RNA degradation in perfused rat liver as determined from the release of [14C]cytidine

Abstract

The degradation of RNA in the cyclically perfused rat liver was determined from the release of labeled cytidine from RNA that had been previously labeled with [6-14C]orotic acid in vivo. Because cytidine is not appreciably degraded in rat liver (its deamination to uridine is virtually nil) or produced in significant amounts from free 5'-nucleotides, its release will directly reflect net RNA breakdown. This conclusion was substantiated by the fact that the specific radioactivity of released cytidine equaled that of CMP in RNA and remained unchanged for 180 min of perfusion. The initial rate of [14C]cytidine accumulation was slow, but after 10-20 min it increased abruptly by more than 4-fold and remained virtually constant. The addition of 0.5 mM unlabeled cytidine effectively prevented the reutilization of label and increased the rate of labeled cytidine release by an amount representing 13% of the maximal rate of cytidine accumulation. Rates of RNA degradation, measured between 20 and 60 min in the presence of 0.5 mM unlabeled cytidine, averaged 1.00 +/- 0.05 mg h-1 liver-1 (100-g rat), the equivalent of 65% of total RNA per day. This accelerated value, which was about 4-fold larger than the initial rate, is believed to be the direct consequence of amino acid deprivation since, in separate experiments, the increase was completely suppressed by the addition of plasma amino acids (Lardeux, B. R., and Mortimore, G. E. (1987) J. Biol. Chem. 262, 14514-14519). These findings demonstrate the potential value of cytidine as a marker for following moment-to-moment regulatory alterations in RNA degradation in the isolated liver or hepatocyte preparation.