Our results, using a new mouse model mimicking an acute reduction of cMLCK, suggest that cMLCK plays a pivotal role in the transition from compensated to decompensated hypertrophy via sarcomeric disorganization.

Background Heterozygous human mutations of NKX2-5 are highly penetrant and associated with varied congenital heart defects. The heterozygous knockout of murine Nkx2-5, in contrast, manifests less profound cardiac malformations, with low disease penetrance. We sought to study this apparent discrepancy between human and mouse genetics. Since missense mutations in the NKX2-5 homeodomain (DNA binding domain) are the most frequently reported type of human mutation, we replicated this genetic defect in a murine knock-in model. Methods and Results We generated a murine model in a 129/Sv genetic background by knocking-in an Nkx2-5 homeodomain missense mutation previously identified in humans. The mutation was located at homeodomain position 52Arg→Gly (R52G). All the heterozygous neonatal Nkx2-5+/R52G mice demonstrated a prominent trabecular layer in the ventricular wall, so called noncompaction, along with diverse cardiac anomalies, including atrioventricular septal defects, Ebstein’s malformation of the tricuspid valve, and perimembranous and/or muscular ventricular septal defects. In addition, P10 Nkx2-5+/R52G mice demonstrated atrial septal anomalies, with significant increase in the size of the inter-atrial communication and fossa ovalis, and decrease in the length of the flap valve compared to control Nkx2-5+/+ or Nkx2-5+/− mice. Conclusion The results of our study demonstrate that heterozygous missense mutation in the murine Nkx2-5 homeodomain (R52G) are highly penetrant, and result in pleiotropic cardiac effects. Thus, in contrast to heterozygous Nkx2-5 knockout mice, the effects of the heterozygous knock-in mimic findings in humans with heterozygous missense mutation in NKX2-5 homeodomain.

Background Heterozygous human NKX2-5 homeodomain (DNA binding domain) missense mutations are highly penetrant for varied congenital heart defects, including progressive atrioventricular block (AVB) requiring pacemaker implantation. We recently replicated this genetic defect in a murine knock-in model, in which we demonstrated highly penetrant, pleiotropic cardiac anomalies. In this study, we examined postnatal AV conduction in the knock-in mice. Methods and Results A murine knock-in model (Arg52Gly, Nkx2-5+/R52G) in a 129/Sv background was analyzed by histopathology, surface and telemetry ECG, and in vivo electrophysiology studies (EPS), comparing with control Nkx2-5+/+ mice at diverse postnatal stages, ranging from postnatal day 1 (P1) to 17 months. PR-prolongation (1st degree AVB) was present at 4 weeks, 7 months, and 17 months of age but not at P1 in the mutant mice. Advanced AVB was also occasionally demonstrated in the mutant mice. EPS showed that AV nodal function, and right ventricular effective refractory period, were impaired in the mutant mice, while sinus nodal function was not affected. AV nodal size was significantly smaller in the mutant mice compared to their controls at 4 weeks of age, corresponding to the presence of PR-prolongation, but not P1, suggesting, at least in part, that the conduction abnormalities are the result of a morphologically atrophic AV node. Conclusions The highly penetrant and progressive AVB phenotype seen in human heterozygous missense mutations in NKX2-5 homeodomain was replicated in mice by knocking-in a comparable missense mutation.