Desmosomes in the Heart: A Review of Clinical and Mechanistic Analyses

Abstract

Desmosomes have long been appreciated as intercellular junctions that are vital for maintaining the structural integrity of stratified epithelia. More recent clinical investigations of patients with diseases such as arrhythmogenic cardiomyopathy have further highlighted the importance of desmosomes in cardiac tissue, where they help to maintain coordination of cardiac myocytes. Here, we review clinical and mechanistic studies that provide insight into the functions of desmosomal proteins in skin and heart during homeostasis and in disease. While intercellular junctions are organized differently in cardiac and epithelial tissues, studies conducted in epithelial systems may inform our understanding of cardiac desmosomes. We explore traditional and non-traditional roles of desmosomal proteins, ranging from adhesive capacities to nuclear functions. Finally, we discuss how these studies can guide future investigations focused on determining the molecular mechanisms by which desmosomal mutations promote the development of cardiac diseases.

Keywords::

• desmosome
• area composita
• arrhythmogenic cardiomyopathy

INTRODUCTION: CLINICAL ANALYSIS DRAWS ATTENTION TO CARDIAC DESMOSOMES

In 1986, a clinical report emerged describing a group of patients in the Greek island of Naxos with familial palmoplantar keratoderma and hair abnormalities (Protonotarios et al., 1986). Unlike previous reports of epidermal diseases, the majority of these patients were also found to have symptoms or signs of cardiac disease. Patients experienced ventricular tachycardias and were found to have enlarged cardiac ventricles, with one case resulting in sudden cardiac death. In 1996 and 1998, two reports described similar findings of woolly hair, palmoplantar keratoderma, and dilated cardiomyopathy (DCM) in patients in India and Ecuador (Carvajal-Huerta, 1998; Rao et al., 1996).

A major step toward understanding molecular mechanisms underlying these cardiocutaneous syndromes, respectively termed Naxos disease and Carvajal syndrome, was taken upon the discovery that many cases were associated with mutations in proteins of intercellular junctions termed desmosomes, reviewed in detail earlier in this series (Harmon & Green, 2013). Naxos disease was mapped to a 2-base-pair deletion in the gene encoding plakoglobin (Pg) causing a truncation of its C-terminus (McKoy et al., 2000), whereas Carvajal syndrome was mapped to a truncation in the gene encoding desmoplakin (DP) (Norgett et al., 2000).

The emergence of these clinical syndromes sparked a number of reports demonstrating the importance of desmosomal proteins to the structure and function of cardiac tissue. Perhaps most notable are those describing arrhythmogenic cardiomyopathy (AC), a cardiac disease characterized by fibro-fatty replacement of healthy myocardium leading to arrhythmias and in some cases, sudden cardiac death (Basso et al., 2011; Herren et al., 2009). The majority of mutations linked to AC are autosomal dominant and target desmosomal proteins; thus, AC has been dubbed a “disease of the desmosome.” Naxos disease and Carvajal syndrome are considered to be autosomal recessive variants of AC.

While much is yet to be elucidated regarding the roles of desmosomal proteins in the heart, insights can be drawn from studies conducted in epithelial models, where the roles of desmosomes in regulating mechanical integrity and cell signaling are better understood (Simpson et al., 2011). We review those studies here and highlight potential differences between desmosomal proteins in heart and skin that should be accounted for when analyzing the functions of cardiac desmosomes.

CARDIAC DESMOSOMES: A COMPONENT OF THE AREA COMPOSITA

As reviewed earlier in this series (Harmon & Green, 2013), desmosomes are a type of adhesive intercellular junction appearing as electron-dense plaques at regions of cell–cell contact. Desmosomes are composed of proteins from three gene families: cadherins, armadillo proteins, and plakins. Desmosomal cadherins (Desmoglein, Dsg; Desmocollin, Dsc) link desmosomes of opposing cells through their ectodomains, while their cytoplasmic tails connect to desmosomal armadillo proteins (Plakoglobin, Pg; Plakophilin, PKP). These armadillo proteins enable the desmosomal plaque protein desmoplakin (DP) to tether intermediate filaments (IFs) to sites of cell–cell contact (Figure 1A). While complex stratified tissues such as epidermis express multiple cadherin and armadillo isoforms, cardiac tissue isoforms are limited to Dsg2, Dsc2, and PKP2, which along with Pg and DP anchor the IF desmin to the cardiac myocyte plasma membrane (Bolling & Jonkman, 2009).

Figure 1. Intercellular junctions in epithelial and cardiac tissues. (A) In epithelial tissues, adherens junctions and desmosomes comprise discrete plaques that respectively tether actin and intermediate filaments to sites of cell–cell contact (Harmon & Green, 2013; Simpson et al., 2011). (B) In cardiac tissue, components of desmosomes and adherens junctions intermingle as a hybrid area composita that stabilizes adjacent gap junctions (Bolling & Jonkman, 2009; Franke et al., 2006). Desmoplakin may localize to N-cadherin via plakoglobin (Cowin et al., 1986), while αT-catenin could associate with desmosomal cadherins through plakophilin 2 (Goossens et al., 2007). Both desmocollin 2 and plakophilin 2 have been shown to associate with connexin43 (Gehmlich et al., 2011a; Li et al., 2009; Oxford et al., 2007), potentially forming direct links between the area composita and gap junction plaques.

In epithelial cells, desmosomes are discrete structures that tether IF keratins to sites of cell–cell contact, whereas adhesive adherens junctions anchor actin filaments (Simpson et al., 2011). Cardiac desmosomes and adherens junctions, in addition to gap junctions that provide open channels for electrical communication, are traditionally described as discrete components of intercalated discs connecting adjacent cardiac cells. However, more recent studies have revealed that in higher organisms, components of desmosomes and adherens junctions actually intermingle to form a hybrid area composita (Franke et al., 2006) (Figure 1B). This finding necessitates careful consideration of functional differences that may exist for these junctional proteins in different tissue types.

Cardiac muscle architecture

Cardiac tissue is a complex three-dimensional network of cardiac myocytes interspersed with endothelial cells, fibroblasts, and vascular smooth muscle (Harvey & Leinwand, 2011). Cardiac myocytes must maintain mechanical and electrical connections for proper cardiac contraction. Desmosomes and adherens junctions are well known for maintaining structural connections, while gap junctions and ion channels facilitate electrical communication between cells (Simpson et al., 2011).

Disruption of these networks can produce a variety of cardiac diseases (Elliott, 2008; Harvey & Leinwand, 2011; Maron et al., 2006). Irregular heartbeats associated with conditions such as arrhythmias can stem from post-ischemic scar tissue, which disrupts electrical coordination of cardiac cell networks. Cardiomyopathies, or diseases of cardiac muscle, are associated with a wide range of inherited or acquired factors (Table 1). Many cases of cardiomyopathy are linked to mutations of sarcomeric proteins internal to each cardiac myocyte, which are vital for actin–myosin contraction. AC is associated with mutations in desmosomal proteins, though the extent to which cardiomyopathy arises from a loss of desmosome-based structural coordination or other less understood signaling changes is unclear. The potential bases for AC will be further discussed in this review.

Table 1. Cardiomyopathy (CM) classifications. Generalized clinical findings and etiology of each condition, in addition to those of unclassified cardiomyopathies (left ventricular noncompaction, stress-induced, cirrhotic, etc.) are further described in referenced reviews (Elliott, 2008; Harvey & Leinwand, 2011; Maron et al., 2006).

Classification	Tissue characteristics	Common etiologies
Arrhythmogenic CM	Fibro-fatty replacement of ventricular muscle, sparing interventricular septum.	Mutations in desmosomal proteins. Non-familial forms may be linked to inflammation.
Dilated CM	Dilatation and impaired contraction of ventricles, most significantly in the left ventricle.	Idiopathic or secondary to systemic disorders (i.e. infection or systemic toxins). Familial forms due to mutations in cytoskeletal, sarcomeric, nuclear, or junctional proteins.
Hypertrophic CM	Ventricular hypertrophy (involves interventricular septum) with disorganization of cardiac myocytes.	Mutations in sarcomeric proteins; also linked to metabolic disorders.
Restrictive CM	Increased stiffness of myocardium (without alterations in myocyte arrangement), causing elevated ventricular pressures.	Idiopathic or secondary to systemic disorders/injury to cardiac tissue. Familial forms due to mutations in sarcomeric proteins.

Desmosomes in the heart

From 2006 to 2008, Werner Franke and colleagues published a series of papers describing the discovery of the area composita (Borrmann et al., 2006; Franke et al., 2006, 2007; Pieperhoff et al., 2008, 2010; Pieperhoff & Franke, 2007, 2008). Previously, it had been thought that desmosomes and adherens junctions represented discrete intercellular junctions, with the exception of Pg having dual localization to both junction types. Upon closer examination, electron microscopy demonstrated that immunogold-labeled desmosomal components (DP, Pg, and PKP2) could localize to over 85% of total cell–cell contact area, including regions resembling adherens junctions (Franke et al., 2006). Thus, desmosomes and adherens junctions are proposed to comprise a hybrid area composita.

Subsequent studies elucidated the timing of area composita development (Pieperhoff & Franke, 2007). During early stages of embryogenesis, localization of DP is mutually exclusive with the adherens junction component N-cadherin. These proteins begin to converge at E13, though desmosomal cadherins do not co-localize with N-cadherin until birth. Coalescence of adherens junctions and desmosomes progressively increases from 6 days until 1 year after birth. Non-mammalian vertebrates do not exhibit this mixing of junctions, suggesting that the development of the area composita is a late process of vertebrate evolution (Pieperhoff et al., 2010).

Crosstalk between intercellular junctions

A few studies have provided clues as to how desmosomal and adherens junctions components intermingle (Figure 1B). Pg is known to localize to both desmosomal and classical cadherins (Cowin et al., 1986), providing a mechanism by which DP could localize to adherens junctions. In fact, Pg has been shown to form an association between DP and VE-cadherin in endothelial cells (Kowalczyk et al., 1998). The desmosomal armadillo protein PKP2 interacts with αT-catenin (Goossens et al., 2007), explaining a potential association of actin with desmosomes.

While the area composita in itself refers to a mixing of proteins of adhesive junctions, additional studies have established interactions of these proteins with components of cardiac electrical machinery. Both Dsc2 and PKP2 have been found to co-immunoprecipitate with the gap junction component connexin 43 (Cx43) (Gehmlich et al., 2011a; Li et al., 2009; Oxford et al., 2007); in fact, PKP2 can be found to localize inside gap junction plaques (Agullo-Pascual et al., 2013). Subsequent studies demonstrated that PKP2 and Dsg2 can associate with and regulate the activity of the cardiac voltage-gated sodium channel Na_v1.5 (Rizzo et al., 2012; Sato et al., 2009). This interplay of desmosomal proteins with electrical machinery may be coordinated by the cytoskeletal adaptor protein ankyrin-G (Sato et al., 2011) and could potentially contribute to crosstalk between gap junctions and sodium channel complexes (Desplantez et al., 2012; Jansen et al., 2012). These interactions provide a potential explanation for the observed electrophysiological abnormalities in disorders associated with mutations in desmosomal components, including Naxos disease, Carvajal syndrome, and AC (Basso et al., 2011; Kaplan et al., 2004a, 2004b).

AC: A DISEASE OF THE DESMOSOME

Historical perspective

The first descriptions of AC may have occurred as early as the mid-1700s, when a physician in Rome described a family with palpitations, dilatation and aneurysms of the right ventricle, and sudden death recurring over four generations (Lancisi, 1736). The more widely recognized clinical descriptions associated with AC emerged in the late 1970s when Fontaine and colleagues described a condition characterized by replacement of cardiac musculature with fatty and fibrous tissue (Frank et al., 1978; Marcus et al., 1982). At later stages of disease, patients were noted to have enlarged hearts with paper-thin right ventricles. This morphology led clinicians to associate the disease with Uhl's anomaly in which the right ventricular myocardium fails to develop during embryogenesis (Uhl, 1952). Due to the belief that the disease was a developmental abnormality, this condition was initially referred to as right ventricular (RV) dysplasia. Fontaine and colleagues proposed a subset of this condition when associated with arrhythmias: arrhythmogenic RV dysplasia (ARVD).

Upon further investigation, however, it was recognized that associated structural defects were not present at birth, but instead represented progressive damage of the myocardium. Together with the growing recognition of arrhythmias as a common finding of the disease, the classification of the condition was changed from a type of dysplasia to a type of cardiomyopathy: arrhythmogenic right ventricular cardiomyopathy (ARVC) (Basso et al., 2009, 2011). The World Health Organization recognized ARVC as a distinct class of cardiomyopathy in 1995 (Richardson et al., 1996).

While myocardial damage is most typically found in the right ventricle, involvement of the left ventricle has been observed in as high as half of reported cases (Basso et al., 2009). Left ventricular involvement may be associated with later stages of disease progression (Corrado et al., 1997). However, in many cases morphological changes occur predominantly or solely in the left ventricle (Bauce et al., 2005; Norman et al., 2005; Okabe et al., 1995; Sen-Chowdhry et al., 2008; Suzuki et al., 2000). While the factors dictating ventricular localization are unknown, these cases have once again prompted a renaming of the disease. Use of ARVC and ARVD are still common, but the disease is increasingly being referred to, broadly, as AC (Basso et al., 2011).

Clinical findings

Patients with AC commonly present in the second to fourth decades of life with syncope, palpitations, and/or arrhythmias resulting in depolarization/repolarization abnormalities on electrocardiogram (Basso et al., 2009). AC more frequently affects men than women. Diagnosis of AC is based on cardiac structural alterations, tissue characterization of ventricular walls, repolarization abnormalities, depolarization/conduction abnormalities, arrhythmias, and family history (Marcus et al., 2010; McKenna et al., 1994). Athletes with AC are at a higher risk of triggering an arrhythmic event leading to sudden cardiac death, potentially due to acute volume overload and stretching of the right ventricle caused by strenuous activity (Basso et al., 2009; Corrado et al., 2006b).

Compared to other cardiac diseases, AC is considered to be a rare form of cardiomyopathy. Still, AC has a prevalence of up to 1:1000 in certain regions of Europe and 1:5000 in the United States. Further, AC is likely to be under-recognized, remaining unnoticed throughout a patient's life or resulting in sudden cardiac death (Basso et al., 2009, 2011; Corrado et al., 2011). This mortality highlights the importance of elucidating disease pathogenesis and improving disease detection/diagnosis. The original AC diagnostic criteria formulated in 1994 were recently revised in an effort to improve sensitivity for detection of early stages and familial forms of AC (Marcus et al., 2010). Screening programs in Italy, in which competitive athletes undergo clinical examination and take prescreening questionnaires to assess the risk of cardiovascular events, have led to a significant decline in sudden cardiac death in athletes aged 12–35 years (Corrado et al., 2006a). The implementation of such programs worldwide will likely result in similar declines in mortality (Corrado et al., 2008), though variable penetrance of AC mutations necessitates more rigorous criteria to determine if it is beneficial to limit sports activity for these patients (Corrado et al., 2006b).

Histologic and molecular characteristics

On histologic examination, AC is associated with replacement of myocardial cells with fatty and fibrous tissue (Basso et al., 2009; Herren et al., 2009; Marcus et al., 1982). This infiltrate typically begins near the epicardium and progressively extends into the endocardium, becoming transmural but leaving the interventricular septum intact. Wall thinning is found to be most severe in the right ventricular “triangle of dysplasia” (anterior infundibulum, right ventricular apex, and inferior aspect of the right ventricle), though the posterolateral left ventricle may be more readily involved than the right ventricular apex (Te Riele et al., 2013). These regions are vulnerable to aneurysm formation from continuous cycles of mechanical stress. While the presence of fatty tissue may be nonspecific, a degenerative change of myocytes with replacement fibrosis is indicative of AC.

AC is hereditary in up to 50% of patients, and studies over the past 15 years have identified mutations in the desmosomal proteins Dsg2, Dsc2, Pg, PKP2, and DP as being responsible for the vast majority of these cases (Table 2). Thus, AC is often referred to as a disease of the desmosome (Basso et al., 2009; Herren et al., 2009; Rickelt & Pieperhoff, 2012). Mutations in PKP2 appear to be most frequent and are associated with founder groups in regions of the Netherlands (Basso et al., 2009; Cox et al., 2011). However, mutations in any of these proteins may be non-pathogenic polymorphisms, and variable disease penetrance may impede predictions of phenotypic outcome based on genetics. The online ARVC database (http://www.arvcdatabase.info) is a valuable tool that uses in silico prediction models and clinical data to assess mutation pathogenicity (van der Zwaag et al., 2009). Additionally, a study recently identified the N-terminal regions of DP and Dsg2, in addition to conserved residues of PKP2 and Dsg2, as “hot spots” for radical mutations associated with AC development (Kapplinger et al., 2011). However, little is known regarding the mechanisms by which these mutations promote disease pathogenesis.

Table 2. Mutations associated with AC and AC-overlap syndromes. Detailed descriptions of the listed mutations can be found in the literature cited in this text and elsewhere (Rickelt & Pieperhoff, 2012; van der Zwaag et al., 2009).

Gene target	Protein function
DSG2	Desmoglein 2 (Dsg2): desmosomal cadherin with extracellular domains linking desmosomes of neighboring cells
DSC2	Desmocollin 2 (Dsc2): desmosomal cadherin with extracellular domains linking desmosomes of neighboring cells
PG	Plakoglobin (Pg): desmosomal armadillo protein that links DP to desmosomal cadherins. Has dual localization to classical cadherins of adherens junctions
PKP2	Plakophilin 2 (PKP2): desmosomal armadillo protein that links DP to desmosomal cadherins
DSP	Desmoplakin (DP): desmosomal plakin family protein known for tethering intermediate filaments to desmosomal plaques
TGFB3	Transforming growth factor β3 (TGF-β3): secreted factor involved in extracellular matrix deposition and cell signaling
CTNNA3	alphaT-catenin (αT-catenin): adherens junction component linking actin to sites of cell-cell contact
PLN	Phospholamban (PLN/PLB): membrane protein of the sarcoplasmic reticulum that regulates the SERCA2a calcium ATPase
TMEM43	Transmembrane protein 43/LUMA: protein of inner nuclear membrane that binds lamins and emerin
LMNA	Lamin A/C: proteins of inner nuclear membrane involved in nuclear stability and chromatin structure
TTN	Titin/connectin: sarcomeric protein that serves as a molecular spring between sarcomeric Z and M lines
DES	Desmin: intermediate filament protein expressed in muscle cells

Mutations in other non-desmosomal proteins have also been associated with AC development, though they are less common than desmosomal mutations (Table 2). For example, mutations have been found in transforming growth factor beta 3 (TGF-β3) (Bao et al., 2013; Beffagna et al., 2005). TGF-β3 is a key regulator of fibrotic signaling pathways (Leask & Abraham, 2004; Varga & Jimenez, 1986); thus, these mutations could induce fibrosis by stimulating proliferation of mesenchymal cells. Alternatively, TGFB3 mutations could act by altering expression of Pg (Kapoun et al., 2004) and DP (Yoshida et al., 1992), supporting a theory that non-desmosomal mutations can still act through misregulation of desmosomes.

Following this theory, AC was recently associated with a mutation in the adherens junction component αT-catenin (van Hengel et al., 2013). αT-catenin knockout mouse models exhibit a redistribution of PKP2 (Li et al., 2012b). As αT-catenin can associate with PKPs (Goossens et al., 2007; van Hengel et al., 2013), it is plausible that αT-catenin mutations in AC ultimately act through misregulation of desmosomal proteins.

Screening of AC patients revealed a founder mutation in phospholamban, a regulator of the sarcoplasmic reticulum Ca²⁺ (SERCA2a) pump in cardiac muscle (Groeneweg et al., 2013; van der Zwaag et al., 2012, 2013). Interestingly, SERCA2 regulates desmosomal adhesive strength by promoting membrane localization of DP (Hobbs et al., 2011), once again raising the possibility that desmosomal misregulation is a key step in AC development.

A cohort of patients in Newfoundland, Canada expresses a fully penetrant, lethal version of AC associated with a mutation targeting TMEM43, which encodes the inner nuclear membrane protein LUMA (Merner et al., 2008). TMEM43 mutations were subsequently identified in patients outside of this cohort (Bao et al., 2013; Baskin et al., 2013; Christensen et al., 2011; Haywood et al., 2013; Larsen et al., 2012). TMEM43 has been proposed to be part of the adipogenic peroxisome proliferator-activated receptor γ (PPARγ) pathway (Merner et al., 2008), suggesting that mutations may hyperactivate adipogenesis. LUMA binds lamins and emerins to contribute to nuclear envelope integrity (Bengtsson & Otto, 2008), and a mutation in lamin A/C has recently been associated with AC (Quarta et al., 2012). A potential relationship between these mutations and desmosomal misregulation has not been established.

Mutations in the sarcomeric protein titin (TTN) (Taylor et al., 2011) and the IF desmin (DES) (Lorenzon et al., 2013; Hedberg et al., 2012; Klauke et al., 2010; van Tintelen et al., 2009) have been associated with AC but are sometimes considered to produce phenocopies or AC-overlap syndromes (Basso et al., 2011; McLendon & Robbins, 2011; Otten et al., 2010; van Spaendonck-Zwarts et al., 2011). It is likely that future studies will reveal additional non-desmosomal mutations associated with AC. Further screening of large patient cohorts is needed to assess the prevalence of these mutations in AC patients, to determine the specificity of mutations to AC versus other cardiac diseases, and to elucidate if and how their pathogenicities relate to desmosomal misregulation.

Theories of AC pathogenesis

Though histological findings of replacement-type fibrosis, fat deposition, and myocyte degeneration are hallmarks of AC, the mechanisms by which these changes arise are poorly understood. Possible causes are grouped into three theories of pathogenesis lacking mutual exclusivity: degeneration, inflammation, and transdifferentiation (Figure 2) (Basso et al., 2011; Herren et al., 2009).

Figure 2. Theories of AC pathogenesis. AC may arise from cardiac myocyte degeneration, inflammation of myocardium from infection by cardiotropic pathogens, and/or transdifferentiation of cardiac myocytes or progenitor cells into fat- and fibrotic scar-producing cells (Basso et al., 2011; Herren et al., 2009).

In the degenerative theory, it is proposed that cardiac myocyte death enables replacement with fibrous and fatty tissue. A number of studies have pointed to apoptotic death (Campian et al., 2011; Garcia-Gras et al., 2006; Nagata et al., 2000; Nishikawa et al., 1999; Valente et al., 1998; Yamaji et al., 2005). Supporting this idea, fragmented DNA and expression of protease CPP-32 were present in 75% of myocardial samples in one study of AC patients (Mallat et al., 1996), and expression of an AC PKP2 mutation in cardiac cells increased levels of apoptosis (Joshi-Mukherjee et al., 2008). However, there are also examples of necrosis in human samples and animal models of AC (Gattenlohner et al., 2009; Li et al., 2011b; Pilichou et al., 2009). It is plausible that more than one cell death pathway is involved in AC development (Li et al., 2011a).

Inflammatory infiltrates are commonly found in AC, suggesting that inflammation could contribute to AC development (Asimaki et al., 2011; Basso et al., 1996; Calabrese et al., 2006; Fornes et al., 1998). Non-familial AC may be linked to inflammation from infections by viruses including enterovirus, adenovirus, cytomegalovirus, parvovirus B19, and coxsackievirus (Bowles et al., 2002; Grumbach et al., 1998), or by the bacterium Bartonella henselae (H Fischer et al., 2008). It has been proposed that this inflammation can be noninvasively detected by assaying plasma levels of IL-1β, IL-6, and TNF-α (Campian et al., 2010). However, AC patients sometimes have very little evidence of inflammation (Tabib et al., 2003). Inflammatory infiltrate could be secondary to cell death, or could be limited to severe biventricular forms of AC (Campuzano et al., 1986). Thus, the source of inflammation and the value of its assessment remain unclear.

Finally, observations of myocardial cells expressing altered cytological markers have led to the development of a transdifferentiation theory. Since adult cardiac myocytes have limited capabilities for dedifferentiation, it is possible that pro-adipogenic cells found in AC samples are derived from progenitor cells in the subepicardial or second heart fields (Gittenberger-de Groot et al., 2010; Lombardi et al., 2009; Matthes et al., 2011). Multiple AC studies have noted an accumulation of fat droplets or activation of adipogenic pathways in cardiac myocytes (Chen et al., 2014; D’Amati et al., 2000; Fujita et al., 2008; Garcia-Gras et al., 2006; Kim et al., 2013), though it is unclear if these changes are indicative of transdifferentiation.

There is much to be understood regarding the specific mechanisms by which desmosomal mutations may trigger these pathways of disease pathogenesis. Recent work has started to evaluate how loss of desmosomal adhesion and/or signaling promotes cell death, increases susceptibility for viral infection, and/or promotes fibro-fatty change. These pathways will be eplained later in this review.

Desmosomal misregulation in other cardiac diseases

Clinical reports have demonstrated that desmosomal proteins are mutated or otherwise misregulated in a broader spectrum of cardiac diseases. A growing number of studies have established links between desmosomal proteins and components of cardiac electrical machinery including the gap junction subunit connexin43 (Cx43) (Agullo-Pascual et al., 2013; Gehmlich et al., 2011a; Oxford et al., 2007; Sato et al., 2011) and the sodium channel isoform Na_v1.5 (Cerrone et al., 2012; Rizzo et al., 2012; Sato et al., 2009). Therefore, it is possible that desmosomal mutations may contribute more generally to cardiac diseases involving arrhythmia. For example, mutations in PKP2 proposed to cause Na_v1.5 mislocalization were recently described in patients with Brugada syndrome (Cerrone et al., 2013). Similarly, mutations in PKP2 have been associated with sudden unexplained death with negative autopsy (SUDNA) in which cardiac tissue is morphologically normal and therefore not definitively marked as AC (Zhang et al., 2012).

PKP2 was also found to be a marker of cardiac myxomata derived from mesenchymal cells (Rickelt et al., 2010). PKP2 rapidly accumulates in abnormal adherens junction complexes in myxoma samples, without accumulation of other desmosomal proteins. The mechanisms by which PKP2 localizes to adherens junctions in these samples, and the significance of this finding as being a cause versus marker of disease, are yet to be determined.

Desmosomal mutations have also been associated with DCM (Elliott et al., 2010; Pugh et al., 2014). One study noted that 13% of DCM patients harbored desmosomal mutations predicted to be pathogenic, and an additional 6% had desmosomal mutations of unknown significance (Garcia-Pavia et al., 2011).

The IF desmin is a vital cytoskeletal component of striated muscle, and desmin mutations have been associated with DCM (Li et al., 1999) and with a more generalized “desmin myopathy” characterized by skeletal muscle weakness, arrhythmias, and restrictive heart failure (Goldfarb et al., 2004). Desmin myopathy can be caused by mutations in the desmin rod or tail domains, and is typically associated with intracytoplasmic desmin-reactive deposits (Goldfarb et al., 1998; Dalakas et al., 2003).

Finally, while the loss of Pg from intercalated discs has been proposed to be prognostic for AC (Asimaki et al., 2009), a reduction in junctional Pg was also noted in granulomatous myocarditis and sarcoidosis (Asimaki et al., 2011). Pathogenesis of these diseases may be linked to increased cytokine signaling (Asimaki et al., 2011) and/or activation of adipogenic and fibrotic pathways (Garcia-Gras et al., 2006), as can occur upon mislocalization of Pg.

The spectrum of conditions associated with desmosomal misregulation highlights the need to better understand how desmosomal mutations are a basis for disease development.

DESMOSOMES IN DISEASE: MECHANISTIC INSIGHTS

Biochemical analyses, molecular biology, and physiological studies have revealed non-traditional roles for desmosomal proteins that may provide insight to understanding cardiac disease pathogenesis. We review those studies here, highlighting mouse models and studies of individual disease mutations (Table 3). We refer readers to other reviews for additional information on the basic biology of desmosomes (Delva et al., 2009; Harmon & Green, 2013) and their roles in human disease (Delmar & McKenna, 2010; McGrath, 1999).

Table 3. Potential mechanisms of AC mutation pathogenicity. Select findings from mouse models and/or in vitro studies are highlighted here, with further information available in the text.

Mechanism	Targeted Protein	Summary of Findings	Citation
Cell death	Dsg2	Transgenic expression of N271S (equivalent of human Dsg2 N266S) increases cell necrosis	Pilichou et al. (2009)
	Dsg2	Mice lacking Dsg2 ectodomain exhibit increased cardiac myocyte death and fibrosis	Krusche et al. (2011)
	DP	Transgenic expression of DP R2834H increases cell apoptosis	Yang et al. (2006)
Transcriptional activity	Dsg2	Mice lacking Dsg2 ectodomain have increased mRNA expression of c-Myc, ANF, BNF, CTGF, GDF15	Krusche et al. (2011)
	Pg	Cardiac-restricted deletion of Pg leads to increase of β-catenin signaling in nucleus	Li et al. (2011b)
	Pg	Cardiac-restricted deletion of Pg leads to increase in TGF-β-mediated signaling	Li et al. (2011a)
	DP/Pg	Cardiac-restricted deletion of DP leads to nuclear translocation of Pg with expression of pro-adipogenic and pro-fibrotic genes	Garcia-Gras et al. (2006)
	Pg	Transgenic expression of truncated Pg increases expression of adipogenic factors	Lombardi et al. (2011)
	PKP2	iPSC-derived cardiac myocytes expressing PKP2 c.2484C> T demonstrate nuclear translocation of Pg with decreased β-catenin transcriptional activity	Kim et al. (2013)
Gap junction or ion channel abnormalities	Dsg2	Transgenic expression of N271S (equivalent of human Dsg2 N266S) compromises ventricular activation time	Rizzo et al. (2012)
	Dsc2	Q851fsX855 compromises association of Dsc2a with Cx43 and Pg	Gehmlich et al. (2011a)
	PKP2	Mouse models of PKP2 haploinsufficiency exhibit sodium current deficiencies	Cerrone et al. (2012)
Protein stability and/or modification	Dsg2	C813R targeting ICS alters Dsg2 protein migration on gel electrophoresis	Gehmlich et al. (2010)
	Dsg2	V977fsX1006 targeting DUR increases rate of Dsg2 endocytosis	Chen et al. (2012)
	Dsg2	R46Q targeting proprotein convertase cleavage site impairs Dsg2 prodomain cleavage	Gaertner et al. (2012)
	Dsc2	R203C and T275M impair Dsc2 prodomain cleavage	Gehmlich et al. (2011b)
	Dsc2	E102K and I345T increase Dsc2 cytoplasmic localization	Beffagna et al. (2007)
	Pg	S39_K40insS increases ubiquitylation and cytoplasmic localization of Pg	Asimaki et al. (2007)
	PKP2	c.2386T> C increases degradation of PKP2 by calpain proteases	Kirchner et al. (2012)
	DP	R1269X and K324_E325del lead to degradation of DP, producing functional haploinsufficiency	Rasmussen et al. (2013a)

Desmosomal cadherins

Similar to classical cadherins of adherens junctions, the extracellular domains of desmosomal cadherins form adhesive bonds between desmosomal plaques of neighboring cells. As such, mutation or autoimmune-mediated targeting of desmosomal cadherins can lead to blistering disorders including pemphigus vulgaris, pemphigus foliaceus, and keratodermas, reviewed elsewhere (Amagai, 2010; Getsios et al., 2010; Waschke, 2008). Misregulation of desmosomal cadherins has also been associated with defects in hair growth/cycling (McGrath & Wessagowit, 2005). Here, we will focus on misregulation of desmosomal cadherins expressed in cardiac tissue: Dsg2 and Dsc2.

Dsg

Dsg2-knockout mouse models demonstrate a vital function for Dsg2 in development. Dsg2 is needed for embryonic stem cell proliferation, and Dsg2-/- mice die at or shortly after implantation (Eshkind et al., 2002). More restricted targeting of Dsg2 through transgenic expression of Dsg2 N271S, the mouse equivalent of the human AC mutation Dsg2 N266S, reproduces features of AC with myocyte necrosis thought to initiate myocardial damage (Pilichou et al., 2009). Mice expressing a Dsg2 mutant lacking the Dsg2 ectodomain demonstrate cell death and fibrosis associated with increased mRNA expression of cardiac stress and remodeling markers, including c-Myc, ANF, BNF, CTGF, and GDF15 (Krusche et al., 2011). Importantly, necrosis in Dsg2 mutant models is preceded by widening of intercellular spaces with abnormalities in ventricular activation time, attributed to the novel finding of an interaction between Dsg2 and the sodium channel Na_v1.5 (Rizzo et al., 2012). Thus, necrosis may be a secondary, late stage of disease following more specific signaling changes.

Alterations in molecular pathways that may contribute to disease pathogenesis induced by expression of Dsg2 mutations have been revealed by recent biochemical analyses. AC mutation C813R targeting the intracellular cadherin segment (ICS) domain of Dsg2 causes a shift in protein migration detected by gel electrophoresis (Gehmlich et al., 2010). Migration patterns were restored by the mutant C813A, suggesting that mutations may introduce polar residues that alter posttranslational modifications of Dsg2. The V977fsX1006 frameshift mutation in the Dsg2 unique region (DUR) of Dsg2 compromised tail–tail interactions of homodimerized Dsg2, leading to an increased rate of Dsg2 endocytosis and thus loss of cell surface Dsg2 (Chen et al., 2012). Further, the R46Q mutation in the proprotein convertase cleavage site of Dsg2 impairs proper prodomain cleavage, prerequisite to normal Dsg2 function in desmosomes (Gaertner et al., 2012). Because it has been observed that mutated Dsg2 proteins can be incorporated into desmosomes (Rasmussen et al., 2013b), it is likely that phenotypes induced by these mutations promote disease development even when co-expressed with wild-type Dsg2.

Interestingly, Dsg2 has been identified as a high-affinity receptor used by multiple adenovirus serotypes (Wang et al., 2011a,b, 2013). More information is needed to establish if and how mutated Dsg2 may enhance AC development upon viral infection.

Dsc

Similar to some Dsg2 mutations, missense mutations R203C and T275M in Dsc2 hinder proteolytic cleavage of the Dsc2 prodomain, causing a reduction in functional Dsc2 (Gehmlich et al., 2011b). Missense mutations E102K targeting the Dsc2 prodomain and I345T targeting the Dsc2 ectodomain result in cytoplasmic localization of Dsc2, likely by respectively impairing proteolytic cleavage and adhesive functions of Dsc2 (Beffagna et al., 2007). Interestingly, a recent study found that the Dsc2a splice variant can directly interact with the gap junction component Cx43, and that expression of the mutation Q851fsX855 compromises binding of Dsc2a with both Cx43 and Pg (Gehmlich et al., 2011a). Thus, mutations in desmosomal cadherins can impair mechanical and electrical coordination of cardiac cells through distinct mechanisms.

Armadillo proteins

The structural similarities between desmosomal armadillo proteins (Pg and PKPs) and their respective counterparts in adherens junctions (β-catenin and p120 catenin) have prompted analysis of the unique and overlapping functions of these junctional proteins (Hatzfeld, 2007; Bass-Zubek et al., 2009; Zhurinsky et al., 2000b). Armadillo proteins not only play structural roles in junctions but also activate and repress transcription of nuclear genes (Miller et al., 2013; Schmidt & Koch, 2007). The extent to which their junctional and nonjunctional functions contribute to human disease pathogenesis is yet to be fully determined.

Pg

Pg (also known as γ-catenin) is homologous to the adherens junction component β-catenin (Butz et al., 1992; McCrea et al., 1991), an effector of the canonical Wnt signaling pathway that promotes T-cell/lymphoid-enhancing binding (TCF/LEF) transcriptional activity. Pg-null mouse models demonstrate embryonic lethality between E12 and 16 with a “broken heart” phenotype: cardiac ventricles burst and blood fills the pericardium (Ruiz et al., 1996). The architecture of intercellular junctions is severely altered in these mice, with a diffuse localization of Dsg2, abnormally elongated junctions, and an enhanced co-localization of DP with β-catenin at early time points of development. β-catenin can partially compensate for the absence of Pg in the skin, where it associates with desmosomal cadherins and reduces the severity of skin phenotypes in Pg-null mice (Bierkamp et al., 1999; Li et al., 2012a).

Importantly, while Pg and β-catenin may replace each other in junctions, their structural similarities may actually lead to functional competition in the nucleus (Ben-Ze'ev & Geiger, 1998; Hu et al., 2003; Simcha et al., 1998; Zhurinsky et al., 2000a) since the arm domain mediates binding of both proteins to TCF/LEF nuclear partners (Behrens et al., 1996; Huber et al., 1996; Simcha et al., 1998; Miravet et al., 2002). For example, cardiac-restricted deletion of Pg leads to a concomitant increase in β-catenin at junctions and in the nucleus, where it associates with TCF-4 to activate transcription of factors including c-Myc, c-Fos, and cyclin D1 (Li et al., 2011b). In contrast, Pg has been shown to form a complex with LEF-1 to repress transcription of c-Myc in the epidermis (Williamson et al., 2006).

Interestingly, cardiac deletion of DP results in relocalization of Pg to the nucleus, where it suppresses Wnt/β-catenin signaling and activates expression of adipogenic and fibrotic genes including C/EBP-α, PPARγ, adiponectin, lipoprotein lipase, and procollagens (Garcia-Gras et al., 2006). However, a separate study of Pg ablation in cardiac myocytes revealed an AC phenocopy with increased cell death but without a change in Wnt/β-catenin-mediated signaling (Li et al., 2011a). Instead, these mice were found to have elevated levels of TGF-β-mediated signaling. Thus, mouse models have not fully clarified mechanisms by which Pg and β-catenin are regulated, or by which they may affect each other's activities (Zhurinsky et al., 2000b).

Since a Pg mutation associated with Naxos disease (Pg2157del2) produces a truncated form of Pg that is still detectable in patient biopsies (McKoy et al., 2000), further information may be gained by studying mutated Pg rather than Pg-null models. C-terminal truncation mutants of Pg can be incorporated into desmosomes but alter junction morphology and produce extremely long desmosomes (Palka & Green, 1997). A transgenic mouse model overexpressing truncated Pg confirmed a loss of binding to DP and Dsg2 (Lombardi et al., 2011). As truncated Pg localizes primarily to the nucleus, cardiac progenitor cells isolated from these mice demonstrate increased levels of adipogenic factors including KLF15, C/EBP-α, and Wnt5β, and reduced levels of the anti-adipogenic factor CTGF. Thus, these more specific mutant models are useful in determining mechanisms of disease pathogenesis.

Expression of an N-terminal S39_K40insS Pg mutation associated with AC has demonstrated increased ubiquitylation and cytoplasmic localization of Pg (Asimaki et al., 2007). Further, this Pg mutant revealed novel interactions with histidine-rich calcium-binding protein (HRC-BP) and TGF-β-induced apoptosis protein 2 (TAIP-2), producing higher rates of proliferation and lower rates of apoptosis. While expression of either Naxos-associated Pg2157del2 or S39_K470insS increases cell motility in epithelial cells, 2157del2 does so by compromising cell–cell adhesion while S39_K470insS decreases cell stiffness (Huang et al., 2008). In vitro studies have demonstrated that Pg can regulate fibronectin production and Src signaling via the small GTPases Rac1 and RhoA in epithelial cells (Yin et al., 2005; Todorovic et al., 2010). Together, these observations suggest that Pg mutations could act by impairing desmosomal and non-desmosomal functions.

PKP

PKPs 1, 2, and 3 have variable expression patterns in different tissues: PKP1 is most highly expressed in suprabasal layers of stratified epithelia, PKP2 is expressed in all simple and complex epithelia in addition to cardiac myocytes and lymph node follicle cells, and PKP3 is expressed in most simple epithelia in addition to all layers of stratified epithelia (Bass-Zubek et al., 2009; Hatzfeld, 2007). PKPs localize to desmosomes in addition to cell nuclei, with this differential localization potentially dictated by alternative splice products (Schmidt et al., 1997) or by the interaction of PKPs with scaffolding proteins such as 14-3-3 (Muller et al., 2003).

While a potential role for PKPs in controlling gene transcription is uncertain, it is clear that they have functions in addition to their roles in adhesion. PKP2 can interact with the RNA polymerase III subunit of the transcription factor TFIIIB (Mertens et al., 2001) and can augment β-catenin/TCF-mediated transcription (Chen et al., 2002). PKP3 forms cytoplasmic stress-induced granules with RNA-binding proteins PABPC1, FXR1, and G3BP (Hofmann et al., 2006), and has recently been shown to associate with the ETV1/ER81 transcription factor (Munoz et al., 2014). While all PKPs may be important in the pathogenesis of human diseases including ectodermal dysplasia-skin fragility syndrome (McGrath & Wessagowit, 2005), here we will focus on studies of PKP2, the sole PKP expressed in heart.

PKP2-null mice lack obvious epithelial defects, likely due to compensation from other PKP isoforms; yet, these mice experience embryonic lethality at E10.5–E11 due to severe alterations in cardiac morphology (Grossmann et al., 2004). Mice with PKP2 haploinsufficiency are viable but exhibit sodium current deficiencies (Cerrone et al., 2012), supporting data on the association of PKP2 with the Na_v1.5 channel in cells (Sato et al., 2009, 2011). In cardiac cell cultures, PKP2 knockdown compromises gap junction signaling and Cx43 localization to sites of cell–cell contact (Oxford et al., 2007; Fidler et al., 2009). In fact, the cytoskeletal adaptor protein ankyrin-G may coordinate interactions between PKP2, Cx43, and Na_v1.5 (Sato et al., 2011).

In epithelial cells, PKP2 was shown to regulate the localization of RhoA to sites of cell–cell contact, with PKP2 deficiency resulting in an impairment of cortical actin remodeling, an increase in contractile signaling, and alterations in focal contact structure and integrin expression (Godsel et al., 2010; Koetsier et al., 2014). Similar alterations in actin have been observed in PKP2-deficient HL-1 and neonatal rat cardiac myocytes (Dubash et al., unpublished observations). This work raises the possibility that PKP2 could control matrix remodeling in AC. Further, PKP2 forms a complex with DP and PKCα to regulate the recruitment of DP to cell–cell borders (Bass-Zubek et al., 2008). PKCα activity is thought to be responsible for a switch between dynamic and “hyperadhesive” states of desmosomes (Wallis et al., 2000; van Hengel et al., 1997), suggesting that misregulation of PKCα localization and/or activity could compromise structural integrity of cell networks including that of cardiac muscle. Thus, misregulation of PKP2 could act in part by affecting its downstream regulation of the desmosomal plaque protein, DP. Further, PKP2-knockdown cardiac cells exhibit reduced levels of active PKCα with subsequent hyperactivation of the Hippo pathway, leading to enhanced adipogenesis that likely contributes to AC (Chen et al., 2014).

Some PKP2 mutations are thought to result in loss of protein: for example, the PKP2 mutant c.2386T> C exhibits enhanced degradation by calpain proteases (Kirchner et al., 2012). However, other studies have demonstrated that PKP2 mutations can result in stable, dysfunctional PKP2 (van der Smagt et al., 2012), highlighting the importance of studying both knockdown and mutant PKP2 models. With recent advances in stem cell biology, researchers have begun to explore the pathogenicities of specific mutations using induced pluripotent stem cell (iPSC)-derived cardiac myocytes grown from fibroblasts of patients with PKP2 mutations (Ma et al., 2013). iPSC-derived cardiac myocytes expressing the PKP2 mutation c.1841T> C demonstrate an accumulation of lipid droplets. Similarly, cells derived from a patient with the PKP2 c.2484C> T mutation exhibit nuclear translocation of Pg with decreased β-catenin activity (Kim et al., 2013). When altering metabolic conditions in culture by modulating PPAR-γ activity, these cells reproduce aspects of AC in vitro with increased lipogenesis and apoptosis. Application of an inhibitor of glycogen synthase kinase 3β (GSK3β) prevents adipogenesis in iPSC-derived cardiac myocytes expressing the c.972InsT/N PKP2 mutation (Caspi et al., 2013), suggesting a potential therapeutic strategy for patients with AC.

DP

DP is an obligate component of desmosomes vital for integrity of multiple tissue types including the skin and heart. DP ablation leads to embryonic lethality by E6.5 with defects in extraembryonic tissue integrity (Gallicano et al., 1998). Extra-embryonic rescue of DP via a tetraploid aggregation method delays lethality to E12.5, with cardiac hemorrhages and defective capillary formation (Gallicano et al., 2001). Conditional depletion of DP from the epidermis enables embryonic survival but leads to death soon after birth due to severe sloughing of skin (Vasioukhin et al., 2001). Interestingly, while adherens junctions are typically thought to regulate desmosome assembly, conditional DP-knockout mice demonstrate abnormalities in adherens junctions. Thus, the assembly, maturation, and maintenance of desmosomes and adherens junctions may be more interdependent than previously recognized. Conditional ablation of DP from cardiac tissue recapitulates AC with defects in connexin expression (Lyon et al., 2013) and electrophysiological abnormalities before the onset of overt structural changes (Gomes et al., 2012). Changes in gap junction and sodium channel signaling upon DP knockdown have been reproduced in cardiac cell systems (Zhang et al., 2013).

In humans, expression of a truncating mutation in the DP C-terminus completely ablates the function of DP in tethering IFs to desmosomes, resulting in the fatal condition of lethal acantholytic epidermolysis bullosa (Bolling et al., 2010; Hobbs et al., 2010; Jonkman et al., 2005). DP mutations and haploinsufficiency have also been associated with less severe epidermal conditions including skin fragility/woolly hair syndrome, hypo/oligodontia, and striate palmoplantar keratoderma, some of which can be associated with cardiac findings (Armstrong et al., 1999; Chalabreysse et al., 2011; Mahoney et al., 2010; Smith et al., 2012; Whittock et al., 1999, 2002).

Biological analysis of individual DP mutations is needed to fully understand the pathogenesis of AC and these epidermal diseases. AC mutations V30M and Q90R targeting the N-terminus of DP display an inability to bind Pg in cells; however, transgenic expression of these mutations leads to embryonic lethality, preventing further understanding of how these mutations might contribute to gradual AC development (Yang et al., 2006). In this same study, transgenic expression of R2834H targeting the C-terminus of DP recapitulates AC findings by 6 months of age with lipogenesis, fibrosis, and an absence of desmin at intercalated discs. These mice also demonstrate an increase in cell apoptosis; however, a separate study has shown that RNAi-mediated silencing of DP in keratinocytes actually increases cell proliferation (Wan et al., 2007).

It is possible that the domain of DP targeted by AC mutations may dictate pathogenic effects and potential lethality. Mutations targeting the N-terminus of DP could completely prevent localization of DP to desmosomes, whereas mutations targeting the DP C-terminus could allow for partial function of DP in desmosomal plaques but impair tethering of IFs to sites of cell–cell contact. AC mutations may also have different effects on DP expression levels: one study demonstrated that mutant proteins could either be expressed in cells or lead to degradation/instability producing functional haploinsufficiency (R1269X, K324_E325del) (Rasmussen et al., 2013a).

There is much to be understood regarding novel functions of DP that stand apart from its tethering of IFs. As mentioned, DP knockout mice activate adipogenesis and fibrosis via nuclear translocation of Pg, antagonistic of canonical Wnt/β-catenin signaling (Garcia-Gras et al., 2006). In support of this finding, DP was shown to act as a tumor suppressor by inhibiting Wnt/β-catenin signaling in a model of non-small cell lung cancer (Yang et al., 2012). Recent studies also implicate DP in binding the microtubule plus-end binding protein EB1 to promote Cx43 membrane localization (Patel and Green, in revision). These findings highlight potential novel mechanisms by which DP mutations can contribute to AC development.

IFs

Misregulation of desmosome-associated IFs is implicated in a number of diseases (Omary, 2009). It is unclear if mutations in desmin, the IF expressed in cardiac muscle, primarily cause AC (Lorenzon et al., 2013; Klauke et al., 2010; Hedberg et al., 2012) or AC phenocopies (van Tintelen et al., 2009; van Spaendonck-Zwarts et al., 2011; McLendon & Robbins, 2011; Basso et al., 2011). Similar to broad findings of desmin myopathy, desmin-null mice develop defects in multiple muscle types (Milner et al., 1996; Li et al., 1996). The determinants of desmin misregulation affecting multiple tissue types as opposed to cardiac muscle alone are not well understood; nonetheless, some valuable information has emerged regarding the pathogenicities of desmin mutations (Bar et al., 2004; Clemen et al., 2013).

Several desmin mutations located in the central α-helical rod domain of desmin affect filament formation (Brodehl et al., 2013; Bar et al., 2005; Dagvadorj et al., 2003; Sjoberg et al., 1999; Munoz-Marmol et al., 1998; Goldfarb et al., 1998; Goudeau et al., 2001) or can compromise association of desmin with nebulin, through which it attaches to cardiac Z-discs (Conover & Gregorio, 2011). Interestingly, disease mutations have been reported to target putative PKC sites in the desmin head domain (Pica et al., 2008), a region whose phosphorylation is proposed to regulate cardiac myocyte differentiation (Hollrigl et al., 2007).

Truncation of the tail domain of desmin can abolish interactions with DP (Lapouge et al., 2006). Mice transgenically expressing the I451M mutation targeting the desmin tail demonstrate a loss of desmin localization to Z-discs (Mavroidis et al., 2008). Though desmin I451M can still localize to intercalated discs, junctions exhibit altered morphology. Interestingly, expression of the desmin mutation R454W compromises junctional localization of PKP2 and DP (Otten et al., 2010). Further, a deletion mutation in desmin has been shown to impair membrane localization of Cx43 (Gard et al., 2005). Thus, though IFs are an “end-point” of desmosomes, it is important to consider them as potential regulators of junction assembly and stability. More information on these pathways is likely to be obtained from the use of iPSC-derived cardiac myocytes from patients with desmin mutations (Tse et al., 2013).

CONCLUDING REMARKS

Historically, desmosomes have been considered as rigid plaques that maintain adhesive integrity of cells in contact. However, as outlined here, the dynamic natures of desmosomal components and their novel non-desmosomal roles are becoming increasingly recognized as key determinants of tissue homeostasis and disease pathogenesis. Researchers and clinicians are able to draw from studies conducted in vitro and in vivo to understand how mutations in these desmosomal proteins can misregulate various cellular processes.

Since the discovery of AC as a “disease of the desmosome,” many patients with cardiac findings have been screened for mutations in desmosomal proteins. However, there are unfortunately a number of confounding factors complicating this approach. Most obviously, the variable penetrance of mutations is poorly understood and may be related to exercise-induced stress, gene–dose effects, and compound and/or digenic heterozygosity (Basso et al., 2011; Corrado et al., 1997, 2006b; Kirchhof et al., 2006; Rigato et al., 2013; Xu et al., 2010). Systems analysis may be needed to better determine if AC-associated mutations are non-pathogenic, disease modifying, or highly pathogenic (van der Zwaag et al., 2009; Silvano et al., 2013; Kapplinger et al., 2011; Christensen et al., 2010). Traditional screening methods may lack sensitivity for copy number variations or cryptic splice site mutations in desmosomal genes (Awad et al., 2006; Li Mura et al., 2013), and often neglect consideration of mutations in lesser-studied genes.

Despite these issues, studies analyzing the pathogenicities of individual mutations are essential for gaining a better understanding of disease development. A number of studies have used structural modeling to predict potential mechanisms by which AC mutations could alter protein function (Al-Jassar et al., 2013; Choi & Weis, 2011), though very few studies have expressed these mutations to analyze their effects in vitro or in vivo. Use of iPSC-derived cardiac myocytes obtained from patients with AC mutations, with consideration of epigenetic factors that may contribute to disease development, is a specific method to test misregulation of adhesive and signaling pathways. In the long-term, an integration of molecular modeling, in vitro analysis, and iPSC-based biology may provide key information regarding mechanisms by which desmosomal mutations serve as a basis for disease development.

Declaration of interest: The authors report no declarations of interest. The authors alone are responsible for the content and writing of the paper.