Cellular and Molecular Analysis of Body Wall Closure

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Mammalian ventral body wall closure defects, including gastroschisis, thoracoabdominoschisis, and omphalocele are linked with the morphogenesis of the ventral body wall, the amnion, and the umbilical ring. Malformations of the ventral body wall comprise one of the leading categories of human birth defects and are present in about one out of every 2000 live births. Although the occurrence of these defects is relatively common, there is a surprising lack of knowldege concerning the development and closure of the ventral body wall in mouse or human. This field is further complicated by the array of theories on the pathogenesis of human body wall defects that have not been experimentally tested. This proposal aims to produce a paradigm shift in our understanding of mammalian ventral body closure that can provide a mechanistic framework and comprehensive resource for future understanding of how this process goes awry because of genetic and/or environmental causes. The transcription factor AP-2, encoded by the gene Tfap2a, has an essential role in mouse body wall closure and previous studies have shown that it regulates ectodermal, mesenchymal, peripheral nervous system, and mesodermal interactions that drive development of the ventral body wall. Tfap2a null mice have a severe form of ventral closure defect, a thoracoabdominoschisis, in which the ventral covering of the chest and abdomen fails to form so that the heart, lungs, liver, and gut are exposed. Critically, this is one of the few simple mouse models that gives a fully penetrant and consistent body wall closure defect that can be used to understand how this important developmental process can fail. In the first Aim, relevant tissue will be collected from control and Tfap2a null mice at specific embryonic times and subjected to single cell RNA seq (scRNAseq) analysis. This will identify the normal gene expression patterns of mouse body wall closure as well as cell types and genes that are impacted by loss of Tfap2a. Several additional genes are known to affect body wall closure and we will be able to attribute these genes to relevant tissue populations to understand how they may be influencing this critical developmental process. In the second Aim, we will perform scATACseq analysis on all control time points as well as the most relevant time points for the mutant. We will then integrate all the scRNAseq and scATACseq data to identify the tissues, cell types, and gene expression signatures that are normally associated with body wall closure. Further, we will identify how chromatin accessibility in the body wall is altered by loss of Tfap2a. Verification analyses will be performed to identify critical changes in cell populations and gene expression profiles associated with normal and abnormal body wall closure. Subsequently, the information from these studies will be synthesised into a novel and powerful model for normal body wall closure that can serve as an important framework for future understanding of this important class of human birth defect.