Uncovering the cellular and molecular mechanisms of folic acid fortification in neural tube defects

总结 这项建议的目的是阐明叶酸如何防止神经管缺陷的细胞和 分子水平，并开发一个平台，以功能验证和测试人NTD的FA反应性 候选基因神经管缺陷是第二常见的先天性畸形， 在美国的10，000例妊娠中。未能关闭神经管，大脑和脊髓的前身 脊髓灰质炎是一种由遗传和环境压力引起的复杂疾病。因为多种因素影响着 这种疾病的发展，关键是要了解环境因素如何与遗传学相互作用， NTD的病因。而全民叶酸强化使NTD的总体发生率降低了28%， 在美国，叶酸是如何预防NTD的仍不清楚。我的建议旨在解决两个假设， 叶酸和神经管缺陷首先，我假设FA通过作用于细胞而有助于NTD的预防 运动水平以及诱导转录级联以补偿失调的基因。到 为了揭示FA是否可以通过在细胞或分子水平上起作用来拯救NT关闭，我将描述FA的特征。 补充FA的Alx 1突变小鼠模型的细胞运动和转录谱。的 Alx 1基因敲除模型导致100%的突变胚胎发生NTD，然而产前FA补充可以 将发病率降至13%。Alx 1突变体是分析FA强化作用的一个易于处理的模型 恢复NT闭合，因为该模型中的NTD是由转录因子的失调引起的， 异常的细胞运动我们将使用基于成像的技术来表征细胞形态发生 方法和scRNA-Seq和scATAC-Seq，以鉴定Alx 1-null中FA诱导的基因表达 背景这些补充研究的结果将告知FA是否通过支持细胞运动而起作用， 诱导转录活性或其组合。其次，我假设测试更多的基因突变， FA反应性将揭示从FA强化作为预防策略中获益的基因类型模式。 为了解决这个假设，我将开发一个基于课程的本科生研究经验（CURE）， 什么，为什么，以及如何神经管缺陷。本课程的科学目标是从功能上 验证候选人NTD基因在鸡胚NT闭合中的未知意义，并测试叶酸 酸反应CURE的社会目标是增加相关研究机会的数量 为本科生，同时满足公共卫生需要，筛选候选基因，并传达回 临床医生和家庭。我希望筛选更多NTD和FA反应性的候选基因， 新的高风险基因，并增加我们对可能反应的失调基因类型的预测能力。 补充FA。总之，这项建议的结果将对加强我们对以下问题的理解至关重要： FA如何预防神经管缺陷并改善神经管缺陷预防策略的风险评估。

SUMMARY The objective of this proposal is to elucidate how folic acid prevents neural tube defects on the cellular and molecular level, and to develop a platform to functionally validate and test the FA responsiveness of human NTD candidate genes. Neural tube defects (NTDs) are the second most common congenital abnormality, affecting 6 in 10,000 pregnancies in the United States. Failure to close the neural tube, the precursor to the brain and spinal cord, is complex disorder resulting from genetic and environmental stressors. Because multiple factors play into the development of this disorder, it is critical to understand how environmental factors interact with genetics in the etiology of NTDs. While population-wide folic acid (FA) fortification has reduced the overall NTD rate by 28% in the US, it remains unclear how folic acid prevents NTDs. My proposal aims to address two hypotheses about folic acid and neural tube defects. First, I hypothesize that FA contributes to NTD prevention by acting at the cell movement level as well as inducing transcriptional cascades to compensate for the dysregulated genes. To uncover whether FA can rescue NT closure by acting at the cellular or molecular level, I will characterize the cellular movements and transcriptional profiles for the Alx1 mutant mouse model supplemented with FA. The Alx1 knockout model results in NTDs in 100% of mutant embryos, however prenatal FA supplementation can reduce the incidence to 13%. The Alx1 mutant is a tractable model to dissect the contribution of FA fortification to restoring NT closure because NTDs in this model result from dysregulation of a transcription factor and abnormal cellular movements. We will use imaging-based techniques to characterize cell morphogenesis processes and scRNA-Seq and scATAC-Seq to identify FA-induced gene expression in the Alx1-null background. Results from these complimentary studies will inform if FA acts by supporting cellular movements, inducing transcriptional activity, or a combination. Second, I hypothesize that testing more gene mutations for FA responsiveness will reveal patterns of gene types that benefit from FA fortification as a prevention strategy. To address this hypothesis, I will develop a course-based undergraduate research experience (CURE) entitled The What, the Why, and the How of neural tube defects. The scientific aim of this course will be to functionally validate candidate human NTD genes of unknown significance in NT closure in chick embryos and test the folic acid responsiveness. The societal aim of the CURE is to increase the number of relevant research opportunities for undergraduates while meeting a public health need to screen candidate genes and communicate back to clinicians and families. I expect screening more candidate genes for NTDs and FA responsiveness will reveal new high-risk genes and increase our predictive power over the types of dysregulated genes likely responsive to FA supplementation. Together, the results from this proposal will be important to bolster our understanding of how FA can prevent NTDs and improve risk assessment of preventative strategies against neural tube defects.