Free pyrimidine nucleotide pool of Ehrlich ascites-tumour cells. Characteristics related to quantitative studies of RNA metabolism

Ehrlich ascites-tumour cells incubated in a mineral medium supplemented with glucose and glutamine intensely incorporated labelled uridine into free nucleotides and RNA. Detailed kinetic studies of the labelling of total acid-soluble pools of uridine and cytidine nucleotides and of RNA under standard conditions and after chase with non-radioactive uridine were carried out. The relative distribution of the radioactivity between the individual uridine phosphates as well as their total cellular contents were also determined under standard and chase conditions. The labelling kinetics of the mononucleotides and RNA by radioactive uridine and orotate were compared, and some turnover parameters were estimated by mathematical analysis of a steady-state model. The following conclusions were drawn. (1) The observed chase effect of addition of non-radioactive uridine is due to a rapid expansion of the acid-soluble uridine nucleotide pool and consequently a severalfold lowering of its specific radioactivity. (2) The free uridine nucleotides are partly separated into a large and a small compartment, labelled preferentially by exogeneous orotate and uridine respectively. (3) The half-life of the mononucleotides, at least those located in the smaller compartment, is a few minutes only, owing to a rapid exchange with the uridylic acid residues of unstable RNA species. (4) This process of reutilization of RNA-degradation products accounts for 85% (a minimum estimate) of the overall turnover of the nucleotide pool in resting cells.