Dysregulated Ribonucleoprotein Granules Impair Mitochondrial Function in <i>RBM20</i>-Related Dilated Cardiomyopathy.

Abstract

Pathogenic variants in <i>RBM20</i> cause severe dilated cardiomyopathy. Loss-of-function variants disrupt splicing; neomorphic gain-of-function (GoF) variants also mislocalize RBM20 to cytoplasmic ribonucleoprotein granules and are associated with more aggressive disease. The mechanism by which RBM20 mislocalization drives cardiac dysfunction remains unknown. We investigated the effects of <i>Rbm20</i> GoF and loss-of-function (LoF) variants using proteomic profiling, protein solubility assays, mitochondrial respiration and calcium flux analyses, and ultrastructural imaging in mouse models. Human induced pluripotent stem cell-derived cardioids were used to validate variant-specific phenotypes. <i>Rbm20</i> GoF, but not LoF, variants caused posttranscriptional downregulation of soluble mitochondrial proteins, including the calcium efflux regulator TMEM65 (transmembrane protein 65), and reduced solubility of mitochondrial membrane proteins. Electron microscopy revealed enlarged mitochondria with cristae disorganization. Functional assays confirmed impaired oxidative phosphorylation, reduced mitochondrial membrane potential, and abnormal calcium handling in <i>Rbm20</i> GoF models. Human cardioids reproduced these findings, demonstrating that cytoplasmic mislocalization, rather than splicing deficiency, drives mitochondrial dysfunction. Cytoplasmic mislocalization of RBM20 disrupts mitochondrial function by reducing mitochondrial protein abundance, leading to oxidative phosphorylation failure and abnormal mitochondrial calcium handling. This mechanism distinguishes <i>RBM20</i> GoF from LoF variants and may explain the more severe heart failure phenotype observed in patients with <i>RBM20</i> GoF variants. These insights advance the mechanistic understanding of <i>RBM20</i>-related cardiomyopathy and identify mitochondrial mRNA/protein regulation as a key node in cardiac energetics.