Tetrahydrofolate (THF) is a central cofactor for one-carbon transfer reactions in all living organisms. In this study, we analyzed the expression of dihydropterin pyrophosphokinase-dihydropteroate synthase (HPPK-DHPS) in pea (Pisum sativum) organs during development, and so the capacity to synthesize dihydropteroate, an intermediate in the de novo THF biosynthetic pathway. During seedling development, all of the examined organs/tissues contain THF coenzymes, collectively termed folate, and express the HPPK-DHPS enzyme. This suggests that each organ/tissue is autonomous for the synthesis of THF. During germination, folate accumulates in cotyledons and embryos, but high amounts of HPPK-DHPS are only observed in embryos. During organ differentiation, folate is synthesized preferentially in highly dividing tissues and in photosynthetic leaves. This is associated with high levels of the HPPK-DHPS mRNA and protein, and a pool of folate 3-to 5-fold higher than in the rest of the plant. In germinating embryos and in meristematic tissues, the high capacity to synthesize and accumulate folate correlates with the general resumption of cell metabolism and the high requirement for nucleotide synthesis, major cellular processes involving folate coenzymes. The particular status of folate synthesis in leaves is related to light. Thus, when illuminated, etiolated leaves gradually accumulate the HPPK-DHPS enzyme and folate. This suggests that folate synthesis plays an important role in the transition from heterotrophic to photoautotrophic growth. Analysis of the intracellular distribution of folate in green and etiolated leaves indicates that the coenzymes accumulate mainly in the cytosol, where they can supply the high demand for methyl groups.The synthesis of numerous biological compounds and the regulation of many metabolic processes require the addition or removal of one-carbon units (C1 metabolism). Tetrahydrofolate (THF) coenzymes mediate these C1 transfer reactions that are involved in several major cellular processes, including the synthesis of purines and thymidylate, amino acid metabolism, pantothenate synthesis, mitochondrial and chloroplastic protein biogenesis, and Met synthesis (Fig. 1). Met is the direct precursor of S-adenosyl-Met (Ado-Met), which in turn is the source of methyl units for the synthesis of a myriad of molecules such as choline, chlorophyll, or lignin (for reviews, see Cossins, 2000;Scott et al., 2000;Hanson and Roje, 2001). In plants, THF is also involved in the photorespiratory cycle, a specific pathway that occurs at very high rates in green leaves from C3 plants. Photorespiration relies on two THF-dependent enzymes present in the matrix space of leaf mitochondria, the Gly decarboxylase complex (GDC) and Ser hydroxymethyltransferase (SHMT; for reviews, see Oliver, 1994;Douce et al., 2001).THF is composed of three distinct parts, namely a pterin ring, a p-aminobenzoic acid, and a poly-Glu chain of variable length (1-8 residues). Its function is to bind, transport, and donate C1 units that differ i...

methionine ␥-lyase ͉ NMR profiling ͉ plant T he sulfur-containing amino acid methionine (Met) is an essential metabolite in all living organisms (1). Besides its role as protein constituent, Met is the precursor of S-adenosylmethionine (AdoMet), which is the major methyl-group donor in transmethylation reactions and an intermediate in the biosynthesis of biotin, polyamines, and the phytohormone ethylene. Met is synthesized de novo by plants, fungi, and microbes. It is the only sulfur-containing amino acid that is essential for human and monogastric livestock. In these organisms, Met is metabolized through the reverse transsulfuration pathway where the ␥-cleavage of the cystathionine intermediate, a reaction that does not exist in plants, allows synthesis of cysteine (Cys) (2). Therefore, Met must be supplied from the diet and, because it is the most limiting essential amino acid in legume seeds, metabolic engineering strategies have been developed to increase the carbon flux into free Met as well as the incorporation of this amino acid into proteins (3).In higher plants, the neo-synthesized Met molecule originates from three convergent pathways with the sulfur atom deriving from Cys, the nitrogen͞carbon backbone from aspartate, and the methyl moiety from the ␤-carbon of serine via the pool of folates (1). De novo Met synthesis consists of three consecutive reactions localized in plastids (4) and catalyzed by cystathionine ␥-synthase, cystathionine ␤-lyase, and methionine synthase. Several recent studies have indicated that Met synthesis and accumulation are subject to complex regulatory controls in which cystathionine ␥-synthase plays a crucial role (5-7).Studies of metabolic fates of Met using the aquatic plant Lemna paucicostata indicated that the synthesis and turnover of AdoMet accounts for Ϸ80% of Met metabolism, whereas the synthesis of proteins drives Ϸ20% of Met (for a review, see ref. 8). More than 90% of AdoMet then is used for transmethylation, leading to nucleic acid, protein, lipid, and other metabolites modifications. The resulting homocysteinyl moiety is recycled back to Met and AdoMet by a set of cytosolic reactions designated as the activated methyl cycle. The use of AdoMet for the synthesis of polyamines and, in some plant tissues, ethylene also is accompanied by recycling of the methylthio moiety and regeneration of Met. The last known metabolic fate of Met is unique to plants and leads to the production of S-methylmethionine (SMM), a compound resulting from the AdoMet-dependent methylation of Met. SMM then can donate a methyl group to homocysteine (Hcy) through the reaction catalyzed by Hcy S-methyltransferase, yielding two molecules of Met. The SMM cycle might serve to achieve short-term control of AdoMet levels in plant cells, thus controlling the commitment of one-carbon units into methyl-group synthesis (9-11).Despite recent progress in elucidating the synthesis of Met and its complex regulation, little is known about the catabolism of this amino acid in plant cells. Several authors have ...