Generation of a biallelic NRAP-knockout mutant from a human iPSC line

Abstract

Cardiomyopathies, a leading cause of mortality, are associated with dysfunctional intercalated discs, which connect neighbouring cardiomyocytes and ensure proper contractility. In human cardiac diseases, loss-of-function mutations of the intercalated disc-associated protein Nebulin-Related Anchoring Protein (NRAP) have been reported. NRAP plays a crucial role in myofibril assembly and mechanotransduction, however, its regulatory functions remain unclear. To investigate the effects of NRAP loss-of-function in cardiac disease, a human induced pluripotent stem cell (hiPSC) line was generated carrying a biallelic NRAP-knockout (KO) using the CRISPR-Cas9 technology. Control and mutant cell lines were assessed for karyotype integrity, pluripotency, off-target effects, mycoplasma contamination, and differentiation into ectoderm, mesoderm, and endoderm. This hiPSC line provides a valuable tool to study how NRAP modulates cardiac function and contributes to disease progression.