Accumulation of 5-methyltetrahydrofolic acid and folylpolyglutamate synthetase expression by mitogen stimulated human lymphocytes

D. W. Fort, R. H. Lark, A. K. Smith, M. Marling Cason, Steven D Weitman, B. Shane, B. A. Kamen

Research output: Contribution to journalArticle

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Abstract

The accumulation of 5-methyl[3H]tetrahydrofolic acid (5CH3[3H]FH4) by phytohaemagglutinin stimulated lymphocytes (PHA-L) cultured in folate free media was investigated to determine the mechanism of uptake of 5CH3FH4 and the requirement of the cells for this vitamin as assessed by monitoring de novo thymidine synthesis. When grown in 20 nM 5CH3[3H]FH4 PHA-L accumulate radiolabel at a rate of 0.04 pmol/h/106 cells. This doubles the endogenous folate pool of unstimulated cells (0.6 ± 0.16 pmol/106 cells) in about 15 h. Uptake proceeded via a saturable process, independent of a high affinity folate receptor as assessed by ligand binding and by Northern and Western blot analysis. However, transport was blocked by probenecid, which is consistent with an anion carrier mechanism. Unstimulated cells lacked folylpolyglutamate synthetase (FPGS) activity and did not express significant amounts of FPGS mRNA. After 48 h of mitogen stimulation there was a 4-10-fold increase in FPGS mRNA and folylpolyglutamate formation (Glu(≥5)) was essentially simultaneous with 5CH3[3H]FH4 transport. Increasing extracellular folate to 2μM only increased intracellular folate 8-fold, but the length of the folylpolyglutamates decreased. The increased folate did not increase de novo thymidine synthesis compared to cells grown in physiological folate. We conclude that mitogen stimulation activates the process(es) for folate accumulation, especially FPGS, and that physiological uptake (0.04 pmol/h/106 cells) is adequate for meeting the cells' need for the vitamin.

Original languageEnglish (US)
Pages (from-to)595-601
Number of pages7
JournalBritish Journal of Haematology
Volume84
Issue number4
StatePublished - 1993
Externally publishedYes

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Mitogens
Folic Acid
Lymphocytes
Phytohemagglutinins
Vitamins
Thymidine
Probenecid
Messenger RNA
5-methyltetrahydrofolate
folylpolyglutamate synthetase
Northern Blotting
Anions
Western Blotting
Ligands

ASJC Scopus subject areas

  • Hematology

Cite this

Fort, D. W., Lark, R. H., Smith, A. K., Marling Cason, M., Weitman, S. D., Shane, B., & Kamen, B. A. (1993). Accumulation of 5-methyltetrahydrofolic acid and folylpolyglutamate synthetase expression by mitogen stimulated human lymphocytes. British Journal of Haematology, 84(4), 595-601.

Accumulation of 5-methyltetrahydrofolic acid and folylpolyglutamate synthetase expression by mitogen stimulated human lymphocytes. / Fort, D. W.; Lark, R. H.; Smith, A. K.; Marling Cason, M.; Weitman, Steven D; Shane, B.; Kamen, B. A.

In: British Journal of Haematology, Vol. 84, No. 4, 1993, p. 595-601.

Research output: Contribution to journalArticle

Fort, DW, Lark, RH, Smith, AK, Marling Cason, M, Weitman, SD, Shane, B & Kamen, BA 1993, 'Accumulation of 5-methyltetrahydrofolic acid and folylpolyglutamate synthetase expression by mitogen stimulated human lymphocytes', British Journal of Haematology, vol. 84, no. 4, pp. 595-601.
Fort, D. W. ; Lark, R. H. ; Smith, A. K. ; Marling Cason, M. ; Weitman, Steven D ; Shane, B. ; Kamen, B. A. / Accumulation of 5-methyltetrahydrofolic acid and folylpolyglutamate synthetase expression by mitogen stimulated human lymphocytes. In: British Journal of Haematology. 1993 ; Vol. 84, No. 4. pp. 595-601.
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