| Abstract |
Polyamines (spermidine and spermine) and their precursor putrescine are
ubiquitous in biological systems. Prokaryotes have a higher concentration
of putrescine than of spermidine, and have no or little spermine. On the
other hand eukaryotic cells usually have little putrescine but significant
amounts of spermine as well as spermidine. Concentrations of these amines
were found to be elevated in rapidly proliferating tissues and to increase
when growth was induced. A close correlation between polyamines and
nucleic acid synthesis and content has been observed, and recent studies
using specific inhibitors of polyamine synthesis suggest that the amines
and specially spermidine are involved in these processes. Amphibian
embryos have putrescine and spermidine at higher concentrations than
spermine. An euploid cell line, XB694, was initiated by us from Xenopus
borealis embryos. We have measured the concentrations of putrescine and
spermidine, the synthesis of DNA and the activity of ornithine
decarboxylase (the enzyme that produces putrescine from ornithine) at two
stages of culture: log-phase (4 days) and plateau-phase (9 days). Culture
conditions were previously described. Contents in putrescine and
spermidine were determined by fluorimetry following HP-TLC chromatography
of DNS derivatives. Spermine was present as trace amount. The mean level
of putrescine was very high at log-phase (6.3 nmol/10^6 cells). At plateau-
phase it was reduced more than 9-fold (0.68 nmol/10^6 cells). Spermidine
level was 5.5 nmol/10^6 cells at log-phase and 2.16 nmol/10^6 cells at
plateau-phase, decreasing more slowly (2.5-fold). These differences were
repeatedly observed in other experiments. DNA synthesis was mesured after
a 6.5-h incorporation of 3H-TdR (0.3 muCi/ml; 23 Ci/mmol). PCA-insoluble
material displayed 46 850 +- 4687 cpm/10^6 cells at log-phase and 3 263 +-
284 cpm/10^6 cells at plateau-phase. Autoradiography of the two cell
preparations showed that 60.9% and 7.6% of the nuclei were labelled.
respectively. Ornithine decarboxylase activity was determined in situ.
Incubation mixtures contained 0.1 muCi [I-14C] DL-ornithine (56 mCi/mmol)
and 0.1 mM L-ornithine. Values obtained were 35 513 +- 3 361 cpm (log-
phase) and 5 509 +- 517 cpm (plateau-phase) per h per mg of protein.
Plateau-phase cells display still high putrescine content as compared to
other eukaryotic cells. A good correlation is observed between the levels
of putrescine and the percentage of 3H-TdR-labelled nuclei. The question
of a particular role of the diamine in the proliferation of XB694 cells is
currently under investigation.
|
|---|
