In Vitro Functional Analyses of Arrhythmogenic Right Ventricular Cardiomyopathy-Associated Desmoglein-2-Missense Variations

Abstract

BackgroundAlthough numerous sequence variants in desmoglein-2 (DSG2) have been associated with arrhythmogenic right ventricular cardiomyopathy (ARVC), the functional impact of new sequence variations is difficult to estimate.Methodology/Principal FindingsTo test the functional consequences of DSG2-variants, we established an expression system for the extracellular domain and the full-length DSG2 using the human cell line HT1080. We established new tools to investigate ARVC-associated DSG2 variations and compared wild-type proteins and proteins with one of the five selected variations (DSG2-p.R46Q, -p.D154E, -p.D187G, -p.K294E, -p.V392I) with respect to prodomain cleavage, adhesion properties and cellular localisation.Conclusions/SignificanceThe ARVC-associated DSG2-p.R46Q variation was predicted to be probably damaging by bioinformatics tools and to concern a conserved proprotein convertase cleavage site. In this study an impaired prodomain cleavage and an influence on the DSG2-properties could be demonstrated for the R46Q-variant leading to the classification of the variant as a potential gain-of-function mutant. In contrast, the variants DSG2-p.K294E and -p.V392I, which have an arguable impact on ARVC pathogenesis and are predicted to be benign, did not show functional differences to the wild-type protein in our study. Notably, the variants DSG2-p.D154E and -p.D187G, which were predicted to be damaging by bioinformatics tools, had no detectable effects on the DSG2 protein properties in our study.

Whole Genome Sequence Identified a Rare Homozygous Pathogenic Mutation of the DSG2 Gene in a Familial Arrhythmogenic Cardiomyopathy Involving Both Ventricles

“…Firstly, the mutation of DSG2 p.F531C, which changes an amino acid in the extracellular domain, may have an effect on mechanical, connective, and adhesive characteristics, but it may not influence the expression and localization of the DSG2 protein in cells, according to previous in vitro researches [22,23,24]. Thus, the underlying mechanism of how DSG2 p.F531C contributes to the development of ARVC/D needs to be explored in a transgenic animal model, similar, for example, to the transgenic mouse that carries the DSG2 p.N271S mutation [17].…”

Desmoglein 2–Dependent Arrhythmogenic Cardiomyopathy Is Caused by a Loss of Adhesive Function

“…10,12,29 In vitro experimentation further supports an adhesive function of Dsg2. Expression of different human AC-related Dsg2 mutants and inhibition of Dsg2 adhesion were reported to reduce adhesion upon expression in HL-1 cardiomyocytes 22 (for contrasting results on another mutant see Gaertner et al 30 ). The situation in other AC types is less clear.…”

“…In all instances, interference seems to occur at the ID because mutant Dsg2 is efficiently localized to the ID in the different murine models and human Dsg2-related AC. 8,11,20,30,41 Multiple studies have assigned a crucial role of plakoglobin signaling to the initiation of AC in mouse and man. 17,[42][43][44] Comparing disease manifestation with the available binding sites of desmosomal cadherins for Pg or other desmosomal cadherin-dependent signaling molecules in the various murine models, however, shows that there is no clear correlation (Figure 8).…”

Does p.Q247X in TRIM63 Cause Human Hypertrophic Cardiomyopathy?

“…Of interest, Gaertner et al 12 did not show any functional differences comparing the DSG2 V392I protein with the wild-type variant. In silico prediction classifies the V392I in the DSG2 as benign and in the arrhythmogenic right ventricular cardiomyopathy database (http://www.arvcdatabase.info); this variant is currently classified as a variant of unknown significance.…”