Studies on the regulation of ornithine decarboxylase in yeast: Effect of deletion in the MEU1 gene

Abstract

Methylthioadenosine is formed during the biosynthesis of spermidine and of spermine and is metabolized by methylthioadenosine phosphorylase, an enzyme missing in several tumor cell lines. In Saccharomyces cerevisiae, this enzyme is coded by the MEU1 gene. We have now studied the effect of the meu1 deletion on polyamine metabolism in yeast. We found that the effects of the meu1⌬ mutation mostly depend on the stage of cell growth. As the cell density increases, there is a marked fall in the level of ornithine decarboxylase (ODC) in the MEU1 ؉ cells, which we show is caused by an antizyme-requiring degradation system. In contrast, there is only a small decrease in the ODC level in the meu1⌬ cells. The meu1⌬ cells have a higher putrescine and a lower spermidine level than MEU1 ؉ cells, suggesting that the decreased spermidine level in the meu1⌬ cultures is responsible for the greater apparent stability of ODC in the meu1⌬ cells. The lower spermidine level in the meu1⌬ cells probably results from an inhibition of spermidine synthase by the methylthioadenosine that presumably accumulates in these mutants. In both MEU1 ؉ and the meu1⌬ cultures, the ODC levels were markedly decreased by the addition of spermidine to the media, and thus our results contradict the postulation of Subhi et al. methylthioadenosine ͉ methylthioadenosine phosphorylase ͉ polyamines ͉ spermidine ͉ putrescine O rnithine decarboxylase (ODC) is the key regulatory enzyme of the polyamine biosynthetic pathway and catalyzes the decarboxylation of ornithine to 1,4-diaminobutane (putrescine). Putrescine is then converted to spermidine and spermine by the addition of aminopropyl groups from decarboxylated Sadenosylmethionine (1). Methylthioadenosine is formed as a byproduct of these aminopropyl transferase reactions (spermidine synthase and spermine synthase) (2-4) and is then converted to adenine and to methionine by the ''methionine salvage'' pathway (5-9). The first step in the methionine salvage pathway is the phosphorolysis of methylthioadenosine to methylthioribose-1-phosphate and adenine by methylthioadenosine phosphorylase (EC 2.4.2.28). The metabolism of methylthioadenosine in mammalian cells has been of special interest because methylthioadenosine phosphorylase is absent in many tumors (10)(11)(12)(13)(14)(15). Thomas et al. (16) and Subhi et al. (17) have shown that, in Saccharomyces cerevisiae, methylthioadenosine phosphorylase is coded by the MEU1 gene and have described meu1⌬ mutants. The availability of these yeast mutants deleted in the methylthioadenosine phosphorylase gene permits more extensive studies on the role of this gene and the methionine salvage pathway in polyamine metabolism in yeast, and a comparison of these effects in yeast with those reported in mammalian cells. Therefore, as an extension of our work on the regulation of ornithine decarboxylase and of polyamine pools in yeast (18), we have now studied the effect of the meu1⌬ deletion on these parameters. Our findings were different from those reported by Subhi et al.

Metabolic characteristics and importance of the universal methionine salvage pathway recycling methionine from 5′‐methylthioadenosine

“…In mutants accumulating MTA, on the other hand, the levels will be sufficiently high that an effect on polyamine synthesis is detectable, for yeast at 0.03-3.7 mM (15,30,81). In yeast mutants lacking the phosphorylase, ornithine decarboxylase levels are increased, which is caused by at least partly an enlarged stability of the enzyme (19,81). The last metabolite, MTOB, is also involved in the regulation of ornithine decarboxylase.…”

“…This relationship may be expected thinking of that the starting point of polyamine metabolism, ornithine, is an intermediate in arginine biosynthesis. Furthermore, it has been found that MTA inhibits the polyamine synthetic enzymes, spermidine synthase, and spermine synthase, whereas MTA increases the activities of ornithine and S-adenosylmethionine decarboxylases (20,(81)(82)(83). The concentration of MTA to inhibit the two synthases by 50% was 10-30 lM.…”

“…However, in wild-type cells of at least yeast it seems that MTA plays a limited role because of a quick conversion in the MTA cycle resulting in very low intracellular levels (15). In mutants accumulating MTA, on the other hand, the levels will be sufficiently high that an effect on polyamine synthesis is detectable, for yeast at 0.03-3.7 mM (15,30,81). In yeast mutants lacking the phosphorylase, ornithine decarboxylase levels are increased, which is caused by at least partly an enlarged stability of the enzyme (19,81).…”

Functional Identification of APIP as Human mtnB, a Key Enzyme in the Methionine Salvage Pathway

“…The methionine salvage pathway and the polyamine synthesis seem to be tightly coupled, probably in order to maintain intracellular levels of SAM. For example, it has been shown that the level and activity of ornithine decarboxylase, the rate-controlling enzyme in polyamine synthesis, can be modulated by the first and last metabolites of the methionine salvage pathway: MTA and 4-methhylthio-2-oxo-butanoate (MTOB) [5]–[7]. The methionine salvage pathway may also have an important role in apoptotic processes as both MTA and MTOB were found to induce apoptosis [5], [8].…”

Yeast ornithine decarboxylase and antizyme form a 1:1 complex in vitro: Purification and characterization of the inhibitory complex

“…Ornithine decarboxylase was assayed as described earlier (20). Antizyme activity was measured on the basis of inhibition of ornithine decarboxylase in the same assay mixture as ODC.…”