine is linked with a cinnamic acid. Some 24 hydroxycinnamic acid derivatives of the 5 amines: putrescine, spermidine, spermine, cadaverine and tyramine have been characterized [ 1 ]. Thus, HCA appear to be common constituents of flowering plants. Within the plant, these compounds were found to occur only in the reproductive organs, where they appear to be the main phenolic compounds. They are apparently absent from the green parts (leaves, stems) petals and sepals [ 11. For example, in Nicotiuna tabacum, we have reported the presence of caffeylputrescine, caffeylspermidine in the meristems [2] and demonstrated that there is an increase in the amount of these HCA in the apical part of tobacco plants at the time of floral induction [2,3]. Accumulation of HCA can also be induced in lower leaves by the topping of flowering plants [3]; This indicates that their production may be correlated with floral induction. In Nicotiuna tabacum cv. Xanthi n.c., an increase in temperature which

Polyamine metabolism and in vitro cell multiplication and differentiation in leaf explants of Chrysanthemum morifolium Ramat

Arginine decarboxylase (ADC), ornithine decarboxylase (ODC), diamine oxydase (DAO) free amine and conjugated amine titers were estimated in leaf explants of Chrysanthemum morifolium Ramat. var. Spinder cultivated in vitro in relation to hormone treatment. Addition of benzyladenine (BA) to a basal medium caused the formation of buds on the explants. BA plus 2,4 dichlorophenoxyacetic acid (2,4 D) caused callus formation and proliferation. Formation of roots was obtained by addition of indolylacetic acid (IAA). Arginine decarboxylase (ADC) ornithine decarboxylase (ODC) and diamine oxidase (DAO) activities increased during the first days of culture when cell multiplication was rapid, followed by a sharp decline as the rate of cell division decreased and differentiation took place. DAO activities increased rapidly in proliferating and growing organs and decreased during maturity. This increase was concomitant with ADC and ODC activities and polyamine content (free and conjugated polyamines). The biosynthesis and oxidation of polyamines which occurred simultaneously in physiological states of intense metabolism such as cell division or organ formation were directly correlated. In callus cultures DAO activity was blocked throughout development and regulated neither the cellular levels of polyamines nor polyamine conjugates. Levels of polyamine conjugates were high in callus cultures throughout development. In foliar explants cultivated on a medium promoting callus, inhibition of ODC activity by DFMO (α-DL-difluoromethylornithine, a specific enzyme-activated ODC inhibitor) resulting in an amide deficiency facilated the expression of differentiated cell function; substantial activation of DAO was observed until the emergence of the buds. On a medium promoting bud formation, β-OH ethylhydrazine (DAO inhibitor) promoted callus formation without differentiation. In this system DAO activity was blocked and there were high levels of polyamines, especially polyamine conjugates, throughout the culture period. The relationship among free and conjugated polyamines related biosynthetic enzyme activities, DAO activities, cell division and organ formation is discussed.

Polyamines and related enzymes in rice seeds differing in germination potential

In ungerminated rice seeds, (Japonica rice variety, CV Tapei 309), the content of free amines (putrescine, spermidine, spermine, tyramine) was higher in seed lots having a low germination frequency compared to those with high germination potential. Conversely, amine conjugates (di-feruloylputrescine, di-feruloylspermidine, diferuloyldiaminopropane and feruloyltyramine) decreased with loss of viability. Thus, these compounds appeared to constitute biochemical markers of seed viability. In seeds with high germination potential, conjugates decreased drastically during germination, with an early and rapid increase in free amines (putrescine, spermidine, tyramine). Arginine decarboxylase (ADC) activity was highest during the germination of high germination potential seeds, its activity gradually declining with loss of viability and being closely correlated with agmatine content. The polyamine biosynthetic inhibitors (α-DL-difluoromethylarginine, DFMA, a specific and irreversible inhibitor of ADC; α-DL-difluoromethylornithine, DFMO, a specific irreversible inhibitor of ornithine decarboxylase (ODC); cyclohexylammonium sulfate, CHA, inhibitor of spermidine synthase) neither depleted putrescine and spermidine levels nor inhibited germination in high germination potential seeds. In low germination potential seeds, the germination process was inhibited by DFMA or CHA. Application of agmatine resulted in a reversal of inhibition. DFMA inhibited ADC activity in both categories of seeds. In low germination potential seeds treated with CHA no ADC activity was found. These results suggest that amines are involved in the germination process of rice seeds. It appears that amine conjugates may serve as a storage form of amines which, upon enzymatic hydrolysis, could supply the cell with an additional amine reserve and influence cell division and/or cell elongation.

Genetic transformation with a derivative of rolC from Agrobacterium rhizogenes and treatment with α-aminoisobutyric acid produce similar phenotypes and reduce ethylene production and the accumulation of water-insoluble polyamine-hydroxycinnamic acid conjugates in tobacco flowers

Abstract Tobacco plants were either transformed by rolA (ORF 10) or rolC (ORF 12) from the Ri TL-DNA of Agrobacterium rhizogenes , or they were treated with an inhibitor of ethylene production, α-AIB (α-aminoisobutyric acid). We recorded changes in phenotype, ethylene production and the accumulation of di- and polyamines, tyramine and their derivatives in excised flowers, as a function of floral development. Both α-AIB and the rolC gene (under the control of the 35S promoter from the CaMV virus) caused male sterility and a reduction in flower size, accompanied by a depression in the accumulation of water-insoluble polyamine and tyramine conjugates and a reduction in ethylene production. In plants transformed by 35S- rolC , we also observed narrowed leaves, reduced internode distance and changes in root system architecture. Transformation by 35S- rolA caused leaf wrinkling, internode shortening and downward bending of stamen filaments. We discuss these observations in the light of proposed functions for rolA and rolC and known relationships between polyamine and ethylene metabolism.

Changing root system architecture through inhibition of putrescine and feruloyl putrescine accumulation

Plant roots provide anchorage and absorb the water and minerals necessary for photosynthesis in the aerial parts of the plant. Since plants are sessile organisms, their root systems must forage for resources in heterogeneous soils through differential branching and elongation [(1988) Funct. Ecol. 2, 345‐351; (1991) Plant Roots: The Hidden Half, pp. 3‐25, Marcel Dekker, NY]. Adaptation to drought, for instance, can be facilitated by increased root growth and penetration. Root systems thus develop as a function of environmental variables and the needs of the plant [(1988) Funct. Ecol. 2, 345‐351; (1986) Bot. Gaz. 147, 137‐147; (1991) Plant Roots: The Hidden Half, pp. 309‐330, Marcel Dekker, NY], We show, in a model system consisting of excised tobacco roots, that both α‐dl‐difluoromethylornithine (an inhibitor of putrescine biosynthesis) and the rolA gene (from the root‐inducing transferred DNA of Agrobacterium rhizogenes) stimulate overall root growth and cause a conversion in the pattern of root system formation, producing a dominant or ‘tap’ root. These morphological changes are correlated with a depression in the accumulation of polyamines and their conjugates.

Effects of inhibitors of polyamine biosynthesis and of polyamines on strawberry microcutting growth and development

The primary free polyamines identified during growth and development of strawberry (Fragaria × ananassa Duch.) microcuttings cultivated in vitro were putrescine, spermidine and spermine. Polyamine composition differed according to tissue and stages of development; putrescine was predominant in aerial green tissues and roots. α-DL-difluoromethylarginine (DFMA), a specific and irreversible inhibitor of the putrescine-synthesizing enzyme, arginine decarboxylase (ADC), strongly inhibited growth and development. Application of agmatine or putrescine to the inhibited system resulted in a reversal of inhibition, indicating that polyamines are involved in regulating the growth and development of strawberry microcuttings. α-DL-difluoromethylornithine (DFMO), a specific and irreversible inhibitor of putrescine biosynthesis by ornithine decarboxylase, promoted growth and development. We propose that ADC regulates putrescine biosynthesis during microcutting development. The application of exogenous polyamines (agmatine, putrescine, spermidine) stimulated development and growth of microcuttings, suggesting that the endogenous concentrations of these polyamines can be growth limiting.