Mechanism of regulation of ornithine decarboxylase gene expression by asparagine in a variant mouse neuroblastoma cell line.

Z.P Chen,K.Y Chen

doi:10.1016/s0021-9258(19)50520-8

Z.P Chen, K.Y Chen

Open Access

https://doi.org/10.1016/s0021-9258(19)50520-8

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Abstract

We have developed a clonal variant, named DF-40, from the N2a mouse neuroblastoma cell line, which has the ornithine decarboxylase (L-ornithine carboxylase, EC 4.1.1.17, ODC) gene amplified. When DF-40 cells were maintained in a simple salt glucose medium (e.g. Earle's blanced salt solution), L-asparagine alone was sufficient to induce a maximal increase in ODC activity. The increase in ODC activity correlated well with an increase in the amount of ODC protein. Northern blot analysis indicated that asparagine caused a 12-15-fold increase in ODC mRNA. The half-life of ODC mRNA induced by asparagine in DF-40 cells changed from more than 8 h to about 25 min upon removal of asparagine from the culture in the presence of actinomycin D. In contrast, asparagine had little or no effect on the rate of transcription of the ODC gene. Pulse labeling of cells for 15 min with [35S]methionine showed a 90-140-fold increase in the synthesis of ODC protein after 4-8 h of incubation with asparagine. The removal of asparagine from the medium resulted in a rapid loss of ODC protein with a half-life as short as 12 min. The presence of asparagine increased the half-life of ODC protein by 3-5-fold when measured in the presence of cycloheximide. Taken together, our data show that asparagine induced ODC gene expression in DF-40 cells, primarily by post-transcriptional stabilization of ODC mRNA. In addition, asparagine specifically stimulated the synthesis and suppressed the degradation of ODC protein.

Highlights

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Wehave previously shown that a single amino acid, asparagine, is necessary and sufficient to induce maximal Ornithine decarboxylase (ODC) activity in various tumor cells maintained in a salts/glucose solution (Chen and Canellakis, 1977; Viceps-Madore et al, 1982; Chen and Liu, 1983)
We and others showed that in primary cultures such as hepatocytes or differentiated mouse neuroblastoma cells maintained in the salts/glucose solution, the maximal induction of ODC activity requires the presence of both asparagine and serum, or growth hormones (Chen, 1980; Kanamoto et al, 1987)

Summary

Zong Ping Chen and KuangYu ChenS

The are all ineffective in inducing ODC activity when cells are removal of asparagine from themedium resulted in a maintained in the salts/glucose solution. These rapid loss of ODC protein with a half-life as short as agents can potentiatethe effect of suboptimal concentrations. In view of the unique effect of asparagine including growth stimuli (e.g. Kontula et al (1984), Kahana in inducing ODC activity and thepotential role of asparagine and Nathans (1984), Persson et al (1985), and Hovis et al in regulating ODC under physiological conditions, we have (1986)).It has been shown that ODC activity can be controlled examined the mechanism of the induction of ODC activity by at transcriptional(Kontula et al, 1984; Sertichand Pegg, asparagine in this DF-40 cell line. Were grown in Dulbecco’s modified Eagle’s medium(with 4500 mg of glucose per liter) supplementedwith 10%fetal bovine serum at 37 "C

RESULTS

Polyamreinceosvered in cellular polyamines

EEBBSS S

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Findings

DISCUSSION