REGULATION OF FOLATE AND ONE-CARBON METABOLISM IN MAMMALIAN-CELLS .1.FOLATE METABOLISM IN CHINESE-HAMSTER OVARY CELLS EXPRESSING ESCHERICHIA-COLI OR HUMAN FOLYLPOLY-GAMMA-GLUTAMATE SYNTHETASE-ACTIVITY

Chinese hamster ovary (CHO) cell transfectants expressing various leve ls of human and Escherichia coli folylpoly-gamma-glutamate synthetase (FPGS) activity and possessing different folylpolyglutamate chain leng th distributions have been developed as models for folate and antifola te metabolism. The synthesis of pteroyltriglutamate was sufficient for normal cellular retention of folate and also overcame the phenotypic requirement for purines and thymidine of AUXB1, a CHO cell mutant lack ing FPGS activity and lacking folylpolyglutamates. Only low levels of FPGS are required to enable cellular metabolism of folates to forms th at are retained by mammalian cells. The higher levels found in mammali an cells are required for the synthesis of the long chain polyglutamat e derivatives characteristic of mammalian cells. At low medium folate concentrations, folate accumulation by transfectants expressing human FPGS was not responsive to FPGS levels as the limiting step in metabol ism was beyond the triglutamate, the chain length required for retenti on. The rate-limiting step in folate metabolism in cells expressing th e E. coli enzyme was the conversion of diglutamate to triglutamate, an d, at low FPGS levels, the E. coli enzyme was about 50-fold less effec tive than the human FPGS in enabling cellular folate accumulation. The se data suggest that cellular accumulation of any folate analog whose mono- or diglutamate derivative is a poor substrate for FPGS would be very responsive to the level of FPGS activity.