C-13 NMR ANALYSIS OF INTERCOMPARTMENTAL FLOW OF ONE-CARBON UNITS INTO CHOLINE AND PURINES IN SACCHAROMYCES-CEREVISIAE

In Saccharomyces cerevisiae, the three-carbon of serine is normally the major one-carbon donor, although glycine and formate can substitute for serine. The second carbon of glycine enters via the glycine cleavage system in the mitochondria and can satisfy all cellular one-carbon requirements. It remains unresolved, however, as to the route by which these mitochondrial one-carbon units supply cytosolic anabolic processes. In the present work, we have used yeast mutants blocked at selected sites and C-13 NMR to trace the incorporation of glycine-derived mitochondrial 5, 10-methylenetetrahydrofolate into nonmitochondrial synthesis of choline and purines. Label incorporation into choline traces the methylation pathway of choline synthesis from production of serine to methylation of phosphatidylethanolamine. The active one-carbon unit of S-adenosylmethionine involved in methylation reactions originates almost solely from C3 of serine. On the other hand, flow of mitochondrial one-carbon units to 10-formyltetrahydrofolate for purine synthesis is shown to occur via both serine and formate. Formate transport accounts for at least 25% of the total, even during growth with sufficient serine to provide for the one-carbon requirements of the cell. This work shows that the synthetase function of the cytosolic C1-tetrahydrofolate synthase plays a critical role in the processing of mitochondrial one-carbon units to 10-formyltetrahydrofolate pools. In addition, this study provides evidence of two pools of glycine within the mitochondria and establishes a system of analyzing flux into the different folate derivatives.