Intricate MIB1-NOTCH-GATA6 Interactions in Cardiac Valvular and Septal Development.

Genome-wide association studies and experimental mouse models implicate the MIB1 and GATA6 genes in congenital heart disease (CHD). Their close physical proximity and conserved synteny suggest that these two genes might be involved in analogous cardiac developmental processes. Heterozygous Gata6 loss-of-function mutations alone or humanized Mib1 mutations in a NOTCH1-sensitized genetic background cause bicuspid aortic valve (BAV) and a membranous ventricular septal defect (VSD), consistent with MIB1 and NOTCH1 functioning in the same pathway. To determine if MIB1-NOTCH and GATA6 interact in valvular and septal development, we generated compound heterozygote mice carrying different Mib1 missense ( Mib1 K735R and Mib1 V943F ) or nonsense ( Mib1 R530X ) mutations with the Gata6 STOP/+ heterozygous null mutation. Combining Mib1 R530X/+ or Mib1 K735R/+ with Gata6 STOP/+ does not affect Gata6 STOP/+ single mutant phenotypes. In contrast, combining Mib1 V943F/+ with Gata6 STOP/+ decreases the incidence of BAV and VSD by 50%, suggesting a suppressive effect of Mib1 V943F/+ on Gata6 STOP/+ . Transcriptomic and functional analyses revealed that while the EMT pathway term is depleted in the Gata6 STOP/+ mutant, introducing the Mib1 V943F variant robustly enriches this term, consistent with the Mib1 V943F/+ phenotypic suppression of Gata6 STOP/+ . Interestingly, combined Notch1 and Gata6 insufficiency led to a nearly fully penetrant VSD but did not affect the BAV phenotype, underscoring the complex functional relationship between MIB1, NOTCH, and GATA6 in valvular and septal development.