BPS2026 – Molecular and structural characterization of a pediatric dilated cardiomyopathy mouse model bearing TNNC1-I4M variant

Dilated cardiomyopathy (DCM) is the most common form of cardiomyopathy in children and is frequently caused by genetic mutations in sarcomeric proteins that disrupt cardiac contractility. Variants in TNNC1, encoding cardiac troponin C (cTnC), have been affirmative to cause DCM, yet the mechanisms by which these variants alter cardiac function remain poorly understood. A de novo TNNC1 I4M variant—located in the unique N-helix of cTnC—was identified in a pediatric patient presenting with a heart murmur at birth, progressing to severe DCM by six weeks of age. We developed a heterozygous knock-in mouse model via CRISPR-Cas9 to introduce the I4M mutation into the endogenous TNNC1 locus. Multi-level characterization was performed across organ, tissue, cellular, proteomic, and transcriptomic scales. At 4 weeks of age, echocardiography revealed DCM features, including left ventricular (LV) dilation and reduced systolic function, which corresponded with altered calcium transients and sarcomere contractility in isolated unloaded cTnC-I4M cardiomyocytes relative to wild-type (WT). Absolute Quantitative LC-MS/MS of myofibril fraction and whole-cell lysates showed ∼60% cTnC-I4M incorporation in the LV while mRNA transcript showed ∼50% mutant mRNA. Single-nucleus RNA sequencing of LV myocardium revealed transcriptional remodeling across cardiac cell populations. Therapeutically, administration of danicamtiv, significantly improved ejection fraction in cTnC-I4M mice to comparable levels as WT. Additionally, acute dobutamine stimulation confirmed that β-adrenergic responsiveness remained intact. Histological analysis showed absence of fibrosis, consistent with early-stage cardiac remodeling. To further explore the molecular basis of impaired contractility, high-resolution X-ray diffraction indicated increased interfilament spacing and I1,1/I1,0 equatorial intensity ratio in permeabilized cardiac muscle bundles in relaxing solution. Altogether, these results shed light on how TNNC1 variants impair contractile function and drive remodeling, highlighting the utility of integrated molecular and structural analyses in understanding pediatric DCM mechanisms.