Effects of conditional overexpression of spermidine/spermine N1-acetyltransferase on polyamine pool dynamics, cell growth, and sensitivity to polyamine analogs.
Acetylation of polyamines by spermidine/spermine N(1)-acetyltransferase (SSAT) has been implicated in their degradation and/or export out of the cell. The relationship of SSAT to polyamine pool dynamics and cell growth is not yet clearly understood. MCF-7 human breast carcinoma cells were transfected with tetracycline-regulated (Tet-off) SSAT human cDNA or murine gene. Doxycycline removal for >2 days caused a approximately 20-fold increase in SSAT RNA and a approximately 10-fold increase in enzyme activity. After 4 days, intracellular putrescine and spermidine pools were markedly lowered, and cell growth was inhibited. Growth inhibition could not be prevented with exogenous polyamines due to a previously unrecognized ability of SSAT to rapidly acetylate influxing polyamines and thereby prevent restoration of the endogenous pools. Instead, cells accumulated high levels of N(1)-acetylspermidine, N(1)-acetylspermine, and N(1), N(12)-diacetylspermine, a metabolite not previously reported in mammalian cells. Doxycycline deprivation before treatment with N(1), N(11)-diethylnorspermine markedly increased analog induction of SSAT mRNA and activity and enhanced growth sensitivity to the analog by approximately 100-fold. Overall, the findings demonstrate that conditional overexpression of SSAT lowers polyamine pools, inhibits cell growth, and markedly enhances growth sensitivity to certain analogs. The enzyme also plays a remarkably efficient role in maintaining polyamine pool homeostasis during challenges with exogenous polyamines.
Vujcic
S
S
Grace Cancer Drug Center, Roswell Park Cancer Institute, Buffalo, New York 14263, USA.
Halmekyto
M
M
Diegelman
P
P
Gan
G
G
Kramer
D L
DL
Janne
J
J
Porter
C W
CW
D000107:Acetylation; D000123:Acetyltransferases / Q000235:genetics* / Q000378:metabolism*; D001943:Breast Neoplasms; D002455:Cell Division / Q000187:drug effects / Q000502:physiology*; D002999:Clone Cells; D004318:Doxycycline / Q000494:pharmacology; D005260:Female; D006706:Homeostasis; D006801:Humans; D009955:Ornithine Decarboxylase / Q000378:metabolism; D011073:Polyamines / Q000378:metabolism* / Q000494:pharmacology*; D011700:Putrescine / Q000378:metabolism; D011994:Recombinant Proteins / Q000378:metabolism; D013095:Spermidine / Q000378:metabolism; D014158:Transcription, Genetic; D014162:Transfection; D014407:Tumor Cells, Cultured; D000099228:Diamine N-Acetyltransferase
D000123:Acetyltransferases; D004318:Doxycycline; D009955:Ornithine Decarboxylase; D011073:Polyamines; D011700:Putrescine; D011994:Recombinant Proteins; D013095:Spermidine; D000099228:Diamine N-Acetyltransferase