Amino acid availability regulates polyamine transport in mammalian cells

Abnormal amino acid metabolism characterised by differential alterations in tissue levels of amino acid has been reported in numerous diseases. Supplementation with amino acid is thereby used for treatment and prevention of certain diseases such as atherosclerosis, as well as for improvement of the nutritional status, e.g. in patients critically ill, undergoing bone-marrow transplantation, or resection for colorectal cancer. Additionally, the effects of dietary amino acid restriction have been examined more particularly with the purpose of amino acid imbalance therapy for cancer. Amino acid restriction arrests cell proliferation and decreases rates of protein synthesis in eukaryotic cells. Furthermore, decreased tumorigenesis in arginine-restricted mice might be related to alterations in polyamine homeostasis. Polyamines (putrescine, spermidine and spermine) play an essential role in cell growth. Their intracellular concentrations are tightly regulated through membrane transport and metabolism pathways. Our aim was to characterise the effects of amino acid availability on spermidine transport in cultured human tumour cells. Exposure of colon adenocarcinoma (Caco-2) cells to a medium deprived of L-methionine or L-arginine caused a marked increase in the initial rate of specific transport of spermidine, unlike the activity of the polyamine biosynthesis enzyme ornithine decarboxylase and regardless of changes in the cellular content of polyamines. The increase was measurable as early as 1 h and was maximal 4-6 h after amino acid removal, declining thereafter but still remaining significantly elevated at 24 h. In contrast to D-glucose deprivation, deprivation of a single non-essential amino acid also increased transport, although to a lesser extent than methionine, arginine or leucine deprivation. A kinetic study showed that only the maximal velocity of transport was increased, indicating an augmentation of the number of active transporters. Total protein synthesis was dramatically reduced by 71% or 32% in cells deprived of arginine or leucine, respectively, for 5 h. The amount of free antizyme, or ornithine decarboxylase-antizyme complex, also decreased in these cells, but fell only 16% to 29% below that in normally-fed cells. Treating control cells with cycloheximide, to inhibit the overall protein synthesis, decreased the level of free antizyme by similar to 50% and increased spermidine transport concomitantly. However, when cells were exposed to limited amounts of arginine, leucine or methionine, spermidine transport could be stimulated without alteration of protein synthesis and/or antizyme levels. The present findings indicate that polyamine transport in mammalian cells may be controlled by the availability of amino acids.