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

Abstract

Polyamines are required for optimal growth in most cells; however, polyamine accumulation leads to inhibition of cellular growth. To reduce intracellular polyamine levels, spermidine is monoacetylated in both prokaryotes and eukaryotes. In Escherichia coli, the speG gene encodes the spermidine acetyltransferase, which transfers the acetyl group to either the N-1 or N-8 position. In addition to polyamine accumulation, stress conditions, such as cold shock, cause an increase in the level of spermidine acetylation, suggesting an adaptive role for reduced polyamine levels under stressful growth conditions. The effect of spermidine accumulation on the growth of E. coli at low temperature was examined using a speG mutant. At 37°C, growth of the speG mutant was normal in the presence of 0.5 or 1 mM spermidine. However, following a shift to 7°C, the addition of 0.5 or 1 mM spermidine resulted in inhibition of cellular growth or cell lysis, respectively. Furthermore, at 7°C, spermidine accumulation resulted in a decrease in total protein synthesis accompanied by an increase in the synthesis of the major cold shock proteins CspA, CspB, and CspG. However, the addition of 50 mM Mg 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 N 1 -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 polyamin...

Transcriptome Analysis of Agmatine and Putrescine Catabolism inPseudomonas aeruginosaPAO1

“…In E. coli, polyamine acetylation is catalyzed by SpeG (4,5,19), and acetylated polyamine is readily excreted (27). E. coli might not recycle acetylated polyamine, and no apparent AphA homologue can be found in this organism as revealed by a BLAST search against all available genome sequences of E. coli strains in the NCBI database.…”

“…Recently, we also reported that exogenous polyamines exert a significant effect on antibiotic susceptibility in bacteria (16)(17)(18). While intracellular polyamines play pivotal roles in ensuring optimal growth, their accumulation inhibits protein synthesis and decreases cell viability in the stationary phase and at low temperatures (5,19). Thus, a balance between polyamine synthesis and catabolism appears to be required to adjust the optimal concentration of these compounds in accordance with the growth environment.…”

Genome-Wide Transcriptional Analysis of the Cold Shock Response in Wild-Type and Cold-Sensitive, Quadruple- csp -Deletion Strains of Escherichia coli

“…Genes encoding flagellar proteins were induced and maintained at high levels even after 5 h at 15°C in the wild-type strain and were down-regulated in the quadrupledeletion strain. Spermidine acetyltransferase encoded by speG is required to prevent spermidine toxicity at low temperatures in E. coli (18). Our DNA microarray analysis showed a steady increase in speG levels from three-to fivefold at 1 to 5 h after temperature downshift (Table 3).…”

Growth-related changes in intracellular spermidine and its effect on efflux pump expression and quorum sensing in Burkholderia pseudomallei

“…An E. coli speG mutant lacking spermidine acetyltransferase shows growth inhibition during cold shock due to the accumulation of high levels of spermidine. The growth inhibition is attributed to ribosome inactivation caused by a replacement of ribosome-bound Mg 2+ by spermidine (Limsuwun & Jones, 2000). The E. coli speG mutant is also sensitive to exogenous spermidine and exhibits decreased cell viability, especially when exogenous spermidine is added to 24 h stationary phase cells (Fukuchi et al, 1995).…”

“…Apart from ensuring optimal growth, too many free polyamines are toxic to the bacterium because they can inhibit protein synthesis, especially during stationary phase and at low temperatures (Fukuchi et al, 1995;Limsuwun & Jones, 2000). Hence, a balance between polyamine synthesis and catabolism is needed.…”