Abstract 375: Defining a Unifying Mechanism for Select Cardiomyopathy-Linked Variants of Desmoplakin

Arrhythmogenic cardiomyopathy (AC) is a disease that affects 1 in 2000 Americans every year and segregates with sudden cardiac death (SCD). AC is linked to genetic variants in desmosomal genes, however the molecular mechanisms underlying AC remain unclear. To probe the mechanisms of an AC-linked desmosomal mutation, we identified a candidate family with a history of SCD and an unknown variant (R451G) in the desmoplakin (DSP) gene. Statistically significant linkage between the R451G-variant and symptomatic family members (LOD score = 3.4) was established and heart autopsy tissue from a genotype-phenotype positive family member revealed pronounced fibrofatty scarring and a significant loss of DSP protein at the intercalated disc (ID). Notably, the R451G-variant sits in a pocket containing several clinical DSP variants and we show that endogenous DSP is cleaved via a calpain-dependent mechanism. Together this led us to hypothesize that mutant DSP is more susceptible to calpain cleavage, resulting in cellular haploinsufficiency. To define a unifying mechanism for AC-linked variants, we assessed the biomolecular properties of DSP variants. While these variants do not significantly perturb the global structure of DSP, there are significant changes to local stabilizing intramolecular interactions of select variants, which correlates with augmented calpain degradation due to increased exposure of an auto-inhibited calpain-cleavage site. Taken together we have identified a unifying mechanism for select AC-linked variants of DSP grounded in increased susceptibility to calpain-cleavage leading to decreased functional DSP. To test the hypothesis that DSP haploinsufficiency results in poor ID organization and reduced electrical conduction of the myocardium we created engineered heart tissues (EHTs) from induced pluripotent stem cells generated from a symptomatic R451G-positive family member. Surprisingly, we found that conduction velocities were unchanged however, mutant tissues had a significantly slower rate of contraction despite unchanged peak contractile force. This biomechanical deficiency in DSP haploinsuffient cells may indicate poor coupling between the sarcomere lattice and the ID as a fundamental disease mechanism in AC.