Spermidine Acetyltransferase Is Required To Prevent Spermidine Toxicity at Low Temperatures in Escherichia coli

Polyamines, such as spermidine and putrescine, are present in virtually all cells (25, 33). These polycations have pleiotropic properties, which can influence several cellular processes. They can bind to nucleic acids, stabilize membranes, and stimulate the activity of several enzymes, such as RNA polymerase (1, 31, 34). Found in the ribosomal fraction, polyamines enhance the synthesis of several proteins and stimulate the in vivo assembly of the Escherichia coli 30S ribosomal subunit (7, 13, 21, 35). Although intracellular polyamine is required for optimal growth, polyamine accumulation can lead to inhibition of cellular growth (11, 25, 26, 33). The addition of spermidine to cell cultures of mouse FM3A cells results in a decrease in cell growth accompanied by inhibition of protein synthesis (11). To prevent polyamine toxicity in eukaryotes, polyamine catabolism is initiated by the monoacetylation of spermidine and spermine catalyzed by spermidine/spermine N1-acetyltransferase (SSAT) (4). The acetylpolyamines are then either further oxidized by polyamine oxidase or excreted from the cell. In E. coli, the speG gene encodes spermidine acetyltransferase (SAT), which transfers the acetyl group to either the N-1 or N-8 position of the polyamine (2, 8, 20). SAT is required to reduce the spermidine level, since the addition of exogenous spermidine to a speG mutant results in intracellular accumulation of spermidine (9). Furthermore, the excess spermidine causes decreased protein synthesis and cell viability during the stationary phase of growth (9). Acetylation serves to convert the polyamine to a physiologically inert form; acetylpolyamines cannot substitute for polyamines in RNA binding, in the enhancement of growth of an E. coli polyamine-deficient mutant, or in the stimulation of in vitro translation (18). Neither spermidine-deacetylating activity nor polyamine oxidase activity has been detected in E. coli, suggesting that the N-acetylspermidine is either excreted or kept in the inert acetylated form (14, 24). SAT is induced under nutrient-poor conditions, and its level is higher in cells growing in minimal medium than in those growing in rich medium (8). In addition to high polyamine levels, chemical and physical stresses result in an increased level of spermidine acetylation in E. coli and mammalian cells, suggesting a role for polyamine acetylation in adaptation to stress (2, 4, 10, 27, 32). Shifting E. coli to low temperature results in inhibition of growth and protein synthesis accompanied by the onset of the cold shock response (16). The cold shock response consists of the induction of a set of proteins that has been proposed to aid in cellular adaptation to the low temperature. E. coli also responds to a downshift in temperature by increasing the level of spermidine acetylation (2, 32). However, the role of reduced spermidine levels in growth at low temperature is not known. In this study, we examined the effect of spermidine accumulation on adaptation to low temperatures. We found that the addition of spermidine to a speG mutant resulted in inhibition of growth and protein synthesis at low temperature. However, the inhibition of growth or protein synthesis caused by the addition of 0.5 mM spermidine could be suppressed by the presence of a speG+ multicopy plasmid or the inclusion of magnesium in the growth medium. The data indicate that spermidine acetylation at low temperatures occurs to reduce the toxic effect of spermidine accumulation.