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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 8

Calpain inhibition results in partial rescue of desmoplakin levels in homozygous DSP R451G engineered heart tissues.

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Calpain inhibition results in partial rescue of desmoplakin levels in ho...
(A) Engineered heart tissues were incubated with 25 μM of the calpain inhibitor MDL-28170(MDL) or a vehicle control (DMSO) for 72 hours and then probed for desmoplakin protein levels. Loading across gels was normalized by sarcomeric actin. (Gel images are representative of PGP1 MDL n = 10; PGP1 DMSO, n = 8; HO35 MDL, n = 8; HO35 DMSO, n = 8). (B) Quantification of immunoblot showing significant interaction between genotype and drug treatment (P < 0.05) as well as significant loss of DSP between genotype (*P < 0.05) and drug treatment for 2-way ANOVA with Tukey’s multiple comparisons test. While desmoplakin levels in WT/WT EHTs were significantly decreased in samples exposed to MDL, this decrease was not observed in R451G/R451G EHTs.

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