Project III - Modeling meningomyelocele alleles and response to folic acid diet in mouse

Abstract – Project III Modeling meningomyelocele alleles and response to folic acid diet in mouse The overall hypothesis of the Program Project grant is that de novo mutations contribute to meningomyelocele (MM) risk and that folic acid (FA) influences the genome through chromatin accessibility and modulation of gene expression, to alter this risk. The mouse is considered the gold standard model for human neural tube defects (NTDs), because development is highly conserved in mammals, leading to similar types of cranial and caudal NTDs, utilizing similar genes, and responding to similar environmental factors. The mouse has been invaluable in defining genes required for neural tube closure, now numbering over 250 genes, and highlighting the need for exquisite precision of cell proliferation, actin regulation, planar cell polarity/WNT signaling, cilia/Hedgehog signaling, and FA metabolism. Many mouse NTD models show allele-specific effects, where for example, homozygous null mice are embryonic lethal, heterozygous null mice are healthy, but specific point mutations, some heterozygous, some homozygous, show NTD. In many of these mouse models, NTDs often occur in less than 100% of offspring, even in an entirely pure genetic background, reflecting background-independent partial penetrance. Like in human, the penetrance and expressivity of NTD phenotypes can be influenced by FA exposure, with about half of genes tested to date showing FA responsiveness. FA has been supplemented in grains in the US diet since 1998, regulating biosynthesis of nucleotides and S-adenosylmethionine, the universal methyl donor for DNA methylation. To account for these observations, we hypothesize that NTD risk is established by mutations in a core set of genes, with risk modified by FA-dependent changes in the epigenome. Project III has already assembled the following preliminary data: 1] Performed recessive ENU- mutagenesis screens for genes essential for neural tube closure. 2] Assembled a growing collection of genes and alleles from more than 30 NTD mutants. 3] Demonstrated mutations in primary ciliary genes underlying several mouse NTD mutants. 4] Demonstrated an impact of FA on penetrance and expressivity of murine NTDs. 5] Developed live-cell imaging platform to observe mammalian neural tube closure in real-time. 6] Modeled meningomyelocele in mouse for the human MM gene WLS. This proposal focuses on evaluating genes and alleles emerging from Project I and II as risk factors for MM, as well as the FA-dependent changes in the methylome and transcriptome that can influence this risk. Altogether, the goal of Project III is to define the relationships and interplay between mammalian MM mutations and FA influences on penetrance and expressivity. Aim 1. Model human meningomyelocele (MM) variants and genetic interactions to assess expressivity. Aim 2. Assess impact of folic acid on NTD expressivity on a gene-by-gene basis. Aim 3. Assess impact of folic acid on the neural tube at the level of single cell transcriptome, methylome and chromatin accessibility landscape.