IACTN2/I Mutant Causes Proteopathy in Human iPSC-Derived Cardiomyocytes

• Published in: Cells (Basel, Switzerland) Vol. 11; no. 17
• Main Authors: Zech, Antonia T. L, Prondzynski, Maksymilian, Singh, Sonia R, Pietsch, Niels, Orthey, Ellen, Alizoti, Erda, Busch, Josefine, Madsen, Alexandra, Behrens, Charlotta S, Meyer-Jens, Moritz, Mearini, Giulia, Lemoine, Marc D, Krämer, Elisabeth, Mosqueira, Diogo, Virdi, Sanamjeet, Indenbirken, Daniela, Depke, Maren, Salazar, Manuela Gesell, Völker, Uwe, Braren, Ingke, Pu, William T, Eschenhagen, Thomas, Hammer, Elke, Schlossarek, Saskia, Carrier, Lucie
• Format: Journal Article
• Language: English
• Published: MDPI AG 01-09-2022
• Subjects: Cellular proteins; Genetic aspects; Health aspects; Heart cells; Mutation (Biology); Stem cells; Germany
• ISSN: 2073-4409; 2073-4409
• DOI: 10.3390/cells11172745

Genetic variants in α-actinin-2 (ACTN2) are associated with several forms of (cardio)myopathy. We previously reported a heterozygous missense (c.740C>T) ACTN2 gene variant, associated with hypertrophic cardiomyopathy, and characterized by an electro-mechanical phenotype in human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs). Here, we created with CRISPR/Cas9 genetic tools two heterozygous functional knock-out hiPSC lines with a second wild-type (ACTN2wt) and missense ACTN2 (ACTN2mut) allele, respectively. We evaluated their impact on cardiomyocyte structure and function, using a combination of different technologies, including immunofluorescence and live cell imaging, RNA-seq, and mass spectrometry. This study showed that ACTN2mut presents a higher percentage of multinucleation, protein aggregation, hypertrophy, myofibrillar disarray, and activation of both the ubiquitin-proteasome system and the autophagy-lysosomal pathway as compared to ACTN2wt in 2D-cultured hiPSC-CMs. Furthermore, the expression of ACTN2mut was associated with a marked reduction of sarcomere-associated protein levels in 2D-cultured hiPSC-CMs and force impairment in engineered heart tissues. In conclusion, our study highlights the activation of proteolytic systems in ACTN2mut hiPSC-CMs likely to cope with ACTN2 aggregation and therefore directs towards proteopathy as an additional cellular pathology caused by this ACTN2 variant, which may contribute to human ACTN2-associated cardiomyopathies.