Stability of DNA Methylation Patterns in Mouse Spermatogonia Under Conditions of MTHFR Deficiency and Methionine Supplementation 1 Running Title: DNA methylation in MTHFR-deficient male germ cells

Little is known about the conditions contributing to the stability of DNA methylation patterns in male germ cells. Altered folate pathway enzyme activity and methyl donor supply are two clinically significant factors that can affect the methylation of DNA. 5,10Methylenetetrahydrofolate reductase (MTHFR) is a key folate pathway enzyme involved in providing methyl groups from dietary folate for DNA methylation. Mice heterozygous for a targeted mutation in the Mthfr gene (Mthfr +/- ) are a good model for humans homozygous for the MTHFR677C>T polymorphism which is found in 10% of the population and associated with decreased MTHFR activity and infertility. High dose folic acid is administered as an empirical treatment for male infertility. Here, we examined MTHFR expression in developing male germ cells and evaluated DNA methylation patterns and effects of a range of concentrations of methionine in spermatogonia from Mthfr +/- as compared to wildtype, Mthfr +/+ mice. MTHFR