Impairment of LEM-domain function in LEMD2-associated cardiomyopathy

LEMD2 is an inner nuclear membrane protein that interacts with LMNA and contributes to nuclear membrane integrity and chromatin organization. Pathogenic LEMD2 variants are associated with progressive cardiomyopathy, but the relationship between Lemd2 protein abundance and disease severity remains unclear. Using computational modeling, we designed a Lemd2 in-frame deletion (p.Glu8_Leu9del, ΔE8_L9) within the LEM domain, predicted to alter its conformation, and generated knock-in (KI) mouse by CRISPR/Cas9. Western blot analysis demonstrated that the ΔE8_L9 deletion was associated with a near-complete loss of Lemd2 protein. Findings were considered alongside those previously reported for the Lemd2 p.L13R KI model, which retains ~68% of Lemd2 protein, to describe the cardiac phenotype and nuclear phenotypes associated with two distinct LEM-domain mutations. ΔE8_L9 homozygous mice exhibited postnatal growth retardation and rapidly progressive dilated cardiomyopathy (DCM) with systolic dysfunction and myocardial fibrosis by postnatal day 21 (P21), whereas the reported p.L13R model exhibited later-onset DCM. Transmission electron microscopy (TEM) revealed dense intranuclear chromatin aggregates and peripheral heterochromatin accumulation in cardiomyocytes. Bulk RNA sequencing identified downregulation of DNA replication- and DNA repair-related pathways at P1.5 and upregulation of mechanical stress-, senescence-, and immune response-related pathways at P21. Consistently, ΔE8_L9 hearts exhibited increased γ-H2AX-positive DNA damage and a numerically lower percentage of Ki-67-positive cardiomyocytes. The ΔE8_L9 model is associated with marked Lemd2 protein reduction, nuclear membrane abnormalities, altered chromatin architecture, DNA damage, and early-onset DCM.