Here, we analyze the expression of predicted microproteins in human heart tissue and iPSC cardiomyocytes, using shot-gun proteomics and SRM. 4 identified candidates were functionally validated by interactome studies using AP-MS (affinity purification coupled to mass spectrometry). In addition, the impact of disease-causing protein-truncating variants on translation was studied in rat heart tissue by performing shot-gun proteomics.