In Vivo Metabolism of 5′-Methylthioadenosine in Lemna

Self Cite

Administration of methionine to growing Lemna had essentially no effect on accumulation of sulfate sulfur in protein cysteine, but decreased accumulation into cystathionine and its products (homocysteine, methionine, S-methylmethioninesulfonium salt, S-adenosylmethionine, and Sadenosylhomocysteine)to as low as 21% that of control plants, suggesting that methionine regulates its own de novo synthesis at cystathionine synthesis. Methionine caused only a slight reduction (to 80% that of control plants) in the accumulation of sucrose carbon into the 4-carbon moieties of cystathionine and products. This observation was puzzling since cystathionine synthesis proceeds by incorporation of equivalent amounts of sulfur (from cysteine) and 4-carbon moieties (from O-phosphohomoserine). The apparent inconsistency was resolved by the demonstration in Lemma (Giovanelli, Datko, Mudd, Thompson 1983 Plant Physiol 71: 319-326) that de novo synthesis of the methionine 4-carbon moiety occurs not only via the established transsulfuration route from 0-phosphohomoserine, but also via the ribose moiety of 5'-methylthioadenosine. It is now clear that the more accurate assessment of the flux of sulfur (and 4-carbon moieties) through transsulfuration is provided by the amount of IS from 35S042-that accumulates in cystathionine and its products, rather than by the corresponding measurements with 14C. These studies therefore unequivocally demonstrate in higher plants that methionine does indeed feedback regulate it own de novo synthesis in vivo, and that cystathionine synthesis is a locus for this regulation.Plants play an indispensable role in producing the ultimate source of most methionine required in the diet of humans and other nonruminant animals (15). In spite of this crucial role, the basic questions ofwhether methionine regulates its own de novo2 synthesis in vivo in plants, and if so, the identity ofthe regulatory loci, remain unanswered (15, 23 (1 1) showed that exogenous methionine decreased the accumulation of`4C from ['4C]glucose into protein methionine to 20% that ofcontrol cells, suggesting that methionine regulates its own de novo synthesis in these cells. However, this interpretation was questioned by Davies (10) on the basis that '4C 'trapped' in soluble methionine was not determined. Neither was account taken in these studies of excretion of '4C-compounds into the medium (27), or the possibility that synthesis of the methyl and 4-carbon moieties of methionine might be regulated independently. Bright et al. (2) observed that exogenous methionine did not affect incorporation of radioactivity from [14C]acetate into soluble plus protein methionine in cultured wheat embryos. These results suggest that methionine biosynthesis is insensitive to feedback control in this issue, but the authors themselves pointed out that other interpretations of the data are possible, since their measurements of combined radioactivity in the methyl and 4-carbon groups of methionine need not provide a valid assay of net synthesis of meth...

Section: Methodssupporting

confidence: 54%

“…Two teine--cystathionine--homocysteine--*methionine (15). The recently discovered synthesis of methionine from 5'-methylthioadenosine (17) results in a net synthesis of only the 4-carbon moiety of methionine, while the methyl and sulfur groups of methionine are independently recycled ( 16).…”

mentioning

confidence: 99%

In Vivo Regulation of De Novo Methionine Biosynthesis in a Higher Plant (Lemna)

Giovanelli¹,

Mudd²,

Datko³

1985

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“…Perhaps the largest source of adenine is generated from the catabolism of MTA which is produced during the biosynthesis of polyamines (22), ethylene (1,2) and methionine (9,10). The observation that polyamine levels first increase during meiosis in the anther tissue of Nicotiana and Datura is consistent with an increase in MTA metabolism at the time pollen development aborts in the APRT-deficient mutants (24,28).…”

Section: Uptake and Metabolism Of Adenine Ba And Bar In Mutant Planmentioning

confidence: 74%

Metabolism of Benzyladenine is Impaired in a Mutant of Arabidopsis thaliana Lacking Adenine Phosphoribosyltransferase Activity

Moffatt¹,

Pethe²,

Laloue³

1991

Formation of the riboside-5'-monophosphate is a general feature of the metabolism of cytokinins in plants. As part of a study of the biological significance of the nucleotide form of cytokinins, we analyzed a mutant of Arabidopsis thaliana deficient in adenine phosphoribosyltransferase (APRT) activity for its ability to metabolize N6-benzyladenine (BA). Formation of Nl-benzyladenosine-5'-monophosphate (BAMP) was assayed in vivo, by feeding tritiated BA to wild-type and mutant plantlets, and in crude plantlet extracts. Metabolites were separated by high performance liquid chromatography and quantitated by on-line liquid scintillation spectrometry. BA was rapidly absorbed by A. thaliana plantlets and pnmanly converted to BAMP and to BA 7-and 9-glucosides. BA was also rapidly absorbed by APRT-deficient plantlets, but its conversion to BAMP was strongly reduced. Formation of BAMP from N6-benzyladenosine was not affected in the mutant plantlets.In vitro conversion of BA to its nucleoside-5'-monophosphate was detected in crude extracts of wild-type plantlets, but not in extracts of APRT-deficient plantlets. Therefore, results of both assays indicate that APRT-deficient tissue does not convert BA to BAMP to a significant extent. Further, nondenaturing isoelectric focusing analysis of APRT activity in leaf extracts indicated that the enzyme activities which metabolize adenine and BA into their corresponding riboside-5'-monophosphate in extracts of wildtype plantlets have the same apparent isoelectric point. These activities were not detected in extracts prepared from APRTdeficient plantlets. Thus, these results demonstrate that APRT is the main enzyme which converts BA to its nucleotide form in young A. thaliana plants and that the ribophosphorylation of BA is not a prerequisite of its absorption by the plantlets. lism, but few of the enzyme activities effecting these steps in vivo have been identified. Furthermore, which modified form(s) constitutes the "active" cytokinin remains an open question. Elucidation of the enzymes involved in the synthesis and metabolism of cytokinins may provide valuable insight into the function of the various cytokinin metabolites, how their level in particular tissues is regulated and how these levels relate to physiology of plants.The interconversion of cytokinin bases, ribosides and nucleotides is a major feature of the metabolism of cytokinins (5,17,20,26), and it may be catalyzed by the enzyme system that metabolizes adenine. Several of these enzymes have been purified and shown to act on N-substituted substrates in vitro, albeit at significantly lower rates than those measured on the corresponding adenine substrates (3-6). Results of in vivo feeding experiments also support the notion that the purine metabolizing enzymes may be the main route by which cytokinins are interconverted (7).

“…Quantitatively less important are adenine and formate, formed during methylthio recycling. We have previously shown that adenine is also efficiently converted to adenine nucleotides (15). Conversion of adenine and its nucleoside probably proceeds by established salvage reactions (17).…”

Section: Discussionmentioning

confidence: 99%

“…The nature of the end products formed from the remaining flux through transmethylation is currently under investigation. Preliminary studies with radioactive methionine suggest that a substantial portion of this flux is accounted for by the and the aminopropyl moiety of spermidine; the 4-carbon moiety of methionine is regenerated from 4 of the ribose carbons of ATP (15,19). Some plant tissues are unusual in metabolizing methionine predominantly to ethylene (1,3), and in these tissues ethylene biosynthesis is the major source of methylthio recycling (19).…”

Section: Discussionmentioning

confidence: 99%

Quantitative Analysis of Pathways of Methionine Metabolism and Their Regulation in Lemna

Giovanelli

Mudd²,

Datko³

1985

Self Cite

118

Individual rates of metabolism of the sulfur, methyl, and 4-carbon moieties of methionine were estimated in Lemna paucicostata Hegelm. 6746 The known major pathways for metabolism of methionine in the great majority of plant tissues not evolving large amounts of ethylene are summarized in Figure 1 (reactions a through k). Studies focused on the pathway for net methionine formation (Fig. 1, ---) have clearly demonstrated that this process occurs via transsulfuration ( 12), is subject to feedback control at cystathionine y-synthase (Fig. 1, reaction b) (14, 18), and that methionine accumulates predominantly in protein (12). By contrast, the quantitative significance and regulatory patterns of the other pathways of methionine metabolism illustrated in Figure 1