title: Increased translation efficiency and antizyme-dependent stabilization of ornithine decarboxylase in amino acid-supplemented human colon adenocarcinoma cells, Caco-2.
abstract: The mechanisms of the response of ornithine decarboxylase(ODC), the rate-limiting enzyme in polyamine biosynthesis, to amino acid supplementation were studied in the human colon adenocarcinoma cell line, Caco-2. Supplementation of serum-deprived, subconfluent Caco-2 cells with any one of a series of amino acids (10 mM) resultedin increased ODC activity, reaching a maximum of approx. 12.5-fold after approx. 4 h, over control cells either not supplemented or supplemented with iso-osmolar D-mannitol. Glycine, L-asparagine and L-serine, as well as their D-enantiomers, were the strongest effectors and acted in a concentration-dependent manner; millimolar concentrations of most of these amino acids being sufficient to significantly increase ODC activity. In contrast, supplementation with D-methionine, L-lysine, L-aspartate or L-glutamate had little or no effect on ODC activity, whereas supplemental L-methionine, L-arginine, L-ornithine or L-cysteine was inhibitory. Polyamine assays showed that the putrescine content of cells varied in accordance with the changes in ODC activity. Western-blot and Northern-blot analyses revealed specifically increased levels of ODC protein but not mRNA,respectively, in response to supplementation with an ODC-inducing amino acid. Suppression of the increase in cycloheximide-treated cellsconfirmed a requirement for protein synthesis. Pulse-labelling of cellswith [(35)S]methionine showed a 3-fold increase in thesynthesis of ODC protein after 4 h of supplementation with glycineor L-serine. Supplemental glycine also augmented, reversibly, the half-life of ODC by almost 4-fold and simultaneously decreased the activity of putrescine-induced free antizyme. These results suggest that translational, but not transcriptional, regulation of ODC takes part in ODC induction by amino acids in Caco-2 cells. However, it also appears to occur in concert with decreased enzyme in activation and/or degradation.
authors.affiliation: Unité de Nutrition Cellulaire et Moléculaire, Institut National de la Recherche Agronomique, 63122-Theix, France.
authors.forename: H
authors.initials: H
authors.lastname: Chabanon
authors.forename: L
authors.initials: L
authors.lastname: Persson
authors.forename: H M
authors.initials: HM
authors.lastname: Wallace
authors.forename: M
authors.initials: M
authors.lastname: Ferrara
authors.forename: P
authors.initials: P
authors.lastname: Brachet
mesh_terms: D000596:Amino Acids / Q000378:metabolism* / Q000494:pharmacology; D018938:Caco-2 Cells; D016895:Culture Media, Serum-Free; D004791:Enzyme Inhibitors / Q000378:metabolism*; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D006801:Humans; D007700:Kinetics; D008715:Methionine / Q000378:metabolism; D009955:Ornithine Decarboxylase / Q000096:biosynthesis / Q000235:genetics* / Q000378:metabolism; D014176:Protein Biosynthesis; D011506:Proteins / Q000378:metabolism*
chemical_list: D000596:Amino Acids; D016895:Culture Media, Serum-Free; D004791:Enzyme Inhibitors; D011506:Proteins; C031017:ornithine decarboxylase antizyme; D008715:Methionine; D009955:Ornithine Decarboxylase