**2. Materials and Methods**

A representative cohort of 327 Czech patients with an unexplained etiology of cardiomyopathy underwent WES between September 2015 and June 2017. The cohort consisted mainly of cases with familial and sporadic DCM (81%), LVNC (13%), and less frequently of RCM (6%) or ACM (6%). Rare and possibly pathogenic missense *DES* variants were identified in 6 (1.8%) index patients from 6 different families (Figure 1).

**Figure 1.** (**A**) shows pedigrees of the affected families and segregation of desmin variants (+/− heterozygous status, −/− wild type). In the fourth family (with P4) we also assessed a segregation of the rare variant of *MYH7* (NM\_000257.3), c.4679G > C, p.(Arg1560Pro), which was present just in P4 (II/1) and absent in II/3, II/4, III/1, and III/2. (**B**) summarizes the structure of the desmin gene with localization of the detected variants. (**C**) illustrates the detection of desmin by western blot in myocardial samples (P2, P4, P6, control sample; 30ug protein aliquots) with an obvious reduction of signal in P4 with left ventricular non-compaction cardiomyopathy (*DES*-p.(Q364H)). Abbreviations: AC = arrhythmogenic cardiomyopathy, AVB = atrioventricular block, DCM = dilated cardiomyopathy, DES = desmin, HTx = heart transplantation, LAH = left anterior hemiblock, LVNC = left ventricular non-compaction cardiomyopathy, LVSD = left ventricular systolic dysfunction, RBBB = right bundle branch block, and RCM = restrictive cardiomyopathy.
