Homeostatic Regulation of Folate Metabolism

描述（由申请人提供）：本研究旨在阐明细胞核叶酸代谢的调控和功能，并确定其在细胞培养和敲除动物模型中对基因组完整性的贡献。维生素缺乏和/或单核苷酸多态性对叶酸代谢的破坏影响AdoMet和dTMP的合成，从而影响DMA的染色质甲基化和尿嘧啶含量。尿嘧啶和甲基化都会影响DNA的稳定性，甲基化调节许多基因的表达。目前尚不清楚叶酸代谢与病理（包括某些癌症、心血管疾病）和发育异常（包括神经管缺陷）之间的关联是否源于AdoMet和/或dTMP合成的改变。最近，我们发现胞浆丝氨酸羟甲基转移酶（cSHMT）是一种代谢开关，指导叶酸活化的碳单元在dTMP和AdoMet生物合成途径之间的通量。cSHMT在与叶酸相关病变相关的组织中表达，包括发育中的神经管和结肠；它的表达受多种营养物质的动态调控。因此，cSHMT可能有助于叶酸相关病理的病因学和通过饮食预防的目标。最近，我们发现cSHMT在细胞周期的S期定位于细胞核，并被SUMO修饰。这些实验旨在在cSHMT表达和sumoylation与核叶酸代谢之间，以及核叶酸代谢与基因组完整性之间进行直接和连贯的翻译。阐明核叶酸代谢的功能将增加我们对叶酸相关病理的理解。具体目的是：1 .确定核叶酸代谢的存在和影响其效率的因素。2。以确定人类cSHMT L474F SNP对细胞核叶酸代谢的影响。3。以确定cSHMT表达降低影响TS蛋白水平的机制。IV.确定叶酸缺乏诱导肠内cSHMT蛋白水平的机制。要检验的主要假设是：1。cSHMT通过与TS和DHFR的核共定位促进dTMP合成。2. sumo酰化对细胞核叶酸代谢至关重要。3. 核叶酸代谢增强基因组稳定性。4. TS和cSHMT的表达是协调调节的。5. 叶酸/胆碱对cSHMT的调控保留了dTMP库，但牺牲了AdoMet的合成。该项目的长期目标是：1。定义细胞调节叶酸代谢的稳态机制。

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.