Journal Article

Methylenetetrahydrofolate reductase C677T polymorphism does not alter folic acid deficiency-induced uracil incorporation into primary human lymphocyte DNA <i>in vitro</i>

Jimmy W. Crott, Susan T. Mashiyama, Bruce N. Ames and Michael F. Fenech

in Carcinogenesis

Volume 22, issue 7, pages 1019-1025
Published in print July 2001 | ISSN: 0143-3334
Published online July 2001 | e-ISSN: 1460-2180 | DOI: http://dx.doi.org/10.1093/carcin/22.7.1019
Methylenetetrahydrofolate reductase C677T polymorphism does not alter folic acid deficiency-induced uracil incorporation into primary human lymphocyte DNA in vitro

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Methylenetetrahydrofolate reductase (MTHFR) is an enzyme which converts 5,10-methylene tetrahydrofolate (5,10-MnTHF) to 5-methyl tetrahydrofolate. A common C to T transition (C677T) in the MTHFR gene is reported to reduce the risk for colorectal cancer and acute lymphocytic leukemia in homozygotes (TTs). It is hypothesized that because TTs have reduced MTHFR activity, more 5,10-MnTHF is available to provide methyl groups for the conversion of uracil to thymidine. Folic acid deficiency causes the intracellular accumulation of dUMP and the subsequent incorporation of uracil into DNA. The removal of uracil from DNA may result in double-stranded DNA breaks, the accumulation of which is a putative risk factor for cancer. We tested whether human lymphocytes taken from TTs (n = 10) were more able to resist uracil incorporation into DNA than controls (n = 14 CCs and 6 CTs) when cultured in medium containing 12–120 nM folic acid for 9 days. DNA uracil content of these lymphocytes was measured by CG-MS. TTs and controls showed a dose-dependent increase in DNA uracil content during folic acid deficiency (P < 0.0001, R2 = 0.23 for TTs and P < 0.0001, R2 = 0.19 for controls). DNA uracil content was not different between the two groups at any of the folic acid concentrations (two-way ANOVA: media [folic acid], P < 0.0001; genotype, P = 0.4). The results show that, in this in vitro system, the MTHFR C677T polymorphism does not affect the cell's ability to resist uracil incorporation into DNA. Chromosome breakage, as measured by micronuclei, was also shown to correlate with folic acid concentration in a preliminary experiment (P < 0.0001). Although the results appear not to support the hypothesis that a reduced risk for certain cancers in TTs is due to diversion of folic acid to thymidine synthesis, differences between the in vivo and in vitro situation make this conclusion not definitive.

Keywords: ALL, acute lymphocytic leukemia; BN, binucleated cell; CBMN, cytokinesis block micronucleus assay; CC/CT/TT, persons nullizygous/heterozygous/homozygous for the C677T polymorphism of MTHFR; DSB, double-stranded (DNA) break; FCS, fetal calf serum; GC-MS, gas chromatography-mass spectrometry; MTHFR, methylenetetrahydrofolate reductase; PHA, phytohaemagglutinin; MNed, micronucleated; MS, methionine synthase; THF, tetrahydrofolate; 5,10-MnTHF/5-MeTHF, 5,10-methylene/5-methyl THF; UDG, uracil DNA glycosylase.

Journal Article.  6593 words.  Illustrated.

Subjects: Clinical Cytogenetics and Molecular Genetics

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