Arrhythmogenic cardiomyopathy (ACM) is an inherited myocardial disease at risk of sudden death. Genetic testing impacts greatly in ACM diagnosis, but gene-disease associations have yet to be determined for the increasing number of genes included in clinical panels. Genetic variants evaluation was undertaken for the most relevant non-desmosomal disease genes. We retrospectively studied 320 unrelated Italian ACM patients, including 243 cases with predominant right-ventricular (ARVC) and 77 cases with predominant left-ventricular (ALVC) involvement, who did not carry pathogenic/likely pathogenic (P/LP) variants in desmosome-coding genes. The aim was to assess rare genetic variants in transmembrane protein 43 (TMEM43), desmin (DES), phospholamban (PLN), filamin c (FLNC), cadherin 2 (CDH2), and tight junction protein 1 (TJP1), based on current adjudication guidelines and reappraisal on reported literature data. Thirty-five rare genetic variants, including 23 (64%) P/LP, were identified in 39 patients (16/243 ARVC; 23/77 ALVC): 22 FLNC, 9 DES, 2 TMEM43, and 2 CDH2. No P/LP variants were found in PLN and TJP1 genes. Gene-based burden analysis, including P/LP variants reported in literature, showed significant enrichment for TMEM43 (3.79-fold), DES (10.31-fold), PLN (117.8-fold) and FLNC (107-fold). A non-desmosomal rare genetic variant is found in a minority of ARVC patients but in about one third of ALVC patients; as such, clinical decision-making should be driven by genes with robust evidence. More than two thirds of non-desmosomal P/LP variants occur in FLNC.

Arrhythmogenic cardiomyopathy (ACM) is a cardiomyopathy that is predominantly inherited and characterized by cardiac arrhythmias and structural abnormalities. TMEM43 (transmembrane protein 43) is one of the well-known genetic culprits behind ACM. In this study, we successfully generated an induced pluripotent stem cell (iPSC) line, YCMi010-A, derived from a male patient diagnosed with ACM. Although these iPSCs harbored a heterozygous intronic splice variant, TMEM43 c.443-2A > G, they still displayed normal cellular morphology and were confirmed to express pluripotency markers. YCMi010-A iPSC line is a promising model for investigating the pathomechanisms associated with ACM and exploring potential therapeutic strategies.

Arrhythmogenic cardiomyopathy is an inherited cardiomyopathy that can involve both ventricles. Several genes have been identified as pathogenic in arrhythmogenic cardiomyopathy, including TMEM43. However, there are limited data on cardiac MRI findings in patients with TMEM43 variants to date. In this case series, cardiac MRI findings and clinical outcomes are described in 14 patients with TMEM43 variants, including eight (57%) with the pathogenic p.Ser358Leu variant (six female patients; mean age, 33 years ± 15 [SD]) and six (43%) with a TMEM43 variant of unknown significance (three female patients; mean age, 38 years ± 11). MRI findings demonstrated left ventricular systolic dysfunction in eight (57%) patients and right ventricular dysfunction in four (29%) patients. Among the nine patients with late gadolinium enhancement imaging, left ventricular late gadolinium enhancement was present in seven (78%; all subepicardial) patients. In summary, TMEM43 variants are associated with high prevalence of subepicardial late gadolinium enhancement and left ventricular dysfunction. Keywords: Arrhythmogenic Cardiomyopathy, Arrhythmogenic Right Ventricular Cardiomyopathy, TMEM43, Cardiac MRI, Genetic Variants Supplemental material is available for this article.

Arrhythmogenic right ventricular cardiomyopathy (ARVC) is a genetic disorder of desmosomal and structural proteins that is characterized by fibro-fatty infiltrate in the ventricles and fatal arrhythmia that can occur early before significant structural abnormalities. Most ARVC mutations interfere with β-catenin-dependent transcription that enhances adipogenesis; however, the mechanistic pathway to arrhythmogenesis is not clear. We hypothesized that adipogenic conditions play an important role in the formation of arrhythmia substrates in ARVC. Cardiac myocyte monolayers co-cultured for 2-4 days with mesenchymal stem cells (MSC) were derived from human-induced pluripotent stem cells with the ARVC5 TMEM43 p.Ser358Leu mutation. The TMEM43 mutation in myocyte co-cultures alone had no significant effect on impulse conduction velocity (CV) or APD. In contrast, when co-cultures were exposed to pro-adipogenic factors for 2-4 days, CV and APD were significantly reduced compared to controls by 49% and 31%, respectively without evidence of adipogenesis. Additionally, these arrhythmia substrates coincided with a significant reduction in IGF-1 expression in MSCs and were mitigated by IGF-1 treatment. These findings suggest that the onset of enhanced adipogenic signaling may be a mechanism of early arrhythmogenesis, which could lead to personalized treatment for arrhythmias associated with TMEM43 and other ARVC mutations.

Arrhythmogenic cardiomyopathy (ACM) is an inherited heart muscle disease caused by heterozygous missense mutations within the gene encoding for the nuclear envelope protein transmembrane protein 43 (TMEM43). The disease is characterized by myocyte loss and fibro-fatty replacement, leading to life-threatening ventricular arrhythmias and sudden cardiac death. However, the role of TMEM43 in the pathogenesis of ACM remains poorly understood. In this study, we generated cardiomyocyte-restricted transgenic zebrafish lines that overexpress eGFP-linked full-length human wild-type (WT) TMEM43 and two genetic variants (c.1073C>T, p.S358L; c.332C>T, p.P111L) using the Tol2-system. Overexpression of WT and p.P111L-mutant TMEM43 was associated with transcriptional activation of the mTOR pathway and ribosome biogenesis, and resulted in enlarged hearts with cardiomyocyte hypertrophy. Intriguingly, mutant p.S358L TMEM43 was found to be unstable and partially redistributed into the cytoplasm in embryonic and adult hearts. Moreover, both TMEM43 variants displayed cardiac morphological defects at juvenile stages and ultrastructural changes within the myocardium, accompanied by dysregulated gene expression profiles in adulthood. Finally, CRISPR/Cas9 mutants demonstrated an age-dependent cardiac phenotype characterized by heart enlargement in adulthood. In conclusion, our findings suggest ultrastructural remodeling and transcriptomic alterations underlying the development of structural and functional cardiac defects in TMEM43-associated cardiomyopathy.

Arrhythmogenic right ventricular cardiomyopathy (ARVC) is a severe cardiac disease that leads to heart failure or sudden cardiac death (SCD). For the pathogenesis of ARVC, various mutations in at least eight different genes have been identified. A rare form of ARVC is associated with the mutation TMEM43 p.S358L, which is a fully penetrant variant in male carriers. TMEM43 p.S358 is homologous to CG8111 p.S333 in Drosophila melanogaster. We established CRISPR/Cas9-mediated CG8111 knock-out mutants in Drosophila, as well as transgenic fly lines carrying an overexpression construct of the CG8111 p.S333L substitution. Knock-out flies developed normally, whereas the overexpression of CG8111 p.S333L caused growth defects, loss of body weight, cardiac arrhythmias, and premature death. An evaluation of a series of model mutants that replaced S333 by selected amino acids proved that the conserved serine is critical for the physiological function of CG8111. Metabolomic and proteomic analyses revealed that the S333 in CG8111 is essential to proper energy homeostasis and lipid metabolism in the fly. Of note, metabolic impairments were also found in the murine Tmem43 disease model, and fibrofatty replacement is a hallmark of human ARVC5. These findings contribute to a more comprehensive understanding of the molecular functions of CG8111 in Drosophila, and can represent a valuable basis to assess the aetiology of the human TMEM43 p.S358L variant in more detail.

BACKGROUND: Transmembrane protein 43 (TMEM43), a member of the transmembrane protein subfamily, plays a critical role in the initiation and development of cancers. However, little is known concerning the biological function and molecular mechanisms of TMEM43 in pancreatic cancer.
METHODS: In this study, TMEM43 expression levels were analyzed in pancreatic cancer samples compared with control samples. The relationship of TMEM43 expression and disease-free survival (DFS) and overall survival (OS) were assessed in pancreatic cancer patients. In vitro and in vivo assays were performed to explore the function and role of TMEM43 in pancreatic cancer. Coimmunoprecipitation (co-IP) followed by protein mass spectrometry was applied to analyze the molecular mechanisms of TMEM43 in pancreatic cancer.
RESULTS: We demonstrated that TMEM43 expression level is elevated in pancreatic cancer samples compared with control group, and is correlated with poor DFS and OS in pancreatic cancer patients. Knockdown of TMEM43 inhibited pancreatic cancer progression in vitro, decreased the percentage of S phase, and inhibited the tumorigenicity of pancreatic cancer in vivo. Moreover, we demonstrated that TMEM43 promoted pancreatic cancer progression by stabilizing PRPF3 and regulating the RAP2B/ERK axis.
CONCLUSIONS: The present study suggests that TMEM43 contributes to pancreatic cancer progression through the PRPF3/RAP2B/ERK axis, and might be a novel therapeutic target for pancreatic cancer.

The TMEM43 has been studied in human diseases such as arrhythmogenic right ventricular cardiomyopathy type 5 (ARVC5) and auditory neuropathy spectrum disorder (ANSD). In the heart, the p.(Ser358Leu) mutation has been shown to alter intercalated disc protein function and disturb beating rhythms. In the cochlea, the p.(Arg372Ter) mutation has been shown to disrupt connexin-linked function in glia-like supporting cells (GLSs), which maintain inner ear homeostasis for hearing. The TMEM43-p.(Arg372Ter) mutant knock-in mice displayed a significantly reduced passive conductance current in the cochlear GLSs, raising a possibility that TMEM43 is essential for mediating the passive conductance current in GLSs. In the brain, the two-pore-domain potassium (K2P) channels are generally known as the \"leak channels\" to mediate background conductance current, raising another possibility that K2P channels might contribute to the passive conductance current in GLSs. However, the possible association between TMEM43 and K2P channels has not been investigated yet. In this study, we examined whether TMEM43 physically interacts with one of the K2P channels in the cochlea, KCNK3 (TASK-1). Utilizing co-immunoprecipitation (IP) assay and Duolink proximity ligation assay (PLA), we revealed that TMEM43 and TASK-1 proteins could directly interact. Genetic modifications further delineated that the intracellular loop domain of TMEM43 is responsible for TASK-1 binding. In the end, gene-silencing of Task-1 resulted in significantly reduced passive conductance current in GLSs. Together, our findings demonstrate that TMEM43 and TASK-1 form a protein-protein interaction in the cochlea and provide the possibility that TASK-1 is a potential contributor to the passive conductance current in GLSs.

Dilated cardiomyopathy (DCM) is an umbrella term entailing a wide variety of genetic and non-genetic etiologies, leading to left ventricular systolic dysfunction and dilatation, not explained by abnormal loading conditions or coronary artery disease. The clinical presentation can vary from asymptomatic to heart failure symptoms or sudden cardiac death (SCD) even in previously asymptomatic individuals. In the last 2 decades, there has been striking progress in the understanding of the complex genetic basis of DCM, with the discovery of additional genes and genotype-phenotype correlation studies. Rigorous clinical work-up of DCM patients, meticulous family screening, and the implementation of advanced imaging techniques pave the way for a more efficient and earlier diagnosis as well as more precise indications for implantable cardioverter defibrillator implantation and prevention of SCD. In the era of precision medicine, genotype-directed therapies have started to emerge. In this review, we focus on updates of the genetic background of DCM, characteristic phenotypes caused by recently described pathogenic variants, specific indications for prevention of SCD in those individuals and genotype-directed treatments under development. Finally, the latest developments in distinguishing athletic heart syndrome from subclinical DCM are described.

High-level exercise has been associated with a malignant phenotype in desmosomal and genotype-negative forms of arrhythmogenic right ventricular cardiomyopathy (ARVC). This is the first study to examine this issue with ARVC secondary to the TMEM43 p.S358L mutation.
The purpose of this study was to evaluate the impact of exercise on arrhythmic risk and cardiac death in TMEM43 p.S358L ARVC.
Individuals with the TMEM43 p.S358L mutation enrolled in a prospective registry who had received a primary prevention implantable cardioverter-defibrillator (ICD) were invited to complete the modified Paffenbarger Physical Activity Questionnaire to assess their physical activity in the year before their ICD implantation. Time-to-event analyses using unadjusted and adjusted Cox proportional hazards models evaluated associations between physical activity and first appropriate ICD discharge secondary to malignant ventricular arrhythmia or cardiac death.
In 80 subjects with the TMEM43 p.S358L mutation, exercise ≥9.0 metabolic equivalent of task (MET)-hours/day (high level) in the year before ICD implantation was associated with an adjusted 9.1-fold increased hazard of first appropriate ICD discharge (there were no deaths) relative to physical activity <9.0 MET-hours/day (moderate level) (95% confidence interval [CI] 3.3-24.6 MET-hours/day; P < .001). The median age from birth to first appropriate ICD discharge was 58.5 years (95% CI 56.5-60.5 years) vs 35.8 years (95% CI 28.2-43.4 years) (P < .001) in subjects in moderate- and high-level exercise groups, respectively.
Exercise ≥9.0 MET-hours/day is associated with an increased risk of malignant ventricular arrhythmias in the TMEM43 p.S358L subtype of ARVC. Extrapolating these data, we suggest molecular testing be offered in early childhood to inform exercise choices reflective of the genotype.