Purine and pyrimidine metabolism in cultured white spruce (Picea glauca) cells: Metabolic fate of C-14-labeled precursors and activity of key enzymes

In order to examine the biosynthesis, interconversion, and degradation of p urine and pyrimidine nucleotides in white spruce cells, radiolabeled adenin e, adenosine, inosine, uracil, uridine, and orotic acid were supplied exoge nously to the cells and the overall metabolism of these compounds was monit ored. [8-C-14]adenine and [8-C-14]adenosine were metabolized to adenylates and part of the adenylates were converted to guanylates and incorporated in to both adenine and guanine bases of nucleic acids, A small amount of [8-C- 14]inosine was converted into nucleotides and incorporated into both adenin e and guanine bases of nucleic acids. High adenosine kinase and adenine pho sphoribosyltransferase activities in the extract suggested that adenosine a nd adenine were converted to AMP by these enzymes. No adenosine nucleosidas e activity was detected. Inosine was apparently converted to AMP by inosine kinase and/or a non-specific nucleoside phosphotransferase, The radioactiv ity of [8-C-14]adenosine, [8-C-14]adenine, and [8-C-14]inosine was also det ected in ureide, especially allantoic acid, and CO2. Among these 3 precurso rs, the radioactivity from [8-C-14]inosine was predominantly incorporated i nto CO2. These results suggest the operation of a conventional degradation pathway, Both [2-C-14]uracil and [2-C-14]uridine were converted to uridine nucleotides and incorporated into uracil and cytosine bases of nucleic acid s. The salvage enzymes, uridine kinase and uracil phosphoribosyltransferase , were detected in white spruce extracts, [6-C-14]orotic acid, an intermedi ate of the de novo pyrimidine biosynthesis, was efficiently converted into uridine nucleotides and also incorporated into uracil and cytosine bases of nucleic acids. High activity of orotate phosphoribosyltransferase was obse rved in the extracts. A large proportion of radioactivity from [2-C-14]urac il was recovered as CO2 and beta-ureidopropionate. Thus, a reductive pathwa y of uracil degradation is functional in these cells. Therefore, white spru ce cells in culture demonstrate both the de novo and salvage pathways of pu rine and pyrimidine metabolism, as well as some degradation of the substrat es into CO2.