Precursor-dependent differences in the incorporation of fluorouracil in RNA

The distribution and level of 5-fluorouracil (FUra) incorporation into RNA were studied in S-180 murine tumor cells for two different fluoropyrimidine nucleosides: 5-fluorouridine (FUrd) and 5'-deoxy-5-fluorouridine (5'-dFUrd) under both cytotoxic (thymidine nonreversible) and nontoxic conditions. Exposure of cells to 200 microM 5'-dFUrd for 6 hr resulted in minor thymidine-nonreversible toxicity (11% cell kill) and produced an FUra incorporation level of 12.5 pmol of FUra/microgram of RNA. Exposure of cells to 1 microM FUrd for 6 hr resulted in considerably more toxicity (74% cell kill) but was found to produce less FUra incorporation (7.8 pmol of FUra/microgram of RNA) than the nontoxic concentration of 5'-dFUrd. Subcellular fractionation (i.e., nucleolar, nucleoplasmic, cytoplasmic) was completed, and the density of FUra incorporation from 5'-dFUrd was found to be approximately equally distributed between nucleoplasmic RNA and nucleolar RNA, independent of toxicity or concentration levels (average nucleolar/nucleoplasmic FUra ratio = 0.75). FUrd, however, was preferentially incorporated into nucleolar RNA (average nucleolar/nucleoplasmic FUra ratio = 22). The nontoxic accumulation of FUra in RNA of cells treated with 200 microM 5'-dFUrd was also associated with the mature cytoplasmic 28 S and 18 S rRNA, the levels of which increased considerably with exposure time. In contrast, in cells exposed to 1 microM FUrd, there was very little FUra measured in the mature 28 S and 18 S rRNA, consistent with an inhibition of rRNA processing. It is concluded that 5'-dFUrd can produce large nontoxic levels of FUra accumulation in cellular RNA compared with FUrd because total FUra incorporation in nuclear RNA was almost equally dispersed between nucleoplasmic and nucleolar RNA, whereas nuclear FUra incorporation from FUrd was preferential for nucleolar RNA. Also, for nontoxic doses of 5'-dFUrd, unaltered rRNA processing over time can result in the large accumulation of FUra-containing rRNA in the cytoplasm.