Malene Lindholm PhD, Han Zhu MD, Yong Huang, Euan Ashley MD, PhD, Matthew Wheeler MD, PhD.
Stanford Cardiovascular Institute, School of Medicine, Stanford University, Stanford, CA
In cardiac and skeletal muscle, alpha-actinins are critical cytoskeletal proteins that anchor actin filaments within the sarcomere. Mutations in ACTN2 have been associated with cardiac abnormalities. However, the mechanisms behind how ACTN2 mutations lead to cardiac dysfunction remain poorly understood. The aim of this study was to investigate the effects of two novel ACTN2 mutations on cardiac and skeletal muscle phenotypes in human tissue and patient-specific iPSC-derived cardiomyocytes.
We identified patients in the Stanford Center for Inherited Cardiovascular Disease database with rare or novel ACTN2 variants using a custom mutation pipeline optimized for rare variant discovery. We identified one patient homozygous for a stop-gain mutation (p.Q860X) in ACTN2 and a family with an exon 8-10 deletion. In heart transplant tissue of the homozygous patient, we observed mild hypertrophy and interstitial fibrosis. There was no variation in ACTN2 protein expression, indicating absence of nonsense mediated decay. siRNA knock down of ACTN2 in neonatal rat ventricular cardiomyocytes and a human myoblast cell line resulted in dramatic changes in cell size and morphology. Patient-derived iPSC-cardiomyocytes were hypertrophic, displayed sarcomeric structural disarray and had a slower contractile velocity. Using Co-Immunoprecipitation for ACTN2, followed by mass-spectrometry, we identified a missing protein-protein interaction with AKAP9 in the patient with the truncated ACTN2 variant.
The molecular effects of ACTN2 on a cellular level and how it causes cardiomyopathy has not been fully elucidated. Here, we provide evidence that two loss of function genetic variants in ACTN2 are associated to contractile dysfunction and lead to cardiac abnormalities.