Folate Status, Genomic Uracil, and the Balance of Base Excision Repair Activity

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DESCRIPTION (provided by applicant): Dietary folic acid is an important nutrient for human health and has received much attention as a means of preventing cancer. Studies have shown that folate supplementation suppresses early stages of carcinogenesis. However, concern is growing that folate supplementation may exacerbate the growth of pre- existing tumors. These observations underscore the need for a better understanding of the molecular mechanisms of pro- and anticancer activities of dietary folate. At a biochemical level, folate derivatives are important sources of one-carbon units for de novo synthesis of nucleotides and are necessary for enzymatic methylation of CpG sequences for gene silencing. A significant mechanism by which folate deficiency is thought to be procarcinogenic is by increasing genome instability. In particular, N5,N10 methylene tetrahydrofolate is required by thymidylate synthase to convert dUMP to TMP, which is the only de novo source of TMP. Thus, folate deficiency decreases TTP, increases dUTP, and increases uracil incorporation into DNA. Uracil in DNA is removed by uracil DNA glycosylases (UDGs) of the Base Excision Repair (BER) pathway. BER removes an array of mutagenic DNA base damage. Unfortunately, BER becomes problematic under folate-deficient conditions. Because BER requires a DNA resynthesis step, elevated dUTP presumptively causes the reintroduction of genomic uracil. BER strand break intermediates are themselves clastogenic, and if not repaired can induce apoptosis. In other words, BER initiated by UDGs appear to unwittingly contribute to genome instability during folate deficiency. There is an additional link that is yet to be explored. MBD4 (MED1) specifically removes uracil or thymine when paired opposite guanine in CpG sequences. In other words, MBD4 is thought to act as a tumor suppressor by preventing CpG to TpG mutagenesis caused by deamination of cytosine or 5-methylcytosine. Thus, several questions arise. During folate deprivation, what happens when BER is initiated at CpG islands when BER cannot be completed accurately? What is the balance between mutagenic consequences if BER is not initiated and clastogenic consequences if BER is initiated but cannot be accurately completed? Furthermore, at what point during tumorigenic transformation would folate deficiency cause BER to exert antiproliferative activity? The balance of these folate-dependent procarcinogenic and anticarcinogenic contributions of BER initiation have not been directly tested. The hypothesis to be tested is that BER initiated by uracil DNA glycosylases during folate depletion causes chromosomal instability, which is procarcinogenic to normal epithelial cells and anticarcinogenic to established adenocarcinoma cells. The studies will define the mechanism of an extremely important interaction between genes, epigenetic events, and diet in cancer prevention. It is well known that there are variations in BER capacity among humans. Knowing the cumulative status of the pathways of folate metabolism and BER should provide a better means devising better prevention and treatment strategies based on genotype. PUBLIC HEALTH RELEVANCE: Dietary folic acid is an important nutrient for human health, but there is concern that folic acid may also contribute to growth of a tumor once it has formed. Base excision repair (BER) corrects DNA damage and is generally considered to be valuable, but when folate deficiency occurs, BER may unwittingly contribute to DNA damage and cause cells to become cancerous. We will investigate the links between BER and folic acid that define the seemingly contradictory pro- and anticancer effects of each.