In Arabidopsis thaliana, silencing of hydroxycinnamoyl-CoA shikimate/quinate hydroxycinnamoyl transferase (HCT), a lignin biosynthetic gene, results in a strong reduction of plant growth. We show that, in HCT-silenced plants, lignin synthesis repression leads to the redirection of the metabolic flux into flavonoids through chalcone synthase activity. Several flavonol glycosides and acylated anthocyanin were shown to accumulate in higher amounts in silenced plants. By contrast, sinapoylmalate levels were barely affected, suggesting that the synthesis of that phenylpropanoid compound might be HCTindependent. The growth phenotype of HCT-silenced plants was shown to be controlled by light and to depend on chalcone synthase expression. Histochemical analysis of silenced stem tissues demonstrated altered tracheary elements. The level of plant growth reduction of HCT-deficient plants was correlated with the inhibition of auxin transport. Suppression of flavonoid accumulation by chalcone synthase repression in HCT-deficient plants restored normal auxin transport and wildtype plant growth. By contrast, the lignin structure of the plants simultaneously repressed for HCT and chalcone synthase remained as severely altered as in HCT-silenced plants, with a large predominance of nonmethoxylated H units. These data demonstrate that the reduced size phenotype of HCT-silenced plants is not due to the alteration of lignin synthesis but to flavonoid accumulation.

Root hairs and rhizoids are cells with rooting functions in land plants. We describe two basic helix-loop-helix transcription factors that control root hair development in the sporophyte (2n) of the angiosperm Arabidopsis thaliana and rhizoid development in the gametophytes (n) of the bryophyte Physcomitrella patens. The phylogeny of land plants supports the hypothesis that early land plants were bryophyte-like and possessed a dominant gametophyte and later the sporophyte rose to dominance. If this hypothesis is correct, our data suggest that the increase in morphological complexity of the sporophyte body in the Paleozoic resulted at least in part from the recruitment of regulatory genes from gametophyte to sporophyte.

The hydroxyl group in the 3-position of the phenylpropanoid compounds is introduced at the level of coumarate shikimate/ quinate esters, whose synthesis implicates an acyltransferase activity. Specific antibodies raised against the recombinant tobacco (Nicotiana tabacum) acyltransferase revealed the accumulation of the enzyme in stem vascular tissues of tobacco, in accordance with a putative role in lignification. For functional analysis, the acyltransferase gene was silenced in Arabidopsis thaliana and N. benthamiana by RNA-mediated posttranscriptional gene silencing. In Arabidopsis, gene silencing resulted in a dwarf phenotype and changes in lignin composition as indicated by histochemical staining. An indepth study of silenced N. benthamiana plants by immunological, histochemical, and chemical methods revealed the impact of acyltransferase silencing on soluble phenylpropanoids and lignin content and composition. In particular, a decrease in syringyl units and an increase in p-hydroxyphenyl units were recorded. Enzyme immunolocalization by confocal microscopy showed a correlation between enzyme accumulation levels and lignin composition in vascular cells. These results demonstrate the function of the acyltransferase in phenylpropanoid biosynthesis.

A protein hydrolyzing hydroxycinnamoyl-CoA esters has been purified from tobacco stem extracts by a series of high pressure liquid chromatography steps. The determination of its N-terminal amino acid sequence allowed design of primers permitting the corresponding cDNA to be cloned by PCR. Sequence analysis revealed that the tobacco gene belongs to a plant acyltransferase gene family, the members of which have various functions. The tobacco cDNA was expressed in bacterial cells as a recombinant protein fused to glutathione S-transferase. The fusion protein was affinity-purified and cleaved to yield the recombinant enzyme for use in the study of catalytic properties. The enzyme catalyzed the synthesis of shikimate and quinate esters shown recently to be substrates of the cytochrome P450 3-hydroxylase involved in phenylpropanoid biosynthesis. The enzyme has been named hydroxycinnamoyl-CoA: shikimate/quinate hydroxycinnamoyltransferase. We show that p-coumaroyl-CoA and caffeoyl-CoA are the best acyl group donors and that the acyl group is transferred more efficiently to shikimate than to quinate. The enzyme also catalyzed the reverse reaction, i.e. the formation of caffeoyl-CoA from chlorogenate (5-O-caffeoyl quinate ester). Thus, hydroxycinnamoyl-CoA:shikimate/ quinate hydroxycinnamoyltransferase appears to control the biosynthesis and turnover of major plant phenolic compounds such as lignin and chlorogenic acid.

SUMMARYBy combining Zinnia elegans in vitro tracheary element genomics with reverse genetics in Arabidopsis, we have identified a new upstream component of secondary wall formation in xylary and interfascicular fibers. Walls are thin 1 (WAT1), an Arabidopsis thaliana homolog of Medicago truncatula NODULIN 21 (MtN21), encodes a plant-specific, predicted integral membrane protein, and is a member of the plant drug/metabolite exporter (P-DME) family (transporter classification number: TC 2.A.7.3). Although WAT1 is ubiquitously expressed throughout the plant, its expression is preferentially associated with vascular tissues, including developing xylem vessels and fibers. WAT1:GFP fusion protein analysis demonstrated that WAT1 is localized to the tonoplast. Analysis of wat1 mutants revealed two cell wall-related phenotypes in stems: a defect in cell elongation, resulting in a dwarfed habit and little to no secondary cell walls in fibers. Secondary walls of vessel elements were unaffected by the mutation. The secondary wall phenotype was supported by comparative transcriptomic and metabolomic analyses of wat1 and wild-type stems, as many transcripts and metabolites involved in secondary wall formation were reduced in abundance. Unexpectedly, these experiments also revealed a modification in tryptophan (Trp) and auxin metabolism that might contribute to the wat1 phenotype. Together, our data demonstrate an essential role for the WAT1 tonoplast protein in the control of secondary cell wall formation in fibers.

In their environment, plants interact with a multitude of living organisms and have to cope with a large variety of aggressions of biotic or abiotic origin. To survive, plants have acquired, during evolution, complex mechanisms to detect their aggressors and defend themselves. Receptors and signaling pathways that are involved in such interactions with the environment are just beginning to be uncovered. What has been known for several decades is the extraordinary variety of chemical compounds the plants are capable to synthesize, and many of these products are implicated in defense responses. The number of natural products occurring in plants may be estimated in the range of hundreds of thousands, but only a fraction have been fully characterized. Despite the great importance of these metabolites for plant and also for human health, our knowledge about their biosynthetic pathways and functions is still fragmentary. Recent progress has been made particularly for phenylpropanoid and oxylipin metabolism, which are emphasized in this review. Both pathways are involved in plant resistance at several levels: by providing building units of physical barriers against pathogen invasion, by synthesizing an array of antibiotic compounds, and by producing signals implicated in the mounting of plant resistance.

Arabidopsis wat1 (walls are thin1)-mediated resistance to the bacterial vascular pathogen, Ralstonia solanacearum, is accompanied by cross-regulation of salicylic acid and tryptophan metabolism SUMMARYInactivation of Arabidopsis WAT1 (Walls Are Thin1), a gene required for secondary cell-wall deposition, conferred broad-spectrum resistance to vascular pathogens, including the bacteria Ralstonia solanacearum and Xanthomonas campestris pv. campestris, and the fungi Verticillium dahliae and Verticillium albo-atrum. Introduction of NahG, the bacterial salicylic acid (SA)-degrading salicylate hydroxylase gene, into the wat1 mutant restored full susceptibility to both R. solanacearum and X. campestris pv. campestris. Moreover, SA content was constitutively higher in wat1 roots, further supporting a role for SA in wat1-mediated resistance to vascular pathogens. By combining transcriptomic and metabolomic data, we demonstrated a general repression of indole metabolism in wat1-1 roots as shown by constitutive down-regulation of several genes encoding proteins of the indole glucosinolate biosynthetic pathway and reduced amounts of tryptophan (Trp), indole-3-acetic acid and neoglucobrassicin, the major form of indole glucosinolate in roots. Furthermore, the susceptibility of the wat1 mutant to R. solanacearum was partially restored when crossed with either the trp5 mutant, an over-accumulator of Trp, or Pro35S:AFB1-myc, in which indole-3-acetic acid signaling is constitutively activated. Our original hypothesis placed cell-wall modifications at the heart of the wat1 resistance phenotype. However, the results presented here suggest a mechanism involving rootlocalized metabolic channeling away from indole metabolites to SA as a central feature of wat1 resistance to R. solanacearum.

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