Inhibition of Coxsackievirus-associated dystrophin cleavage prevents cardiomyopathy
Byung-Kwan Lim, … , Ju Chen, Kirk U. Knowlton
Byung-Kwan Lim, … , Ju Chen, Kirk U. Knowlton
Published November 8, 2013
Citation Information: J Clin Invest. 2013;123(12):5146-5151. https://doi.org/10.1172/JCI66271.
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Inhibition of Coxsackievirus-associated dystrophin cleavage prevents cardiomyopathy

Abstract

Heart failure in children and adults is often the consequence of myocarditis associated with Coxsackievirus (CV) infection. Upon CV infection, enteroviral protease 2A cleaves a small number of host proteins including dystrophin, which links actin filaments to the plasma membrane of muscle fiber cells (sarcolemma). It is unknown whether protease 2A–mediated cleavage of dystrophin and subsequent disruption of the sarcolemma play a role in CV-mediated myocarditis. We generated knockin mice harboring a mutation at the protease 2A cleavage site of the dystrophin gene, which prevents dystrophin cleavage following CV infection. Compared with wild-type mice, we found that mice expressing cleavage-resistant dystrophin had a decrease in sarcolemmal disruption and cardiac virus titer following CV infection. In addition, cleavage-resistant dystrophin inhibited the cardiomyopathy induced by cardiomyocyte-restricted expression of the CV protease 2A transgene. These findings indicate that protease 2A–mediated cleavage of dystrophin is critical for viral propagation, enteroviral-mediated cytopathic effects, and the development of cardiomyopathy.

Authors

Byung-Kwan Lim, Angela K. Peter, Dingding Xiong, Anna Narezkina, Aaron Yung, Nancy D. Dalton, Kyung-Kuk Hwang, Toshitaka Yajima, Ju Chen, Kirk U. Knowlton

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Figure 1

Generation of enteroviral protease 2A cleavage–resistant DysKI mice.

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Generation of enteroviral protease 2A cleavage–resistant DysKI mice. (...
(A) Genomic structure and design of DysKI mouse. We changed the wild-type exon 50 (E50) amino acid residues 2425 and 2426 from leucine to proline (L2425P) and serine to aspartic acid (S2426D), respectively. (B) Confirmation of DysKI recombination by Southern blotting of mouse embryonic stem cells. Genomic DNA isolated from embryonic stem cells was used to confirm that homologous recombination had occurred as opposed to random insertion. Southern blotting with probe 1 was performed with a BglII digest, demonstrating loss of the wild-type genome at 9.8 kb (left panel). Female mice were genotyped using PCR primers P1 and P2 (right panel). Data are shown for mice with wild-type dystrophin (DysWT), heterozygous dystrophin knockin (Dys+/KI), and homozygous dystrophin knockin (DysKI). (C) Sequence analysis of exon 50 showing replacement of nucleotides in DysKI mouse genomic DNA as expected. (D) Western blot showing dystrophin in embryonic cardiac myocytes 24 hours after CVB3 infection, in which the cells were isolated from DysWT and DysKI mice. Dystrophin cleavage was inhibited in myocytes from DysKI mice. Comparable levels of viral infection of the cells are demonstrated by the presence of the CVB3 capsid protein VP1.