Open Access Thesis
Date of Award
Nutrition and Food Science
Diane C. Cabelof
Folate is a water-soluble vitamin B that plays a critical co-enzyme in the de novo nucleotide synthesis and other biochemical processes including DNA metabolism, DNA repair, DNA methylation, and cellular growth. Folate deficiency has been associated to increase the risk of neural tube defects (NTDs) and cancers of the lung, breast, colon, cervix, esposphagus and brain. Most importantly, folate deficiency has been shown to increase uracil misincorporation into DNA and therefore induce DNA damage repaired by the base excision repair (BER) pathway. In response to folate depletion, levels of thmidylate decrease in the deoxyribonucelotide pool, resulting in uracil being misincorporation into DNA instead of thymine during replication and repair. Uracil misincorporation into DNA is believed to be the biological mechanism of how folate affects carcinogenesis. We evaluated the impact of folate depletion on uracil accumulation, BER activity and UDG activity in folate depleted UNG+/+ and UNG-/- mouse embryonic fibroblasts (MEFs). Additionally, cell growth in response to folate deficiency was also determined by completing doubling time in folate depleted UNG+/+ and UNG-/- cells. As expected, levels of uracil significantly increased in UNG+/+ and UNG-/- cells in response to folate depletion. Uracil-DNA glycosylase (Udg) activity, which is responsible for the removal of uracil during the base excision (BER) pathway, significantly decreased in folate depleted UNG-/- cells. Such decreases in Udg activity corresponded to the decrease in nuclear UDG protein levels in response to folate deficiency. Similarly, BER capacity significantly (p<0.001) decreased in response to folate depletion in UNG+/+ cells, suggesting that folate deficiency inhibits BER in mouse embryonic fibroblasts.
Doko, Eneida, "Uracil accumulation in folate depleted mouse embryonic fibroblasts" (2012). Wayne State University Theses. 173.