Quantitative Analysis of the Pyrimidine Metabolism in Pheochromocytoma PC‐12 Cells

Abstract

A detailed quantitative study of pyrimidine metabolism in exponentially growing rat pheochromocytoma PC-12 cells has been performed. The sizes of ribonucleotide pools have been analysed and the pathways and the rates of metabolism of uridine, cytidine and aspartic acid have been determined, based on the incorporation of radioactive label. The fluxes of radioactive label through uridine-cytidine kinase, cytidine deaminase. CTP synthetase, nucleoside monophosphate kinase and nucleoside diphosphate kinase were obtained, as well as the flux through the pyrimidine de novo pathway. Also, the fluxes of radioactive label towards UDP-sugars, CDP-compounds, DNA and RNA were quantified in situ under steady-state conditions in intact PC-12 cells. From these fluxes of radioactivity, distribution ratios at the branch points of the metabolism were obtained. The pyrimidines synthesised via the de novo pathway were preferentially used for the synthesis of UDP-N-acetylhexosamines and UDP-hexoses, whereas the salvage of precursors from the medium contributed, to a large extent, to the synthesis of RNA. Therefore, we postulate that at least two different UTP pools exist in these cancer cells derived from the neural crest. Furthermore, after metabolism of radiolabeled cytidine and uridine into UTP, radiolabel was distributed in a similar manner from UTP towards UDP-N-acetylhexosamines, UDP-hexoses and RNA-UMP. Uridine, as well as cytidine, was channelled towards nucleic acids via small compartmented ribonucleotide pool