The role of DNA methylation in the expression of DNA polymerase beta in folate deficient mice.

摘要

Folate deficiency (FD) is associated with cancers of the lung, cervix, pancreas, breast, colon, and liver. The underlying mechanism connecting folate deficiency to cancer remains unknown. FD induces a phenotype of single strand breaks, mutations and chromosomal instability suggestive of an inability to repair DNA damage. It is suggested that carcinogenic effect of FD are related to alterations in the methylation status of DNA due to reduction in S-adenosylmethionine (SAM) levels and/or increased uracil content of DNA. The DNA repair pathway for removal of uracil is base excision repair (BER). We have previously shown that FD results in an upregulation in Uracil DNA glycosylase (UDG) in the liver of mice with no upregulation in BER pathway and its rate limiting enzyme, DNA polymerase beta (beta-pol). Imbalance in BER capacity in FD mice has been shown to result in an accumulation in DNA repair intermediates, e.g., DNA single strand breaks and aldehydic lesions, and has been shown to be exacerbated in BER haploinsufficient mice. Here we show that FD inhibits the DNA damage inducibility of beta-pol at the level of transcription in liver in response to oxidative stress induced by treatment with 2-nitropropane, 2-NP. Thus, a direct inhibition of the BER response to carcinogens in folate deficient animals appears to be due to transcriptional repression of beta-pol gene. These data indicate that FD alters the ability of the animals to respond to deficiency. FD could alter the expression of beta-pol either through alteration in methylation status of DNA or through alteration in level and/or activity of specific regulatory factors. To elucidate the mechanism by which FD inhibits beta-pol upregulation, we analyzed the methylation status of beta-pol promoter during FD using a bisulfite genomic sequencing technique. We have shown that methylation status of exon 1 and the first 300 bases of the beta-pol promoter are not affected by FD. Since p53 regulates BER, the effect of folate deficiency on the level of p53 and methylation status of p53 CG rich exon 5-8 was also determined. FD resulted in a significant decline in the level of p53 protein however, FD did not alter the methylation status of exon 5-8 of the p53 gene. Hence, DNA methylation does not appear to be the underlying mechanism by which FD alters expression of beta-pol and BER capacity. This study shows no epigenetic changes in the promoter region of the beta-pol promoter during folate deficiency. Therefore, further investigation is needed to know the particular mechanism through which folate deficiency inhibits the expression of beta-pol.