We first performed solubility profiling in RBM20-WT, -P635L+/+ and -R636Q+/+ mice. We lysed murine ventricular tissues using either a mild detergent (NP-40) to extract only the soluble fraction of the proteome, or a strong detergent (SDS) to extract total proteomes, within the same experiment. Solubility is quantified as a proportion of the protein amount identified in the soluble fraction divided by the protein amount in the total fraction. The solubility values of proteins interacting within the same cellular compartment are then expected to be similar. For this experiment, we used N = 3 mice per genotype, with two fractions (total and soluble) for each mouse. We then perfomred proteomics analysis of only soluble proteomes, to gain a deeper understanging of the mechanism causing the downregulation of soluble proteins in RBM20-mutant mice. For that, we lysed left ventricules of RBM20-WT, -P635L+/+ and -R636Q+/+ mice, total N = 6 mice per genotype, using mild detergent (NP-40). In addition, to assess whether our observed findings are specific to these gain-of-fucntion variants, we compared soluble lysates of RBM20-WT and RBM20-KO (Loss-of-function, LoF model) mice, N=4 mice per genotype. All mice for all experiments were between 14-17 weeks of age. To validate our findings in a human caridac model, we performed analyses of total (lysed with SDS) proteomes of human iPSC-derived cardiac organoids with RBM20-WT, -P633L, -R634Q, and -KO, N = 3 biological replicates each.