ORDERED CELL-CYCLE PHASE PERTURBATIONS IN CHINESE-HAMSTER OVARY CELLSTREATED WITH AN S-ADENOSYLMETHIONINE DECARBOXYLASE INHIBITOR

The polyamines putrescine, spermidine, and spermine are needed for nor mal cell cycle progression and polyamine-depleted cells cease to proli ferate. We have investigated cell cycle perturbations in Chinese hamst er ovary cells seeded in the presence of 4-amidinoindan-1-one 2'-amidi nohydrazone (CGP 48664), a potent inhibitor of S-adenosylmethionine de carboxylase, an enzyme which is essential for the synthesis of spermid ine and spermine. At 9 h and at 1, 2, and 3 days after seeding, cells were labelled with the thymidine analog bromodeoxyuridine (BrdUrd) for 30 min, after which the BrdUrd-containing medium was removed and the cells were allowed to progress through the cell cycle in BrdUrd-free m edium before sampling (post-labelling time). Using flow cytometry, cou pled with an indirect immunofluorenscence technique, utilizing monoclo nal anti-BrdUrd and secondary fluorescein-isothiocyanate-conjugated an tibodies, and the DNA stain propidium iodide, cellular BrdUrd and DNA contents were quantified. By investigating the movement of BrdUrd-nonl abelled G(1) cells into S phase during the post-labelling time, a meas ure of the G(1)/S transition was obtained. The time of appearence in G (1) of BrdUrd-labelled cells which had divided was used to monitor the length of the G(2)+M phase. A measure of the S phase length (DNA synt hesis time) was obtained by monitoring the progression of BrdUrd-label led cells through S phrase. CGP 48664-induced spermine depletion signi ficantly increased the length of the S phase already 9 h after seeding cells in the presence of the inhibitor. No effects on the G(1)/S tran sition or on the length of the G(2)+M phase were observed until 2 days after seeding.