Effects of the S-adenosylmethionine decarboxylase inhibitor, 5'-{[(Z)-4-amino-2-butenyl]methylamino}-5' -deoxyadenosine, on cell growth and polyamine metabolism and transport in Chinese hamster ovary cell cultures

T. L. Byers, R. S. Wechter, R. H. Hu, Anthony Pegg

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Abstract

The regulation of polyamine transport and the roles of polyamine transport and synthesis in cell growth were investigated using cultured Chinese hamster ovary (CHO) cells and CHOMG cells which are mutants lacking polyamine-transport activity. Metabolically stable methylated polyamine analogues were used to measure polyamine accumulation, and the irreversible S-adenosyl-L-methionine decarboxylase inhibitor. 5'-{[(Z)-4-amino-2-butenyl]methylamino}-5'-deoxyadenosine (AbeAdo), was used to inhibit synthesis. Exposure to AbeAdo led to a dose-dependent decrease in growth for both cell lines, although CHOMG cells were more sensitive. Intracellular putrescine levels were greatly increased in AbeAdo-treated CHO cells and to a lesser extent in CHOMG cells, whereas intracellular spermidine and spermine levels were substantially reduced in both. Treatment with AbeAdo increased putrescine content in the culture medium to a much greater extent in CHOMG cultures indicating that a portion of the excess putrescine synthesized in response to AbeAdo treatment is excreted, but that CHO cells salvage this putrescine whereas it is lost to CHOMG cells which cannot take up polyamines. AbeAdo treatment increased polyamine transport into CHO cells despite high intracellular putrescine, suggesting that spermidine and/or spermine, and not putrescine, are the major factors regulating transport activity. The accumulation of either 1-methylspermidine or 1,12-dimethylspermine was significantly increased by AbeAdo treatment. Accumulation was increased even further when protein synthesis was blocked by cycloheximide, indicating that a short-lived protein is involved in the regulation of polyamine uptake. In the presence of cycloheximide and AbeAdo or α-difluoromethylornithine methylated polyamine derivatives accumulated to very high levels leading to cell death. These results show that the polyamine-transport system plays an important role in retaining intracellular polyamines and that down-regulation of the transport system in response to increased intracellular polyamine content is necessary to prevent accumulation of toxic levels of polyamines.

Original languageEnglish (US)
Pages (from-to)89-96
Number of pages8
JournalBiochemical Journal
Volume303
Issue number1
DOIs
StatePublished - Jan 1 1994

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Adenosylmethionine Decarboxylase
Polyamines
Cell growth
Cricetulus
Cell culture
Metabolism
Ovary
Cell Culture Techniques
Putrescine
Growth
Spermidine
Spermine
Cycloheximide
MDL 73811
Cells
Eflornithine
Salvaging
S-Adenosylmethionine
Poisons
Cell death

All Science Journal Classification (ASJC) codes

  • Biochemistry
  • Molecular Biology
  • Cell Biology

Cite this

@article{361e723a756b4e969526ace3372ef221,
title = "Effects of the S-adenosylmethionine decarboxylase inhibitor, 5'-{[(Z)-4-amino-2-butenyl]methylamino}-5' -deoxyadenosine, on cell growth and polyamine metabolism and transport in Chinese hamster ovary cell cultures",
abstract = "The regulation of polyamine transport and the roles of polyamine transport and synthesis in cell growth were investigated using cultured Chinese hamster ovary (CHO) cells and CHOMG cells which are mutants lacking polyamine-transport activity. Metabolically stable methylated polyamine analogues were used to measure polyamine accumulation, and the irreversible S-adenosyl-L-methionine decarboxylase inhibitor. 5'-{[(Z)-4-amino-2-butenyl]methylamino}-5'-deoxyadenosine (AbeAdo), was used to inhibit synthesis. Exposure to AbeAdo led to a dose-dependent decrease in growth for both cell lines, although CHOMG cells were more sensitive. Intracellular putrescine levels were greatly increased in AbeAdo-treated CHO cells and to a lesser extent in CHOMG cells, whereas intracellular spermidine and spermine levels were substantially reduced in both. Treatment with AbeAdo increased putrescine content in the culture medium to a much greater extent in CHOMG cultures indicating that a portion of the excess putrescine synthesized in response to AbeAdo treatment is excreted, but that CHO cells salvage this putrescine whereas it is lost to CHOMG cells which cannot take up polyamines. AbeAdo treatment increased polyamine transport into CHO cells despite high intracellular putrescine, suggesting that spermidine and/or spermine, and not putrescine, are the major factors regulating transport activity. The accumulation of either 1-methylspermidine or 1,12-dimethylspermine was significantly increased by AbeAdo treatment. Accumulation was increased even further when protein synthesis was blocked by cycloheximide, indicating that a short-lived protein is involved in the regulation of polyamine uptake. In the presence of cycloheximide and AbeAdo or α-difluoromethylornithine methylated polyamine derivatives accumulated to very high levels leading to cell death. These results show that the polyamine-transport system plays an important role in retaining intracellular polyamines and that down-regulation of the transport system in response to increased intracellular polyamine content is necessary to prevent accumulation of toxic levels of polyamines.",
author = "Byers, {T. L.} and Wechter, {R. S.} and Hu, {R. H.} and Anthony Pegg",
year = "1994",
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T1 - Effects of the S-adenosylmethionine decarboxylase inhibitor, 5'-{[(Z)-4-amino-2-butenyl]methylamino}-5' -deoxyadenosine, on cell growth and polyamine metabolism and transport in Chinese hamster ovary cell cultures

AU - Byers, T. L.

AU - Wechter, R. S.

AU - Hu, R. H.

AU - Pegg, Anthony

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N2 - The regulation of polyamine transport and the roles of polyamine transport and synthesis in cell growth were investigated using cultured Chinese hamster ovary (CHO) cells and CHOMG cells which are mutants lacking polyamine-transport activity. Metabolically stable methylated polyamine analogues were used to measure polyamine accumulation, and the irreversible S-adenosyl-L-methionine decarboxylase inhibitor. 5'-{[(Z)-4-amino-2-butenyl]methylamino}-5'-deoxyadenosine (AbeAdo), was used to inhibit synthesis. Exposure to AbeAdo led to a dose-dependent decrease in growth for both cell lines, although CHOMG cells were more sensitive. Intracellular putrescine levels were greatly increased in AbeAdo-treated CHO cells and to a lesser extent in CHOMG cells, whereas intracellular spermidine and spermine levels were substantially reduced in both. Treatment with AbeAdo increased putrescine content in the culture medium to a much greater extent in CHOMG cultures indicating that a portion of the excess putrescine synthesized in response to AbeAdo treatment is excreted, but that CHO cells salvage this putrescine whereas it is lost to CHOMG cells which cannot take up polyamines. AbeAdo treatment increased polyamine transport into CHO cells despite high intracellular putrescine, suggesting that spermidine and/or spermine, and not putrescine, are the major factors regulating transport activity. The accumulation of either 1-methylspermidine or 1,12-dimethylspermine was significantly increased by AbeAdo treatment. Accumulation was increased even further when protein synthesis was blocked by cycloheximide, indicating that a short-lived protein is involved in the regulation of polyamine uptake. In the presence of cycloheximide and AbeAdo or α-difluoromethylornithine methylated polyamine derivatives accumulated to very high levels leading to cell death. These results show that the polyamine-transport system plays an important role in retaining intracellular polyamines and that down-regulation of the transport system in response to increased intracellular polyamine content is necessary to prevent accumulation of toxic levels of polyamines.

AB - The regulation of polyamine transport and the roles of polyamine transport and synthesis in cell growth were investigated using cultured Chinese hamster ovary (CHO) cells and CHOMG cells which are mutants lacking polyamine-transport activity. Metabolically stable methylated polyamine analogues were used to measure polyamine accumulation, and the irreversible S-adenosyl-L-methionine decarboxylase inhibitor. 5'-{[(Z)-4-amino-2-butenyl]methylamino}-5'-deoxyadenosine (AbeAdo), was used to inhibit synthesis. Exposure to AbeAdo led to a dose-dependent decrease in growth for both cell lines, although CHOMG cells were more sensitive. Intracellular putrescine levels were greatly increased in AbeAdo-treated CHO cells and to a lesser extent in CHOMG cells, whereas intracellular spermidine and spermine levels were substantially reduced in both. Treatment with AbeAdo increased putrescine content in the culture medium to a much greater extent in CHOMG cultures indicating that a portion of the excess putrescine synthesized in response to AbeAdo treatment is excreted, but that CHO cells salvage this putrescine whereas it is lost to CHOMG cells which cannot take up polyamines. AbeAdo treatment increased polyamine transport into CHO cells despite high intracellular putrescine, suggesting that spermidine and/or spermine, and not putrescine, are the major factors regulating transport activity. The accumulation of either 1-methylspermidine or 1,12-dimethylspermine was significantly increased by AbeAdo treatment. Accumulation was increased even further when protein synthesis was blocked by cycloheximide, indicating that a short-lived protein is involved in the regulation of polyamine uptake. In the presence of cycloheximide and AbeAdo or α-difluoromethylornithine methylated polyamine derivatives accumulated to very high levels leading to cell death. These results show that the polyamine-transport system plays an important role in retaining intracellular polyamines and that down-regulation of the transport system in response to increased intracellular polyamine content is necessary to prevent accumulation of toxic levels of polyamines.

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