Apparent post-transcriptional modification of ornithine decarboxylase accounts for its induction in IEC-6 cells in culture

Putrescine, the product of the ornithine decarboxylase (ODC)-catalyzed reaction, stimulates macromolecular synthesis in a duodenal crypt cell line. IEC-6 cells, grown in culture. In addition, supplementation of medium with putrescine alone reverses the inhibition of proliferation produced by inhibition of ODC with difluoromethylornithine (DFMO). A series of experiments was initiated in the IEC-6 cell line to study the regulation of induction of ODC, as this enzyme is rate-limiting in putrescine synthesis. Five percent fetal bovine serum (FBS) and 10 nM IGF-II stimulated a 21-fold and 6-fold induction of ODC activity, respectively. Kinetic analysis indicated that the effect was on the V_max of the reaction and not on the K_m, suggesting an increase in total ODC protein. This was verified by measuring [³H]DFMO binding; serum-stimulated induction of activity was accompanied by a corresponding 20-fold increase in the specific binding of DFMO to ODC. In contrast. Northern analysis demonstrated only a two-fold change in ODC mRNA level during induction. Measurement of enzyme stability showed that the half-life of the ODC protein was increased three-fold above basal level in the induced state. Inhibition of induction was produced by pretreatment with either the calmodulin antagonist, W-7, or the product of the ODC-catalyzed reaction, putrescine. Further analysis illustrated that the inhibition produced by these agents was partly the result of destabilization of the enzyme and not a decrease in message level. These results demonstrate that the induction of ODC by trophic agents is the result of post-transcriptional events rather than at the level of RNA synthesis.