Discovering and Modeling Genetic Causes of Congenital Heart Diseas

Congenital heart disease (CHD) is the most common birth defect and affects around 2% of live births, when including bicuspid aortic valve (BAV). Although surgical care has significantly improved patient outcome, it remains a major contributor to morbidity and mortality. Population and family based studies have identified a strong genetic component to CHD, however the exact etiology by which CHD occurs is not well understood. Clinically, there is a lack of genetic diagnostic tools available for these patients. While whole exome and whole genome sequencing offer potential benefits to this patient population, it has not yet been utilized or tested in a clinical setting. Additionally, pipelines have not yet been developed to systematically analyze the large sequencing datasets that these approaches produce.;As genomic testing continues to become more accessible to clinicians and patients, it is crucial that pipelines are developed that allow for a streamlined clinical testing approach. We propose the utilization of whole exome sequencing, with a candidate gene prioritization approach, to allow for the identification of causative mutations in familial CHD. We performed whole exome sequencing on 9 families with Mendelian inherited CHD and prioritized the variants utilizing a CHD gene list and further filtered based on segregation, rarity, and predicted pathogenicity. This approach was successful in the identification of potentially pathogenic variants in 3 of the 9 families, and included mutations in GATA4, TLL1, and MYH11. This work supports the use of clinical whole exome sequencing in familial cases of CHD, and offers a pipeline for a streamlined approach to identify high-quality, disease causing mutations. Identifying disease associated variants is the first step toward understanding the underlying role of genetics in CHD. However, to determine causality and to determine mechanism one must utilize a model system, allowing for the manipulation of the gene of interest and a subsequent disease readout. We utilized this approach, with a mutation in GATA4 previously identified in a family with atrial septal defects (ASD) and partially penetrant pulmonary valve stenosis, and characterized disease development and molecular deficits underlying this phenotype. Mice were utilized that harbored the orthologous G295S disease-causing mutation and recapitulated the human disease phenotype, exhibiting both ASDs and semilunar valve stenosis. We hypothesized that the GATA4 G295S mutation leads to semilunar valve stenosis due to abnormal valve development. Echocardiographic and histologic examination of adult GATA4 G295Ski/wt mice identified functional semilunar valve stenosis, leaflet thickening, and severe disorganization of extracellular matrix (ECM) proteins. To determine disease onset, 3D-reconstruction was performed on histologic sections of the developing embryonic valves. A reduction in valve leaflet volume was discovered at embryonic day (E)13.5 and morphologic abnormalities were apparent by E15.5. To examine the molecular basis for this phenotype, we performed RNA-seq analysis of E15.5 semilunar valve tissue and identified enrichment in the dysregulation of pathways representing ECM organization and WNT signaling. These findings demonstrate a novel role for GATA4 in semilunar valve development and disease through the utilization of a new mouse model for congenital semilunar valve stenosis.