Young and early‐onset dilated cardiomyopathy with malignant ventricular arrhythmia and sudden cardiac death induced by the heterozygous LDB3, MYH6, and SYNE1 missense mutations

Abstract Background The whole exome sequencing (WES) with targeted gene analysis is an effective diagnostic tool for cardiomyopathy. The early‐onset sudden cardiac death (SCD) was commonly associated with dilated cardiomyopathy (DCM) induced by pathogenic genetic mutations. Methods In a Chinese Han family, the patient of 24 years old occurred with early‐onset and DCM and died of SCD associated with ICD storms induced by repetitive ventricular tachycardia/fibrillation (VT/F). Genomic DNA samples of peripheral blood were conducted for WES and Sanger sequence. Then, we performed bioinformatics analysis for 200 genes susceptible to cardiomyopathies and arrhythmias. Further, we analyzed how the potential pathogenic mutations affecting the secondary structure, hydrophobicity, and phosphorylation of amino acids, protein properties, and their joint pathogenicity by ProtParam, SOPMA, and ORVAL algorisms. The protein–protein interaction was analyzed by STRING algorism. Results The mutations of LDB3 p.M456R, MYH6 p.S180Y, and SYNE1 p.S4607F were identified as “Damaging/Deleterious.” The SYNE1 (p.S4607F) increased one of alpha helix and decreased one of beta sheet. The LDB3 (p.M456R) reduced one of beta sheet and increased one of beta turn. The MYH6 (p.S180Y) decreased two of beta sheets and four of beta turns, but significantly increased twelve coils. The hydrophobicity of amino acid residues and their adjacent sequences were decreased by LDB3 (p.M456R) and MYH6 (p.S180Y), and significantly increased by SYNE1 (p.S4607F). The mutations of LDB3 (p.M456R), SYNE1 (p.S4607F), and MYH6 (p.S180Y) resulted in the phosphorylation changes of the corresponding amino acid sites or the nearby amino acid sites. The pairwise combinations of LDB3, MYH6, and SYNE1 mutations have the high probability of causing disease, especially the highest probability for SYNE1 and LDB3 mutations. There was obviously indirect interaction of the proteins encoded by SYNE1, LDB3, and MYH6. Conclusions The multiple heterozygous mutations of SYNE1, LDB3, and MYH6 may be associated with young and early‐onset of DCM and SCD.

DCM is complicated by incomplete penetrance, high variability in age of onset and disease progression, and high genetic heterogeneity (Haas et al., 2015). Cardiomyopathies are associated with >100 known genes, including genes encoding for sodium and potassium channel in sarcolemma, dystrophin proteins in cytoplasm, myofibril with sarcomere/Z-disk, thin/thick filament, RNA-binding protein, cytoskeletal proteins, desmosome mitochondria, sarcoplas-  (Giglietal.,2019;Lietal.,2020).Here,weaimedtoidentifynovel mutations and genes responsible for young and early-onset DCM with SCD using WES. In doing so, we identified multiple potential pathogenic mutations of LDB3, MYH6, and SYNE1 genes in this patient.

| Myocardial biopsy
The right internal jugular vein was punctured, and a 7-Fr vascular sheath was inserted. A long, flexible catheter with forceps in the head end was inserted into a vein and threaded up into the heart while monitored by X-ray. Five small snips of muscle at the apical septum of the right ventricle were grabbed. Some cardiac tissues, whichwerefixedwithformalinsolutions,thendehydrated,paraffinembeddedandsliced,wereusedforHEandMassonstaining.Some cardiactissueswerefixedwithglutaraldehydeandosmicacid,dehydrated,resinembeddedandsliced,andthenpreparedforuranyl acetate and lead nitrate staining and analyzed using a transmission electron microscope.

| Pathological features of myocardium
The pathological examination showed that some muscle fibers of myocardial tissue were atrophied and reduced in volume, some muscle fibers were hypertrophic; the nucleus of myocardium was large and deeply stained, and obvious interstitial fiber hyperplasia wasfound(Figure1b).Theintercalateddiskofcardiomyocyteswas notconnected(Figure1c-e),andthelocalspacewaswidened.The sizesofmitochondriaweredifferent,andpartofmitochondriawas slightly focal vacuolar degeneration. The local myocardial fibrils were dissolved and disappeared, and many glycogen particles were accumulatedandfilled.

| Genetic screening
In this family (Figure 1a), a set of candidate genes associated with cardiomyopathies and arrhythmias were screened using the WES data of II: 1. The results (

| Study limitations
Furtheranimalorcellexperimentsareneededtoconfirmthemechanisms how these three mutations abnormally change the cardiac structureandfunctionandresultintheDCMandSCD.

ACK N OWLED G M ENTS
We thank Professor Shulin Wu, Yumei Xue, and Fang Rao from Guangdong Cardiovascular Institute. We appreciate Jinsheng Tao andZhipengCaoforassistanceindataanalysisandvisualization.

CO N FLI C T O F I NTE R E S T
The authors declare that they have no competing interest.

DATA AVA I L A B I L I T Y S TAT E M E N T
The data used in this study are not publicly available, but it might be available from the corresponding author upon reasonable request andpermissionfromrelevantChineseAuthorities.