Single-nucleus RNA sequencing of TMEM43 induced arrhythmogenic cardiomyopathy reveals altered cell compositions and transcriptional states in mice

Abstract Introduction and purpose TMEM43 induced Arrhythmogenic cardiomyopathy, also referred to as arrhythmogenic cardiomyopathy type 5 or ACM5, is caused by a point mutation in TMEM43 (p.S358L). The disease is characterised by chamber dilation and a progressive dystrophy of the ventricular myocardium causing life-threatening ventricular arrhythmias and sudden cardiac death. Loss of cardiomyocytes and fibrofatty replacement of the myocardium is clearly observed in our transgenic mouse model of ACM5 overexpressing TMEM43-S358L in postnatal cardiomyocytes. However, the mechanisms underlying cardiomyocyte death and fibroblast activation remain elusive, precluding the development of effective therapeutic options. Here we applied the state-of-the-art analysis on single-nucleus RNA sequencing (snRNA-seq) to characterise the cellular compositions and molecular states in ACM5 mouse hearts. Methods Left ventricular heart tissues of severely affected ACM5 mice and their corresponding transgenic controls overexpressing wild-type TMEM43 were used for snRNA-seq analysis. We identified 10 major cell types, which aggregated into 23 distinct clusters on the basis of transcriptional similarity. Results ACM5 nuclei showed significant depletion of cardiomyocytes and expanded populations of fibroblasts and myeloid cells. Fibroblasts from mutant mice acquired an activated phenotype related to remodelling of the extracellular matrix with a large number of genes like LOX, POSTN, NOX4 or COL1A1 significantly upregulated. In addition, the mutant cardiomyocytes showed altered transcriptional states characterized by cardiac stress and heart failure markers such as NPPB or MYH7. Interestingly, we were able to identify a cluster of mutant cardiomyocytes with a highly apoptotic profile. This cluster is also enriched in genes related to the unfolding protein response (UPR) stress sensors and the DNA damage response (DDR), elucidating a new potential mechanism of myocyte death in ACM5. Furthermore, heterogeneity of the myeloid populations is also observed. The number of tissue resident macrophages is increased in mutant samples and myeloid cells show additional disease-associated transcriptional states that are almost entirely absent in control samples. Conclusion Together these data illuminated shared and distinct cell and molecular architectures of ACM5 in mouse hearts. Further confirmation on diverse gene expression profiles like the UPR and DDR pathways related to cardiomyocyte death will establish a valuable resource to explore novel therapeutic targets and personalized therapies for ACM5.