Tetralogy of Fallot is the most common cyanotic congenital heart defect, but its genetic causes are not fully understood. While deletion of the 22q11.2 region, which includes the gene TBX1 , is a known contributor to the disease, the role of noncoding DNA variants that regulate TBX1 has remained unclear.
A research team led by scientists from Nanjing Medical University has now identified functional noncoding variants within a TBX1 enhancer that contribute to tetralogy of Fallot. The study, published in Science China Life Sciences , combined whole-genome sequencing with human stem cell-derived blood vessel organoids to investigate how these variants may affect disease development.
The researchers performed whole-genome sequencing in 428 patients diagnosed with tetralogy of Fallot. They identified 31 patients with 22q11.2 deletions involving the TBX1 genomic region and detected seven functional noncoding variants in a TBX1 enhancer. Together, these findings accounted for 42 of the 428 cases, or 9.81% of the study cohort.
To explore the biological mechanism, the team deleted the TBX1 enhancer in human embryonic stem cells and generated blood vessel organoids. These organoids provide an in vitro model for studying vessel development. Organoids lacking the TBX1 enhancer showed markedly reduced TBX1 mRNA expression, impaired angiogenesis, vessel regression, decreased pericyte coverage, and abnormal tight junction morphology.
Further analysis suggested that TBX1 acts as a transcription factor in endothelial progenitor cells, influencing their differentiation. The study identified genes involved in angiogenesis-related pathways, including DLL4 and TGFBR2 , as key downstream targets. These genes have previously been implicated in cardiac outflow tract formation.
Importantly, restoring TBX1 expression in enhancer-deficient organoids rescued the expression of downstream target genes and improved vessel features, including tight junction formation and pericyte coverage. These findings support a causal role for TBX1 enhancer disruption in vascular developmental defects relevant to tetralogy of Fallot.
The study highlights the importance of noncoding regulatory variants in congenital heart disease. It also demonstrates the value of organoid models for studying human developmental disorders, particularly when noncoding regions are difficult to evaluate using traditional animal models because of limited sequence conservation.
Science China Life Sciences
Experimental study