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

In order to examine the biosynthesis, interconversion, and degradation of purine and pyrimidine nucleotides in white spruce cells, radiolabeled adenine, adenosine, inosine, uracil, uridine, and orotic acid were supplied exogenously to the cells and the overall metabolism of these compounds was monitored. [8-C-14]adenine and [8-C-14]adenosine were metabolized to adenylates and part of the adenylates were converted to guanylates and incorporated into both adenine and guanine bases of nucleic acids, A small amount of [8-C-14]inosine was converted into nucleotides and incorporated into both adenine and guanine bases of nucleic acids. High adenosine kinase and adenine phosphoribosyltransferase activities in the extract suggested that adenosine and adenine were converted to AMP by these enzymes. No adenosine nucleosidase activity was detected. Inosine was apparently converted to AMP by inosine kinase and/or a non-specific nucleoside phosphotransferase, The radioactivity of [8-C-14]adenosine, [8-C-14]adenine, and [8-C-14]inosine was also detected in ureide, especially allantoic acid, and CO2. Among these 3 precursors, the radioactivity from [8-C-14]inosine was predominantly incorporated into 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 acids. The salvage enzymes, uridine kinase and uracil phosphoribosyltransferase, were detected in white spruce extracts, [6-C-14]orotic acid, an intermediate 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 observed in the extracts. A large proportion of radioactivity from [2-C-14]uracil was recovered as CO2 and beta-ureidopropionate. Thus, a reductive pathway of uracil degradation is functional in these cells. Therefore, white spruce cells in culture demonstrate both the de novo and salvage pathways of purine and pyrimidine metabolism, as well as some degradation of the substrates into CO2.