HORMONAL REGULATION OF FORMIMINOTRANSFERASE Glucagon and epinephrine administered intravenously lead to a rapid increase in hepatic and jejunal formiminotransferase, whereas insulin produces a decrease in this activity in rat and man.
Key Words: formiminotransferase, cyclic AMP, hormonal regulation
folic acid,
Adaptive changes in several rate-limiting enzymes of glycolysis and gluconeogenesis follow dietary intake of carbohydrate' or oral folic acid.* Adaptive increases in several j ej unal fo Ia t e-metaboliz ing enzymes also result from dietary carbohydrate, as well as f ~ l a t e .As ~ several hormones that are mediated by cyclic 3',5'-adenosine monophosphate regulate such changes i n it became of carbohydrate m e t a b ~ l i s m , ~ interest t o see if they would have a similar effect on folate metabolism. F. B. Stifel et al. have now investigated the effects of glucagon, insulin, epinephrine, and cyclic AMP on hepatic and jejunal formiminotransferase activity in rat and man.5 This transferase is a key enzyme in t h e u t i l i z a t i o n of N-formiminoL-glutamate, derived from histidine catabolism, f o r formation of 5-formiminotetrahydrofolate. The latter is ultimately converted t o such important compounds as N1O-formyltetrahydrofolate, which is involved in purine biosynthesis. Deficiency of formiminotransferase results in elevated serum folate, formiminoglutamicaciduria, a subtle megaloblastic anemia, and increased urinary aminoimidazolecarboxamide. In their work, Stifel et al.5 used 150 t o 230 g male Carworth rats maintained on rat chow and water ad libitum. The animals were anesthetized with pentobarbital administered intraperitoneal ly, the abdomens opened, and hormones or 0.9 percent saline injected into the portal vein following the removal of control samples of liver or jejunum. In some cases, puromycin dihy-
drochloride (23 mg) or actinomycin D (0.66 pg per gram) were injected intraperitoneally an hour to two before administration of hormone. In other cases, nucleotides were injected a t 5, 10, or 15 minutes prior to removal of liver samples. Human studies were conducted with five normal male subjects who were fed in six equal portions per day a diet containing in percent: carbohydrate (equal glucose and fructose), 50; corn oil, 30; sodium caseinate, 20. Each person also received 192 mg of ferrous sulfate and one nonavitamin tablet daily. The folic acid content was 14 pg. Jejunal biopsies were obtained on days four and nine. Before the first feeding on day six, 15 units of NPH insulin was injected subcutaneously. Other studies included four children, two of whom had glucose-6-phosphatase deficiency, and the o t h e r t w o debrancher (amylo-l,6glucosidase) deficiency. From these patients, liver biopsies were obtained before and after 2 or 3 minutes of intravenous infusion of glucagon (1 mg per minute). F o r m i m i n o t ransferase was measured spectrophotometrically a t 355 nm essentially as described by T. Arakawa et aL6 on the supernatant solution (60 minutes a t 104,000 x gravity) from homogenates of liver or jejunal mucosa. Formimino-Lglutamate was assayed according t o H. Cyclic AMP was Tabor and L. W~ngarden.~ quantitated in liver following the method of A. G. Gilman' and with the addition of tritium-labeled compound t o monitor recovery. A significant (P < 0.01) increase in hepatic formiminotransferase was found 15 NUTRITION REVIEWSIVOL. 33, NO. 2fFEBRUARY 1975
57
minutes after as little as 1.5 pg of glucagon administered to rats. Maximal response occurred with 100 or more times this level of the hormone, which caused enzyme activity in both liver and jejunal mucosa to nearly double, while the level of hepatic formiminoglutamate fell correspondingly. Pretreatment of animals with puromycin or actinomycin D t o inhibit protein (new enzyme) synthesis had no effect. Insulin elicited the opposite effect, viz. a decrease in formiminotransferaseactivity, which was significant (P < 0.01) with as l i t t l e as 0.01 5 units per kilogram and was nearly maximal with ten times more than this. The hepatic formiminoglutamate increased in inverse correspondence. Again, puromycin and actinomycin D failed to block the effect. Infusion of epinephrine (1 t o 2 pg per minute) produced a rapid stimulatory response on the hepatic transferase and rise in level of cyclic AMP. When 50pmoles of nucleotide was similarly administered, a small, but significant (P < 0.01 1 increase in the transferase activity was found with cyclic AMP, whereas no effect was detected with cyclic GMP, 5'-AMP, or ATP. Although less thoroughly studied, the same general effects on hepatic and jejunal formiminotransferase appear to follow the administration of the hormones to humans. Intravenous glucagon (1 mg per minute) approximately doubled the level of the hepatic transferase of the four child patients within two to three minutes. Subcutaneous insulin (15 units per each of three days) decreased the jejunal transferase activity of the five normal adults fed the high carbohydrate diet. Stifel et aL5 point out that their demonstration of the acute reciprocal hormonal regulation of the formiminotransferase by insulin and glucagon resembles the changes found in the regulation of the gluconeogenic fructose diphosphatase by the same ho r m ~ n e s . ~ The investigators speculate that the control of transferase mediated by cyclic AMP may involve a dephosphorylation-phosphorylation mechanism analogous t o that involved in the regulation of glycogen metabolism. In any event, the 58
NUTRITION REVIEWS/VOL 33, NO. Z/FEBRUARY 1975
hormonal influence on formiminotransferase may well be at the enzyme alteration level, since inhibitors of protein biosynthetic machinery have no effect. The relationship between folate and carbohydrate metabolism may also exist at several levels of operation. 0 1. M. C. Scrutton and M. F. Utter: The Regulation of Glycolysis and Gluconeogenesis in Animal Tissues. Ann, Rev. Biochern. 37: 249-302, 1968 2. R. H. Herman, F. B. Stifel, Y . F. Herman, and N. S. Rosensweig: The Response of Jejunal Glycolytic Enzymes t o a Folate Deficient Diet in Germ-Free and Pathogen-Free Rats. Fed. Proc. 28: 628, 1969 3. F. B. Stifel, R. H. Herman, and N. S. Rosensweig: Dietary Regulation of Glycolytic Enzymes. VII. Effect of Diet and Oral Folate Upon Folate-Metabolizing Enzymes in Rat J e j u n u m . Biochirn. Biophys. Acta 208: 381-386, 1970 4. J. P. Jost and H. V. Rickenberg: Cyclic AMP. Ann. Rev. Biochern. 40: 741-774,1971 5. F. B. Stifel, 0. D. Taunton, H. L. Green, E. G. Lufkin, L. Hagler, and R. H. Herman: Hormonal Regulation of Hepatic and Jejunal Formiminotransferase Activity in Man and Rat. Biochirn. Biophys. Acta 354: 194-205, 1974 6. T. Arakawa, K . Narisawa, K. Tanno, K. Hara, 0. Higashi, Y . Honda, T . Tamura, Y . Wada,T. Mi zuno, and T . Hayashi: Megaloblastic Anemia and Mental Retardation associated with Hyperfolic-Acidemia: Probably due t o N5 Methylterahydrofolate Tranferase Def iciency. Tohoku J. Exp. Med. 93: 1-22, 1967 7. H. Tabor and L. Wyngarden: The Enzymatic F o rrna t i o n of F o r m iminotetrahydrofolic Acid, 5,lO-Methenyltetrahydrofolic Acid, and 10-Formyltetrahydrofolic Acid in the Metabolism of Formiminoglutamic Acid. J. Bid. Chern. 234: 1830-1846,1959 8. A. G. Gilman: A Protein Binding Assay for Adenosine 3':5'-Cyclic Monophosphate. Proc. Hat. Acad. Sci. USA 67: 305-312, 1970 9. 0. D. Taunton, F. B. Stifel, L. H. Greene, and R. H. Herman: Rapid Reciprocal Changes of R a t H epati c G l y c o l y t i c Enzymes and F r u c t ose- 1,6-d i p h osp h a t ase F o I I ow i ng Glucagon and Insulin Injection In Vivo. Biochern. Biophys. Res. Cornrnun. 48: t 663-1670, 1972
Key Words: formiminotransferase, cyclic AMP, hormonal regulation
folic acid,
Adaptive changes in several rate-limiting enzymes of glycolysis and gluconeogenesis follow dietary intake of carbohydrate' or oral folic acid.* Adaptive increases in several j ej unal fo Ia t e-metaboliz ing enzymes also result from dietary carbohydrate, as well as f ~ l a t e .As ~ several hormones that are mediated by cyclic 3',5'-adenosine monophosphate regulate such changes i n it became of carbohydrate m e t a b ~ l i s m , ~ interest t o see if they would have a similar effect on folate metabolism. F. B. Stifel et al. have now investigated the effects of glucagon, insulin, epinephrine, and cyclic AMP on hepatic and jejunal formiminotransferase activity in rat and man.5 This transferase is a key enzyme in t h e u t i l i z a t i o n of N-formiminoL-glutamate, derived from histidine catabolism, f o r formation of 5-formiminotetrahydrofolate. The latter is ultimately converted t o such important compounds as N1O-formyltetrahydrofolate, which is involved in purine biosynthesis. Deficiency of formiminotransferase results in elevated serum folate, formiminoglutamicaciduria, a subtle megaloblastic anemia, and increased urinary aminoimidazolecarboxamide. In their work, Stifel et al.5 used 150 t o 230 g male Carworth rats maintained on rat chow and water ad libitum. The animals were anesthetized with pentobarbital administered intraperitoneal ly, the abdomens opened, and hormones or 0.9 percent saline injected into the portal vein following the removal of control samples of liver or jejunum. In some cases, puromycin dihy-
drochloride (23 mg) or actinomycin D (0.66 pg per gram) were injected intraperitoneally an hour to two before administration of hormone. In other cases, nucleotides were injected a t 5, 10, or 15 minutes prior to removal of liver samples. Human studies were conducted with five normal male subjects who were fed in six equal portions per day a diet containing in percent: carbohydrate (equal glucose and fructose), 50; corn oil, 30; sodium caseinate, 20. Each person also received 192 mg of ferrous sulfate and one nonavitamin tablet daily. The folic acid content was 14 pg. Jejunal biopsies were obtained on days four and nine. Before the first feeding on day six, 15 units of NPH insulin was injected subcutaneously. Other studies included four children, two of whom had glucose-6-phosphatase deficiency, and the o t h e r t w o debrancher (amylo-l,6glucosidase) deficiency. From these patients, liver biopsies were obtained before and after 2 or 3 minutes of intravenous infusion of glucagon (1 mg per minute). F o r m i m i n o t ransferase was measured spectrophotometrically a t 355 nm essentially as described by T. Arakawa et aL6 on the supernatant solution (60 minutes a t 104,000 x gravity) from homogenates of liver or jejunal mucosa. Formimino-Lglutamate was assayed according t o H. Cyclic AMP was Tabor and L. W~ngarden.~ quantitated in liver following the method of A. G. Gilman' and with the addition of tritium-labeled compound t o monitor recovery. A significant (P < 0.01) increase in hepatic formiminotransferase was found 15 NUTRITION REVIEWSIVOL. 33, NO. 2fFEBRUARY 1975
57
minutes after as little as 1.5 pg of glucagon administered to rats. Maximal response occurred with 100 or more times this level of the hormone, which caused enzyme activity in both liver and jejunal mucosa to nearly double, while the level of hepatic formiminoglutamate fell correspondingly. Pretreatment of animals with puromycin or actinomycin D t o inhibit protein (new enzyme) synthesis had no effect. Insulin elicited the opposite effect, viz. a decrease in formiminotransferaseactivity, which was significant (P < 0.01) with as l i t t l e as 0.01 5 units per kilogram and was nearly maximal with ten times more than this. The hepatic formiminoglutamate increased in inverse correspondence. Again, puromycin and actinomycin D failed to block the effect. Infusion of epinephrine (1 t o 2 pg per minute) produced a rapid stimulatory response on the hepatic transferase and rise in level of cyclic AMP. When 50pmoles of nucleotide was similarly administered, a small, but significant (P < 0.01 1 increase in the transferase activity was found with cyclic AMP, whereas no effect was detected with cyclic GMP, 5'-AMP, or ATP. Although less thoroughly studied, the same general effects on hepatic and jejunal formiminotransferase appear to follow the administration of the hormones to humans. Intravenous glucagon (1 mg per minute) approximately doubled the level of the hepatic transferase of the four child patients within two to three minutes. Subcutaneous insulin (15 units per each of three days) decreased the jejunal transferase activity of the five normal adults fed the high carbohydrate diet. Stifel et aL5 point out that their demonstration of the acute reciprocal hormonal regulation of the formiminotransferase by insulin and glucagon resembles the changes found in the regulation of the gluconeogenic fructose diphosphatase by the same ho r m ~ n e s . ~ The investigators speculate that the control of transferase mediated by cyclic AMP may involve a dephosphorylation-phosphorylation mechanism analogous t o that involved in the regulation of glycogen metabolism. In any event, the 58
NUTRITION REVIEWS/VOL 33, NO. Z/FEBRUARY 1975
hormonal influence on formiminotransferase may well be at the enzyme alteration level, since inhibitors of protein biosynthetic machinery have no effect. The relationship between folate and carbohydrate metabolism may also exist at several levels of operation. 0 1. M. C. Scrutton and M. F. Utter: The Regulation of Glycolysis and Gluconeogenesis in Animal Tissues. Ann, Rev. Biochern. 37: 249-302, 1968 2. R. H. Herman, F. B. Stifel, Y . F. Herman, and N. S. Rosensweig: The Response of Jejunal Glycolytic Enzymes t o a Folate Deficient Diet in Germ-Free and Pathogen-Free Rats. Fed. Proc. 28: 628, 1969 3. F. B. Stifel, R. H. Herman, and N. S. Rosensweig: Dietary Regulation of Glycolytic Enzymes. VII. Effect of Diet and Oral Folate Upon Folate-Metabolizing Enzymes in Rat J e j u n u m . Biochirn. Biophys. Acta 208: 381-386, 1970 4. J. P. Jost and H. V. Rickenberg: Cyclic AMP. Ann. Rev. Biochern. 40: 741-774,1971 5. F. B. Stifel, 0. D. Taunton, H. L. Green, E. G. Lufkin, L. Hagler, and R. H. Herman: Hormonal Regulation of Hepatic and Jejunal Formiminotransferase Activity in Man and Rat. Biochirn. Biophys. Acta 354: 194-205, 1974 6. T. Arakawa, K . Narisawa, K. Tanno, K. Hara, 0. Higashi, Y . Honda, T . Tamura, Y . Wada,T. Mi zuno, and T . Hayashi: Megaloblastic Anemia and Mental Retardation associated with Hyperfolic-Acidemia: Probably due t o N5 Methylterahydrofolate Tranferase Def iciency. Tohoku J. Exp. Med. 93: 1-22, 1967 7. H. Tabor and L. Wyngarden: The Enzymatic F o rrna t i o n of F o r m iminotetrahydrofolic Acid, 5,lO-Methenyltetrahydrofolic Acid, and 10-Formyltetrahydrofolic Acid in the Metabolism of Formiminoglutamic Acid. J. Bid. Chern. 234: 1830-1846,1959 8. A. G. Gilman: A Protein Binding Assay for Adenosine 3':5'-Cyclic Monophosphate. Proc. Hat. Acad. Sci. USA 67: 305-312, 1970 9. 0. D. Taunton, F. B. Stifel, L. H. Greene, and R. H. Herman: Rapid Reciprocal Changes of R a t H epati c G l y c o l y t i c Enzymes and F r u c t ose- 1,6-d i p h osp h a t ase F o I I ow i ng Glucagon and Insulin Injection In Vivo. Biochern. Biophys. Res. Cornrnun. 48: t 663-1670, 1972
