Dysregulated ribonucleoprotein granules impair mitochondrial activity in RBM20-dilated cardiomyopathy

Pathogenic variants in the RNA-binding motif protein 20 (RBM20) cause severe dilated cardiomyopathy (DCM). While loss-of-function (LoF) variants lead to splicing dysfunction, gain-of-function (GoF) variants also mislocalize the protein to cytoplasmic ribonucleoprotein (RNP) granules resulting in a more aggressive disease phenotype with a yet unknown pathogenic mechanism. Here, we show that the induction of severe mitochondrial dysfunction is a key consequence of RBM20 mislocalization. Proteomic profiling of RBM20-GoF mouse hearts reveals a predominant reduction in mitochondrial membrane protein solubility and widespread post-transcriptional downregulation of mitochondrial proteins, including Transmembrane Protein 65 (TMEM65), a regulator of mitochondrial calcium efflux. Ultrastructural analysis shows enlarged mitochondria with aberrant morphology. Functional assays confirm impaired mitochondrial respiration, transmembrane potential, and calcium handling in RBM20-GoF, but not LoF models. Using human iPSC-derived cardioids, we validate that RBM20 mislocalization, not splicing deficiency, drives mitochondrial dysfunction in DCM. These findings establish a mechanistic link between dysregulated RNP granules and mitochondrial defects in RBM20-DCM, resembling pathogenic features observed in Fused in Sarcoma protein (FUS)-associated neurodegeneration causing amyotrophic lateral sclerosis (ALS), where cytoplasmic mislocalization of an RNA-binding protein disrupts granule dynamics and organelle function.