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Patient mutations linked to arrhythmogenic cardiomyopathy enhance calpain-mediated desmoplakin degradation
Ronald Ng, … , Maegen A. Ackermann, Stuart G. Campbell
Ronald Ng, … , Maegen A. Ackermann, Stuart G. Campbell
Published June 13, 2019
Citation Information: JCI Insight. 2019;4(14):e128643. https://doi.org/10.1172/jci.insight.128643.
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Research Article Cardiology Genetics

Patient mutations linked to arrhythmogenic cardiomyopathy enhance calpain-mediated desmoplakin degradation

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Abstract

Arrhythmogenic cardiomyopathy (ACM) is an inherited disorder with variable genetic etiologies. Here, we focused on understanding the precise molecular pathology of a single clinical variant in DSP, the gene encoding desmoplakin. We initially identified a potentially novel missense desmoplakin variant (p.R451G) in a patient diagnosed with biventricular ACM. An extensive single-family ACM cohort was assembled, revealing a pattern of coinheritance for R451G desmoplakin and the ACM phenotype. An in vitro model system using patient-derived induced pluripotent stem cell lines showed depressed levels of desmoplakin in the absence of abnormal electrical propagation. Molecular dynamics simulations of desmoplakin R451G revealed no overt structural changes, but a significant loss of intramolecular interactions surrounding a putative calpain target site was observed. Protein degradation assays of recombinant desmoplakin R451G confirmed increased calpain vulnerability. In silico screening identified a subset of 3 additional ACM-linked desmoplakin missense mutations with apparent enhanced calpain susceptibility, predictions that were confirmed experimentally. Similar to R451G, these mutations are found in families with biventricular ACM. We conclude that augmented calpain-mediated degradation of desmoplakin represents a shared pathological mechanism for select ACM-linked missense variants. This approach for identifying variants with shared molecular pathologies may represent a powerful new strategy for understanding and treating inherited cardiomyopathies.

Authors

Ronald Ng, Heather Manring, Nikolaos Papoutsidakis, Taylor Albertelli, Nicole Tsai, Claudia J. See, Xia Li, Jinkyu Park, Tyler L. Stevens, Prameela J. Bobbili, Muhammad Riaz, Yongming Ren, Christopher E. Stoddard, Paul M.L. Janssen, T. Jared Bunch, Stephen P. Hall, Ying-Chun Lo, Daniel L. Jacoby, Yibing Qyang, Nathan Wright, Maegen A. Ackermann, Stuart G. Campbell

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Figure 6

Increased exposure of a calpain target site on desmoplakin due to a glycine substitution at residue 451.

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Increased exposure of a calpain target site on desmoplakin due to a glyc...
(A) Ribbon model of desmoplakin showing location of affected calpain site (resides 447–451). Inset shows intramolecular interactions surrounding the putative calpain site. (B) Immunoblots of lysates from a donor control heart (OSU 294050) for changes in desmoplakin levels following the addition of exogenous Ca2+ or calpain. Addition of 10 mM Ca2+ over endogenous levels results in significant degradation of desmoplakin. Addition of exogenous calpain further reduced levels of desmoplakin at both endogenous levels and increased levels of Ca2+ (*P < 0.05, **P < 0.01 for 2-way ANOVA with Tukey’s multiple comparisons test; n = 3). (C) Histogram of the number of noncovalent interactions surrounding the calpain site, calculated from molecular dynamic simulations of WT desmoplakin (blue) and R451G desmoplakin (red). R451G produced a clear reduction in the average number of noncovalent interactions surrounding the calpain site, possibly increasing solvent accessibility degradation rate. (D) Surface model of desmoplakin showing the exposure of the affected calpain site in WT desmoplakin (left) and R451G desmoplakin (right). Red represents the exposed surface of calpain site residues. (E) Average exposed surface area of the calpain site over the length of the simulation (*P < 0.05, 2-tailed unpaired t test; n = 8). Error bars represent SEM.

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