Regulation of polyamine transport by polyamines and polyamine analogs

Abstract

Regulation of polyamine transport in murine L1210 leukemia cells was characterized in order to better understand its relationship to specific intracellular polyamines and their analogs and to quantitate the sensitivity by which it is controlled. Up-regulation of polyamine uptake was evaluated following a 48-hr treatment with a combination of biosynthetic enzyme inhibitors to deplete intracellular polyamine pools. The latter declined gradually over 48 hr and was accompanied by a steady increase in spermidine (SPD) and spermine (SPM) transport as indicated by rises in Vmax to levels approximately 4.5 times higher than control values. Restoration of individual polyamine pools during a 6-hr period following inhibitor treatment revealed that SPD and SPM uptake could not be selectively affected by specific pool changes. The effectiveness of individual polyamines in reversing inhibitor-induced stimulation of uptake was as follows: putrescine < SPD < SPM = the SPM analog, N1, N12-bis(ethyl)spermine (BESPM). In contrast to stimulation of transport, down-regulation by exogenous polyamines or analogs occurred rapidly and in response to subtle increases in intracellular pools. Following a 1-hr exposure to 10 microM BESPM, Vmax values for SPD and SPM fell by 70%, whereas the analog pool increased to only 400-500 pmol/10(6) cells--about 15-20% of the total polyamine pool (approximately 2.8 nmol/10(6) cells). SPM produced nearly identical regulatory effects on transport kinetics. Both BESPM and SPM were even more effective at down-regulating transport that had been previously stimulated four to fivefold by polyamine depletion achieved with enzyme inhibitors. A dose response with BESPM at 48 hr revealed a biphasic effect on uptake whereby concentrations of analog < 3 microM produced an increase in SPD and SPM Vmax values, whereas concentrations 3 microM and higher produced a marked suppression of these values. Cells treated with 3 microM BESPM for 2 hr and placed in analog-free medium recovered transport capability in only 3 hr. Thus, whereas stimulation of polyamine transport is a relatively insensitive and slowly responsive process that tends to parallel polyamine depletion, down-regulation of polyamine transport by exogenous polyamines and analogs and its reversal are rapidly responsive events that correlate with relatively small (i.e., 15-20%) changes in intracellular polyamine pools.

Differential post‐transcriptional control of ornithine decarboxylase and spermidine‐spermine N1‐acetyltransferase by polyamines

“…The results illustrate how polyamine catabolism as regulated by SSAT and polyamine biosynthesis as regulated by ODC can be rapidly and simultaneously controlled by intracellular pools. These responses, together with that of polyamine transport which is also known to be regulated by polyamines [7][8][9], function in a coordinated manner to sensitively maintain intracellular polyamine pools at a relatively constant level [3]. Such a homeostatic system would ensure an adequate supply of polyamines for cell growth while at the same time preventing cellular toxicity due to polyamine excess.…”

“…These effectors are short-lived and differentially regulated by intracellular polyamine pools [3]. Exposure to exogenous polyamines causes polyamine biosynthesis (reviewed in [5,6]) and uptake [7][8][9] to decrease and polyamine catabolism to increase *Corresponding author. Fax: (1) (716) 845-8857.…”

Regulation of spermidine/spermine N1‐acetyltransferase by intracellular polyamine pools

“…Although the steady-state levels of c-myc and c-fos in COLO 320 cells have been shown to decrease with polyamine depletion [29], the inverse relationship, whereby mRNA levels increase with the addition of polyamines to intact cells, has not been demonstrated. Also, when these same cells are treated with a polyamine analog, N' W2-bis(ethyl)spermine, which is known to mimic namral polyamines in down-regulating ODC [30] and transport [31,32] and by up-regulating SSAT [8-121, the steadystate mRNA levels of these proto-oncogenes decreases instead of increasing [29]. Thus, unlike SSAT, the expression of these genes does not appear to be regulated by polyamines via a truly positive relationship.…”

“…The former was apparent from experiments with exogenous polyamines and the latter by pool depletion with specific enzyme inhibitors. The biosynthetic enzymes, ODC and SAMDC [l], and the polyamine transport system [2,31,32,35], are also sensitively upregulated and down-regulated by changes in intracellular polyamine pools. Unlike SSAT, however, the regulatory relationships for these systems are negative in nature -as pools decrease, enzyme and transport activities increase and vice versa.…”

“…Studies with polyamine analogs are supportive of the model. Cells treated with such analogs respond as though polyamine excess exists, namely ODC and SAMDC are down-regulated [12], transport is down-regulated [31,32], and SSAT is markedly up-regulated . Acetylated polyamines are excreted into the medium [12,36].…”