WNT5a-Mediated Aberrant Actin Filament Dynamics Drive Cardiac Pathogenic Phenotypes in <i>LMNA</i>-Related Emery-Dreifuss Muscular Dystrophy.

Abstract

Emery-Dreifuss muscular dystrophy (EDMD) is a rare genetic disorder characterized by early-onset joint contractures, progressive muscle atrophy, and cardiac abnormalities. Patients with EDMD carrying <i>LMNA</i> sequence variations often exhibit severe cardiac manifestations, including frequent atrioventricular block and ventricular tachycardia. Approximately 20% of those patients may ultimately require heart transplantation. The molecular mechanisms by which <i>LMNA</i> sequence variations lead to EDMD remain unknown. Five clinically diagnosed patients with EDMD carrying <i>LMNA</i> sequence variations were recruited. Patient-specific induced pluripotent stem cells (iPSCs) were generated using a nonintegrating Sendai virus. Previously generated iPSCs, derived from 2 healthy donors, were used as controls. The <i>LMNA</i> L204P sequence variation was corrected by genome editing in EDMD iPSC lines to generate isogenic controls. All iPSC-derived cardiomyocytes (iPSC-CMs) were generated using a monolayer-based differentiation protocol. Three-dimensional, strip-format, and force-generating human engineered heart tissues were generated from iPSC-CMs. A knock-in mouse model carrying the <i>Lmna</i> L204P sequence variation was also generated. EDMD-specific iPSC-CMs exhibited a variety of deleterious phenotypes, including disorganized sarcomeres, abnormal nuclear envelope structure, arrhythmias, and contractile dysfunction, when compared with control and gene-corrected iPSC-CMs. Multi-omics analysis further revealed that <i>LMNA</i> directly binds the <i>WNT5A</i> promoter and the Leu204Pro sequence variation reduces chromatin accessibility and <i>WNT5A</i> transcription in EDMD iPSC-CMs. WNT5a (Wnt family member 5a)/RhoA (Ras homolog family member A) signaling inactivation was shown to lead to actin depolymerization and inhibition of actin polymerization in EDMD iPSC-CMs. This results in a deformed nuclear envelope, contractile dysfunction, and impaired trafficking of Cx43 (connexin 43). The impairment of Cx43 trafficking causes reduced distribution of Cx43 at cell-cell borders, contributing to the arrhythmic phenotype in EDMD iPSC-CMs. Pharmacological interventions of exogenous WNT5a supplementation, RhoA activator, or an actin polymerization stabilizer effectively rescued the pathogenic phenotypes of EDMD iPSC-CMs. EDMD engineered heart tissues displayed dysfunctional contractile force generation, which was significantly alleviated by RhoA activator. <i>Lmna</i> L204P heterozygous knock-in mice exhibited impaired cardiac function and developed cardiac arrhythmias in response to sympathetic stress. We present WNT5a-mediated aberrant actin filament dynamics as a novel mechanism underlying cardiac pathogenic phenotypes in <i>LMNA</i>-related EDMD. Our findings indicate that activating WNT5a/RhoA and stabilizing actin assembly may serve as novel therapeutic strategies for this condition.