Enzyme regulation in neuroblastoma cells in a salts/glucose medium: Induction of ornithine decarboxylase by asparagine and glutamine

Abstract

L-Asparagine is necessary and sufficient for the maximal induction of ornithine decarboxylase (ODC) (Lornithine carboxy-lyase, EC 4.1.1.17) activity in confluent N18 mouse neuroblastoma cells in a salts/glucose medium; L-asparagine also induces maximal ODC activity when added to a tissue culture medium. L-Glutamine is about one-half as effective as asparagine. Cholera toxin and agents that are known to raise intracellular cyclic AMP concentrations have no effect on the induction of ODC activity unless suboptimal concentrations of asparagine are present in the salts/glucose medium. Whereas actinomycin D does not inhibit induction of ODC activity by asparagine, it inhibits the induction of ODC activity in association with cyclic AMP. In the salts/glucose medium, the rate of loss of ODC activity following the inhibition of protein synthesis by cycloheximide or puromycin depends upon the presence or absence of asparagine; loss is rapid only in the absence of asparagine and does not appear to be related to the inhibition of protein synthesis. These results are discussed in the context that the overlay of the growth medium tends to mask the minimal requirements for enzyme induction, because the composition of the medium defines: (a) the requirements for the induction of ODC activity; (b) the effect, orlack of effect, of cyclic AMP (and of inducers of intracellular cyclic AMP) on the induction of ODC activity; (c) the effect, or lack of effect, of actinomycin D on the induction of ODC activity; and (d) the action of puromycin and of cycloheximide on the rate of loss of ODC activity. It will be interesting to determine whether these results are uniquely applicable to ODC, whether many of the reactions attributed to cyclic AMP in the literature may be mediated by asparagine and glutamine, and whether actinomycin D, cycloheximide, and puromycin can be relied upon to differentiate between transcriptional and post-transcriptional control.Most studies on enzyme induction, including our own, especially in relation to ornithine decarboxylase (ODC) (L-ornithine carboxy-lyase, EC 4.1.1.17), have relied heavily on the use of cell cultures in incubation media of varying complexity or on the use of whole animals (1-3). Such studies tend to provide answers that are modified either by the overall physiology of the cell as affected by the complexity of the medium or by the inherent complexity of the animal as modified by any additional conditions of imposed environmental and experimental stress. For example, when enzyme induction by hormones is being determined in a cell culture, if the hormones have differential positive and negative effects on the transport of various substances, the effectiveness or noneffectiveness of a hormone in inducing an enzyme activity will be completely dependent upon the composition of the medium.In order to minimize such extraneous influences, we induce ODC activity in confluent N18 mouse neuroblastoma cells in a minimal salts/glucose medium. Under these conditions, the addition of L-asparagine is...

Antizyme (AZ) regulates intestinal cell growth independent of polyamines

“…ODC activity itself is highly controlled, increasing dramatically from basal levels in response to tissue damage, growth factors, trophic hormones and other proliferative stimuli (Fausto 1971; Hovis et al 1986). Increases in ODC activity are relatively transient, for it has one of the shortest half-lives of any mammalian enzyme (Canellakis et al 1979; Chen et al 1977). The activity of the enzyme is primarily regulated by a negative feedback mechanism involving ODC antizyme (AZ) (Coffino 2001).…”

Amino Acids Regulate Expression of Antizyme-1 to Modulate Ornithine Decarboxylase Activity

“…Fausto (1) found that some amino acids could increase ODC activity in regenerating liver. Chen and Canellakis (13) and Chen and Chen (31) examined the ability of amino acids to induce ODC activity in cells incubated in a simple salt-glucose solution (Earle's balanced salt solution; EBSS), thus eliminating stimulation from growth factors present in serum or other amino acids and nutrients normally found in incubation medium. They showed that ODC activity was increased in neuroblastoma cells by asparagine (Asn) and glutamine.…”

Glucose elevates ornithine decarboxylase expression in vero cells

“…Enzyme levels remained elevated for an additional 8 h r and only began to return to basal levels between 20 and 24 hr. Although this pattern of ODC expression is atypical, a similar pattern of enzyme expression was shown to occur in neuroblastoma cells incubated in EBSS plus 10 mM asparagine (Chen and Canellakis, 1977).…”

“…Previous studies with neuroblastoma cells reported that the addition of group 'A' transported amino acids to EBSS was a necessary and sufficient requirement for ODC expression (Chen and Canellakis, 1977). Relative to basal ODC activity in untreated Vero cells, the amount of 14C02 generated from L-[1-I4C] ornithine was higher not only in homogenates from confluent cells incubated for 5 hr in EBSS plus asparagine but also in cells incubated with EBSS alone (Table 1).…”