Mechanisms of mitochondrial folate metabolism in neural tube closure

描述(由申请人提供)：神经管缺陷(NTDS)是人类最常见的出生缺陷之一，被认为是由多因素引起的。改变NTD易感性的最强联系之一是母亲的叶酸状况。然而，这些叶酸依赖过程背后的生化机制尚不清楚。我们知识上的这种差距阻碍了我们就叶酸强化和预防NTDS和其他人类疾病做出明智的卫生政策决定的能力，这些疾病既包括先天性疾病，也包括晚年发生的疾病。我的长期目标是了解叶酸支持正常神经管发育的机制，以及叶酸代谢改变如何导致NTDS的发展。该建议的目的是确定一种特定的叶酸依赖酶(线粒体Mthfd1)缺失导致NTDS的机制(S)。我的中心假设是，在Mthfd1无合子小鼠中观察到的NTDS是由线粒体叶酸依赖的一碳(1C)代谢缺陷引起的，线粒体叶酸依赖的一碳(1C)代谢为必要的过程提供1C单位，如从头开始的嘌呤、胸腺嘧啶酸、甘氨酸和甲基生物合成。这项研究的基本原理是，Mthfd1小鼠模型提供了一个独特的机会来发现叶酸依赖正常神经管发育的特定代谢机制(S)。更重要的是，更好的机械学理解可能会导致叶酸强化或其他治疗干预的新的和创新的方法，以努力减少或预防人类的NTDS。我将测试我的中心假设， 从而通过一个特定的目标来实现这一建议的目标：识别导致Mthfd1无合子胚胎神经管缺陷的生化缺陷。在这一目标下，我将使用生化分析方法来分析正常(+/+)、杂合子(+/-)和空合子(-/-)胚胎以及来自这三种Mthfd1基因的胚胎干细胞(ES细胞)中叶酸依赖的代谢过程。这一目标的预期结果是确定在Mthfd1无合子胚胎中观察到的NTDS的特定代谢机制。在我看来，本申请中提出的研究是创新的，因为它专注于一种新的小鼠NTD模型(Mthfd1l基因敲除)，该模型紧密复制了人类NTD的表型，并且不需要额外的营养干预来表达疾病的表型。此外，人类Mthfd1的一种常见变异已被证明与某些人群中NTDS风险的增加有关。这一贡献是重要的，因为识别特定的代谢机制将从根本上促进对叶酸反应性NTD的理解，并将为非叶酸反应性NTD提供亟需的新见解。在我们评估当前叶酸强化计划在降低人类NTDS患病率方面的有效性和安全性时，这一详细的机制信息将是必不可少的。

DESCRIPTION (provided by applicant): Neural tube defects (NTDs), among the most common birth defects in humans, are believed to have multifactorial causes. One of the strongest links to modifying NTD susceptibility is to maternal folic acid status. However, the biochemical mechanisms that underlie these folate-dependent processes are not understood. This gap in our knowledge hinders our ability to make informed health policy decisions about folic acid fortification and prevention of NTDs and other human diseases, both congenital and those occurring later in life. My long term goal is to understand the mechanisms by which folic acid supports normal neural tube development, and how altered folate metabolism leads to the development of NTDs. The objective of this proposal is to identify the mechanism(s) by which loss of a specific folate-dependent enzyme (mitochondrial MTHFD1L) leads to NTDs. My central hypothesis is that the NTDs observed in the Mthfd1l nullizygous mouse are caused by defects in mitochondrial folate-dependent one-carbon (1C) metabolism, which supplies 1C units for essential processes such as de novo purine, thymidylate, glycine, and methyl group biosynthesis. The rationale for this research is that the Mthfd1l mouse model provides a unique opportunity to discover the specific metabolic mechanism(s) that underlie the folate dependence of normal neural tube development. More importantly, a better mechanistic understanding is likely to lead to new and innovative approaches to folate fortification or other therapeutic interventions in the effort to reduce or prevent NTDs in humans. I will test my central hypothesis, and thereby accomplish the objective of this proposal, through one Specific Aim: Identify the biochemical defects responsible for neural tube defects in Mthfd1l nullizygous embryos. Under this aim, I will use biochemical assays to analyze the folate-dependent metabolic processes in normal (+/+), heterozygous (+/-), and nullizygous (-/-) embryos and in embryonic stem (ES) cells derived from the three Mthfd1l genotypes. The expected outcome of this aim is the identification of specific metabolic mechanisms responsible for the NTDs observed in Mthfd1l nullizygous embryos. The research proposed in this application is innovative, in my opinion, because it focuses on a new mouse NTD model (Mthfd1l knockout) that closely replicates the human NTD phenotype, and does not require additional nutritional intervention to express the disease phenotype. Moreover, a common variant of human Mthfd1l has been shown to be associated with increased risk of NTDs in some populations. This contribution is significant because identification of specific metabolic mechanisms will fundamentally advance understanding of folate-responsive NTDs, and will provide much needed new insight into non-folate-responsive NTDs as well. This detailed mechanistic information will be essential as we evaluate the efficacy and safety of the current folic acid fortification program in reducing the prevalence of human NTDs.