Desmoplakin Gene Variants and Risk for Arrhythmogenic Cardiomyopathy

SP (Desmoplakin), a cytolinker of the plakin family, is an essential intracellular component of the cell-cell adhesion complexes desmosomes.Its main function is the anchoring of intermediate filaments (IFs), cytokeratins (Ks), and desmin in epithelial cells and cardiomyocytes respectively, to desmosomes.Tissue-specific gene deletion in mouse has shown that DSP is indispensable for maintenance of the cytoarchitecture and cell resilience in both skin and heart. 1 Numerous human DSP variants have been linked to inherited diseases that variably result in skin fragility, palmoplantar keratoderma, woolly hair, and arrhythmogenic or dilated cardiomyopathy.1,2 Homozygous or compound heterozygous mutations that truncate the DSP tail, which bears several domains that synergistically bind to IFs (Figure [A]), are generally pathogenic.For example, severe C-terminal truncations affecting the PRD (plakin repeat domain) 2/B or preceding regions cause lethal acantholytic epidermolysis bullosa, while shorter truncations often result in Carvajal syndrome, characterized by palmoplantar keratoderma, hair defects, and dilated left ventricular cardiomyopathy.1,2 Heterozygous DSP mutations exhibit a complex genotype-phenotype relationship that is sometimes questionable.3 The factors that ultimately determine the apparent skin-or heart-specific impact of certain DSP variants are also unclear.1,2 Furthermore, it is exceptional that a heterozygous mutation specifically weakening the interaction of DSP with IFs is pathogenic in the skin or heart.2,3 Recently, we found that the amino acid substitution p.(Gly2375Arg) in the DSP PRD 2/B drastically reduces the binding of the carboxyl (C) terminus of DSP to Ks, desmin, and vimentin.4 This substitution was associated with arrhythmogenic right ventricular cardiomyopathy, skin fragility, and woolly hair in a homozygous patient.This result suggests that analysis of the IF-binding properties of variants affecting the DSP tail could explain or even predict clinical phenotypes, especially cardiomyopathy that has usually a later onset than skin diseases.To gain better insights into the molecular basis of DSP-related human diseases, we have here assessed the impact of 7 published amino acid substitutions modifying the DSP C-terminus (Figure [A]) on its IF-binding activity by using a novel fluorescence binding assay.4 The mutations were selected based on the variable associated skin and cardiac phenotype(s) (Figure [B]).The difficulty to estimate the pathogenicity of these variants is illustrated by the frequent ambiguous conclusions of the ClinVar and arrhythmogenic right ventricular cardiomyopathy databases (Figure [B]).Prediction of the effect of the tested amino acid substitutions on the protein structure/function with different programs (PolyPhen-2, SIFT, and PROVEAN) was also not univocal (data not shown).Our biochemical assays revealed that like p.(Gly2375Arg) the amino acid substitutions p.(Gln2295His) and p.(Arg2366Cys) in the DSP PRD 2/B almost abolished the IF-binding activity of the DSP C-terminus to both Ks and type III IF proteins (Figure [C and D]).Furthermore, the substitution p.(Ala2655Asp), located in the