Dependence of 5-methyltetrahydrofolate utilization by L5178Y murine leukemia cells in vitro on the presence of hydroxycobalamin and transcobalamin II.

Abstract

Summary L5178Y mouse leukemia cells in tissue culture required a minimum of 10.0 mg l-methionine per liter and 1.0 mg folic acid per liter to maintain a division time of 10 to 10.5 hr during a 72-hr period of growth. When folic acid was withheld, the cells underwent three to four divisions before growth stopped. In contrast, when l-methionine was withheld, there was no growth. N5-Formyltetrahydrofolate was 200 times more effective as a folate source than folic acid. A level of 0.005 mg/liter supported a division time of 10 to 10.5 hr for 72 hr. In contrast, when N5-methyltetrahydrofolate was used as the folate source, only slight growth occurred, even at a level of 0.5 mg/liter. Lower levels of 0.05 and 0.005 mg/liter were without growth-promoting effect. When hydroxycobalamin was added to the culture medium, each of the above levels of N5-methyltetrahydrofolate was able to support moderate growth. If the medium were further supplemented with transcobalamin II plus hydroxycobalamin, 0.005 mg N5-methyltetrahydrofolate per liter was able to support a 10- to 10.5-hr division time for 72 hr. These data support the concept that vitamin B12 is necessary for the conversion of N5-methyltetrahydrofolate to other active folate coenzymes, presumably via methionine biosynthesis through the enzyme N5-methyltetrahydrofolate-homocysteine transmethylase. However, in this system, l-methionine supplementation is required for optimum cell growth, even in the presence of N5-methyltetrahydrofolate, hydroxycobalamin, transcobalamin II, and dl-homocysteine.