Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Article
  • Published:

Forced activation of dystrophin transcription by CRISPR/dCas9 reduced arrhythmia susceptibility via restoring membrane Nav1.5 distribution

Gene Therapy volume 30, pages 142–149 (2023)Cite this article

Subjects

Abstract

Dystrophin deficiency due to genetic mutations causes cardiac abnormalities in Duchenne’s muscular dystrophy. Dystrophin is also shown to be downregulated in conventional failing hearts. Whether restoration of dystrophin expression possesses any therapeutic potential for conventional heart failure (HF) remains to be examined. HF mouse model was generated by transverse aortic constriction (TAC). In vivo activation of dystrophin transcription was achieved by tail-vein injection of adeno-associated virus 9 carrying CRISPR/dCas system for dystrophin. We found that activation of dystrophin expression in TAC mice significantly reduced the susceptibility to arrhythmia of TAC mice and the mortality rate. We further demonstrated that over-expression of dystrophin increased cardiac conduction of hearts in TAC mice by optical mapping evaluation. Activation of dystrophin expression also increased peak sodium current in isolated ventricular myocytes from hearts of TAC mice as recorded by the patch-clamp technique. Immunoblotting and immunofluorescence showed that increased dystrophin transcription restored the membrane distribution of Nav1.5 in the hearts of TAC mice. In summary, correction of dystrophin downregulation by the CRISPR-dCas9 system reduced the susceptibility to arrhythmia of conventional HF mice through restoring Nav1.5 membrane distribution. This study paved the way to develop a new therapeutic strategy for HF-related ventricular arrhythmia.

This is a preview of subscription content, access via your institution

Access options

Buy this article

  • Purchase on Springer Link
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Fig. 1: Forced activation of dystrophin transcription by CRISPR/dCas9 achieved the compensation of downregulated dystrophin in TAC mice.
Fig. 2: Activation of dystrophin transcription reduced the susceptibility to arrhythmia of TAC mice.
Fig. 3: Activation of dystrophin transcription normalized cardiac conduction of hearts from TAC mice.
Fig. 4: Activation of dystrophin transcription restored sodium currents and increased membrane distribution of Nav1.5 in ventricular myocytes of hearts from TAC mice.

Similar content being viewed by others

Data availability

The data that support the findings of this study are available from the corresponding authors upon reasonable request.

References

  1. Hoffman EP, Brown RH Jr, Kunkel LM. Dystrophin: the protein product of the Duchenne muscular dystrophy locus. Cell. 1987;51:919–28.

    Article  CAS  PubMed  Google Scholar 

  2. Koenig M, Hoffman EP, Bertelson CJ, Monaco AP, Feener C, Kunkel LM. Complete cloning of the Duchenne muscular dystrophy (DMD) cDNA and preliminary genomic organization of the DMD gene in normal and affected individuals. Cell. 1987;50:509–17.

    Article  CAS  PubMed  Google Scholar 

  3. Rybakova IN, Patel JR, Ervasti JM. The dystrophin complex forms a mechanically strong link between the sarcolemma and costameric actin. J Cell Biol. 2000;150:1209–14.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  4. Muntoni F, Torelli S, Ferlini A. Dystrophin and mutations: one gene, several proteins, multiple phenotypes. Lancet Neurol. 2003;2:731–40.

    Article  CAS  PubMed  Google Scholar 

  5. Koenig X, Ebner J, Hilber K. Voltage-dependent sarcolemmal ion channel abnormalities in the dystrophin-deficient heart. Int J Mol Sci. 2018;19:3296.

  6. Min YL, Bassel-Duby R, Olson EN. CRISPR correction of Duchenne muscular dystrophy. Annu Rev Med. 2019;70:239–55.

    Article  CAS  PubMed  Google Scholar 

  7. Yoshida H, Takahashi M, Koshimizu M, Tanonaka K, Oikawa R, Toyo-oka T, et al. Decrease in sarcoglycans and dystrophin in failing heart following acute myocardial infarction. Cardiovasc Res. 2003;59:419–27.

    Article  CAS  PubMed  Google Scholar 

  8. Prado FP, Dos Santos DO, Blefari V, Silva CA, Machado J, Kettelhut IDC, et al. Early dystrophin loss is coincident with the transition of compensated cardiac hypertrophy to heart failure. PLoS One. 2017;12:e0189469.

    Article  PubMed  PubMed Central  Google Scholar 

  9. Kawada T, Masui F, Tezuka A, Ebisawa T, Kumagai H, Nakazawa M, et al. A novel scheme of dystrophin disruption for the progression of advanced heart failure. Biochim Biophys Acta. 2005;1751:73–81.

    Article  CAS  PubMed  Google Scholar 

  10. Wang TJ, Larson MG, Levy D, Vasan RS, Leip EP, Wolf PA, et al. Temporal relations of atrial fibrillation and congestive heart failure and their joint influence on mortality: the Framingham Heart Study. Circulation. 2003;107:2920–5.

    Article  PubMed  Google Scholar 

  11. Schoger E, Carroll KJ, Iyer LM, McAnally JR, Tan W, Liu N, et al. CRISPR-mediated activation of endogenous gene expression in the postnatal heart. Circ Res. 2020;126:6–24.

    Article  CAS  PubMed  Google Scholar 

  12. Gilbert LA, Larson MH, Morsut L, Liu Z, Brar GA, Torres SE, et al. CRISPR-mediated modular RNA-guided regulation of transcription in eukaryotes. Cell. 2013;154:442–51.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  13. Shirakabe A, Zhai P, Ikeda Y, Saito T, Maejima Y, Hsu CP, et al. Drp1-dependent mitochondrial autophagy plays a protective role against pressure overload-induced mitochondrial dysfunction and heart failure. Circulation. 2016;133:1249–63.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  14. Maeder ML, Linder SJ, Cascio VM, Fu Y, Ho QH, Joung JK. CRISPR RNA-guided activation of endogenous human genes. Nat Methods. 2013;10:977–9.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  15. Li DS, Xue GL, Yang JM, Li CZ, Zhang RX, Tian T, et al. Knockout of interleukin-17A diminishes ventricular arrhythmia susceptibility in diabetic mice via inhibiting NF-kappaB-mediated electrical remodeling. Acta Pharmacol Sin. 2022;43:307–15.

  16. Zhang Y, Sun L, Xuan L, Pan Z, Hu X, Liu H, et al. Long non-coding RNA CCRR controls cardiac conduction via regulating intercellular coupling. Nat Commun. 2018;9:4176.

    Article  PubMed  PubMed Central  Google Scholar 

  17. Zhan G, Wang F, Ding YQ, Li XH, Li YX, Zhao ZR, et al. Rutaecarpine targets hERG channels and participates in regulating electrophysiological properties leading to ventricular arrhythmia. J Cell Mol Med. 2021;25:4938–49.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  18. Wang Y, Hill JA. Electrophysiological remodeling in heart failure. J Mol Cell Cardiol. 2010;48:619–32.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  19. Kleber AG, Rudy Y. Basic mechanisms of cardiac impulse propagation and associated arrhythmias. Physiol Rev. 2004;84:431–88.

    Article  CAS  PubMed  Google Scholar 

  20. Petitprez S, Zmoos AF, Ogrodnik J, Balse E, Raad N, El-Haou S, et al. SAP97 and dystrophin macromolecular complexes determine two pools of cardiac sodium channels Nav1.5 in cardiomyocytes. Circ Res. 2011;108:294–304.

    Article  CAS  PubMed  Google Scholar 

  21. Ousterout DG, Kabadi AM, Thakore PI, Majoros WH, Reddy TE, Gersbach CA. Multiplex CRISPR/Cas9-based genome editing for correction of dystrophin mutations that cause Duchenne muscular dystrophy. Nat Commun. 2015;6:6244.

    Article  CAS  PubMed  Google Scholar 

  22. Amoasii L, Hildyard JCW, Li H, Sanchez-Ortiz E, Mireault A, Caballero D, et al. Gene editing restores dystrophin expression in a canine model of Duchenne muscular dystrophy. Science. 2018;362:86–91.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  23. Bengtsson NE, Hall JK, Odom GL, Phelps MP, Andrus CR, Hawkins RD, et al. Muscle-specific CRISPR/Cas9 dystrophin gene editing ameliorates pathophysiology in a mouse model for Duchenne muscular dystrophy. Nat Commun. 2017;8:14454.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  24. Armoundas AA, Wu R, Juang G, Marban E, Tomaselli GF. Electrical and structural remodeling of the failing ventricle. Pharmacol Ther. 2001;92:213–30.

    Article  CAS  PubMed  Google Scholar 

  25. Wu QQ, Xiao Y, Yuan Y, Ma ZG, Liao HH, Liu C, et al. Mechanisms contributing to cardiac remodelling. Clin Sci (Lond). 2017;131:2319–45.

    Article  CAS  PubMed  Google Scholar 

  26. Cutler MJ, Jeyaraj D, Rosenbaum DS. Cardiac electrical remodeling in health and disease. Trends Pharmacol Sci. 2011;32:174–80.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  27. Han F, Lu YM, Hasegawa H, Kanai H, Hachimura E, Shirasaki Y. et al. Inhibition of dystrophin breakdown and endothelial nitric-oxide synthase uncoupling accounts for cytoprotection by 3-[2-[4-(3-chloro-2-methylphenyl)-1-piperazinyl]ethyl]-5,6-dimethoxy-1-(4-imidazo lylmethyl)-1H-indazole dihydrochloride 3.5 hydrate (DY-9760e) in left ventricular hypertrophied mice. J Pharmacol Exp Ther. 2010;332:421–8.

    Article  CAS  PubMed  Google Scholar 

  28. Moretti A, Fonteyne L, Giesert F, Hoppmann P, Meier AB, Bozoglu T, et al. Somatic gene editing ameliorates skeletal and cardiac muscle failure in pig and human models of Duchenne muscular dystrophy. Nat Med. 2020;26:207–14.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  29. Luo L, Ning F, Du Y, Song B, Yang D, Salvage SC, et al. Calcium-dependent Nedd4-2 upregulation mediates degradation of the cardiac sodium channel Nav1.5: implications for heart failure. Acta Physiol (Oxf). 2017;221:44–58.

    Article  CAS  PubMed  Google Scholar 

  30. Shy D, Gillet L, Ogrodnik J, Albesa M, Verkerk AO, Wolswinkel R, et al. PDZ domain-binding motif regulates cardiomyocyte compartment-specific NaV1.5 channel expression and function. Circulation. 2014;130:147–60.

    Article  CAS  PubMed  Google Scholar 

  31. Gavillet B, Rougier JS, Domenighetti AA, Behar R, Boixel C, Ruchat P, et al. Cardiac sodium channel Nav1.5 is regulated by a multiprotein complex composed of syntrophins and dystrophin. Circ Res. 2006;99:407–14.

    Article  CAS  PubMed  Google Scholar 

  32. Rougier JS, Gavillet B, Abriel H. Proteasome inhibitor (MG132) rescues Nav1.5 protein content and the cardiac sodium current in dystrophin-deficient mdx (5cv) mice. Front Physiol. 2013;4:51.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  33. Colasante G, Lignani G, Brusco S, Di Berardino C, Carpenter J, Giannelli S, et al. dCas9-based Scn1a gene activation restores inhibitory interneuron excitability and attenuates seizures in Dravet syndrome mice. Mol Ther. 2020;28:235–53.

    Article  CAS  PubMed  Google Scholar 

  34. Liu XS, Wu H, Krzisch M, Wu X, Graef J, Muffat J, et al. Rescue of Fragile X syndrome neurons by DNA methylation editing of the FMR1 gene. Cell. 2018;172:979–92.e6.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

Download references

Funding

This work was supported by National Natural Science Foundation of China (82070344, 81870295 to ZP) and HMU Marshal Initiative Funding (HMUMIF-21017 to ZP).

Author information

Author notes

  1. These authors contributed equally: Ruixin Zhang, Junwu Liu, Genlong Xue.

Authors and Affiliations

  1. Department of Pharmacology (The Key Laboratory of Cardiovascular Research, Ministry of Education) at College of Pharmacy, Harbin Medical University, Harbin, 150086, China

    Ruixin Zhang, Junwu Liu, Genlong Xue, Jiming Yang, Desheng Li, Tao Tian, Xiaofang Zhang, Kangyi Gao & Zhenwei Pan

  2. Institute of Heart and Vascular Diseases, Department of Cardiology, and Central Laboratory, the First Affiliated Hospital of Dalian Medical University, Dalian, China

    Genlong Xue

  3. Research Unit of Noninfectious Chronic Diseases in Frigid Zone, Chinese Academy of Medical Sciences, Beijing, 2019RU070, China

    Zhenwei Pan

  4. NHC Key Laboratory of Cell Transplantation, Harbin Medical University, Harbin, China

    Zhenwei Pan

Authors
  1. Ruixin Zhang
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Junwu Liu
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Genlong Xue
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Jiming Yang
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Desheng Li
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Tao Tian
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Xiaofang Zhang
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Kangyi Gao
    View author publications

    You can also search for this author in PubMed Google Scholar

  9. Zhenwei Pan
    View author publications

    You can also search for this author in PubMed Google Scholar

Contributions

RZ, JL, and GX performed experiments, analyzed data, and prepared the manuscript. JY, DL, TT, XZ, and KG helped perform experiments and collect data. ZP designed the project, oversaw the experiments, and prepared the manuscript.

Corresponding author

Correspondence to Zhenwei Pan.

Ethics declarations

Competing interests

The authors declare no competing interests.

Ethics approval

Compliance with ethical standards.

Additional information

Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Zhang, R., Liu, J., Xue, G. et al. Forced activation of dystrophin transcription by CRISPR/dCas9 reduced arrhythmia susceptibility via restoring membrane Nav1.5 distribution. Gene Ther 30, 142–149 (2023). https://doi.org/10.1038/s41434-022-00348-z

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/s41434-022-00348-z

Search

Advanced search

Quick links