Folate rescues vitamin B₁₂ depletion-induced inhibition of nuclear thymidylate biosynthesis and genome instability.

ABSTRACT: Clinical vitamin B₁₂ deficiency can result in megaloblastic anemia, which results from the inhibition of DNA synthesis by trapping folate cofactors in the form of 5-methyltetrahydrofolate (5-methylTHF) and subsequent inhibition of de novo thymidylate (dTMP) biosynthesis. In the cytosol, vitamin B₁₂ functions in the remethylation of homocysteine to methionine, which regenerates THF from 5-methylTHF. In the nucleus, THF is required for de novo dTMP biosynthesis, but it is not understood how 5-methylTHF accumulation in the cytosol impairs nuclear dTMP biosynthesis. The impact of vitamin B₁₂ depletion on nuclear de novo dTMP biosynthesis was investigated in methionine synthase-null human fibroblast and nitrous oxide-treated HeLa cell models. The nucleus was the most sensitive cellular compartment to 5-methylTHF accumulation, with levels increasing greater than fourfold. Vitamin B₁₂ depletion decreased de novo dTMP biosynthesis capacity by 5-35%, whereas de novo purine synthesis, which occurs in the cytosol, was not affected. Phosphorylated histone H2AX (?H2AX), a marker of DNA double-strand breaks, was increased in vitamin B₁₂ depletion, and this effect was exacerbated by folate depletion. These studies also revealed that 5-formylTHF, a slow, tight-binding inhibitor of serine hydroxymethyltransferase (SHMT), was enriched in nuclei, accounting for 35% of folate cofactors, explaining previous observations that nuclear SHMT is not a robust source of one-carbons for de novo dTMP biosynthesis. These findings indicate that a nuclear 5-methylTHF trap occurs in vitamin B₁₂ depletion, which suppresses de novo dTMP biosynthesis and causes DNA damage, accounting for the pathophysiology of megaloblastic anemia observed in vitamin B₁₂ and folate deficiency.

SUBMITTER: Palmer AM 

PROVIDER: S-EPMC5441772 | biostudies-literature | 2017 May

REPOSITORIES: biostudies-literature