Homeostatic Regulation of Folate Metabolism

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DESCRIPTION (provided by applicant): This research aims to elucidate the regulation and function of nuclear folate metabolism and determine its contribution to genome integrity in both cell culture and knock-out animal models. Disruption of folate metabolism by vitamin deficiency and/or single nucleotide polymorphisms affects AdoMet and dTMP syntheses and thereby influences chromatin methylation and uracil content in DMA. Both uracil and methylation affect DNA stability, and methylation regulates the expression of many genes. It is not known if the associations between folate metabolism and pathologies (including certain cancers, cardiovascular disease) and developmental anomalies (including neural tube defects) result from altered AdoMet and/or dTMP syntheses. Recently, we showed that the enzyme cytoplasmic serine hydroxymethyltransferase (cSHMT) is a metabolic switch that directs the flux of folate-activated one-carbon units between the dTMP and AdoMet biosynthetic pathways. cSHMT is expressed in tissues associated with folate-related pathologies including the developing neural tube and colon; its expression is dynamically regulated by several nutrients. Therefore, cSHMT may contribute to the etiology of folate-related pathologies and a target for prevention through diet. Recently, we discovered that cSHMT localizes to the nucleus during the S phase of the cell cycle and that it modified by SUMO. These experiments are designed to make direct and coherent translations between cSHMT expression & sumoylation and nuclear folate metabolism, and between nuclear folate metabolism and genome integrity. Elucidating the function of nuclear folate metabolism will increase our understanding of folate-related pathologies. The specific aims are: I. to determine the existence of nuclear folate metabolism and the factors that influence its efficiency. II. to determine the impact of the human cSHMT L474F SNP on nuclear folate metabolism. III. to determine the mechanisms whereby decreases in cSHMT expression affect TS protein levels. IV. to determine the mechanism whereby folate deficiency induces cSHMT protein levels in the intestine. The principle hypotheses to be tested are that: 1. cSHMT enhances dTMP synthesis through nuclear co-localization with TS & DHFR. 2. sumoylation is essential for nuclear folate metabolism. 3. nuclear folate metabolism enhances genome stability. 4. TS and cSHMT expression are coordinately regulated. 5. regulation of cSHMT by folate/choline preserves dTMP pools at the expense of AdoMet synthesis. The long-term goals of this project are: 1. to define homeostatic mechanisms used bv cells to regulate folate metabolism.