Clin Res Cardiol (2022). https://doi.org/10.1007/s00392-022-02002-5

Protective Role of Heat Shock Protein A4 (HSPA4) in the Heart

D. Marques Rodrigues1, M. El Kenani2, S. Gersch1, E. T. Sadrach2, S. Lutz3, I. M. Adham4, S. Engelhardt5, G. Hasenfuß1, K. Toischer1, B. A. Mohamed1, für die Studiengruppe: DZHK

1Herzzentrum, Klinik für Kardiologie und Pneumologie, Universitätsmedizin Göttingen, Göttingen; 2Department of Cardiology and Pneumology, University Medical Center Goettingen, Göttingen; 3Institut für Pharmakologie und Toxikologie, Universitätsmedizin Göttingen, Göttingen; 4Institut für Humangenetik, Universitätsmedizin Göttingen, Göttingen; 5Institut für Pharmakologie und Toxikologie, Technische Universität München (TUM), München;

Introduction Myopathies and myocardial dysfunction in cardiac muscle may be caused by an increase of stress due to misfolded proteins in the cell as well as proteinopathies from aggregates. The major chaperone proteins involved homeostasis in the mammalian cell are the Heat Shock Protein-70 complex functioning as an adaptor and transporter of misfolded proteins. HSPA4 is a co-chaperone and nucleotide exchange factor in the HSC-70 chaperone complex, possibly functioning as a rate limiter in the stabilization process. Our previous study on HSPA4 knockout mice showed pathological remodeling of the heart with an underlying hypertrophy and fibrosis but preserved heart function. Additionally, an aggregation of polyubiquitinylated proteins but functional proteasome was present. On the other hand, an accumulation of autophagosomal markers P62/Sequestosome1 and LC3BII in the knockout animals makes an increase in autophagic processes prominent and a target for further investigation. The primary aim of the study is to elucidate the role of the co-chaperone HSPA4 in pathological cardiac remodeling as well as protein quality control of the cell and if a genetic manipulation of the expression in cardiomyocytes can improve protein homeostasis, reduce DOX induced cardiotoxicity and affect remodeling in pressure overload. Results Global HSPA4-KO mice showed 733% Overexpression of HSPA4 in KO mice which rescued the phenotype of cardiac hypertrophy (LVAW;d WT: 0,9625mm -KO:1,134mm p=<0,0001 - OE: 0,9900mm p=0,4086); fibrosis(rel. fibrotic area WT: 0,8276% - KO: 6,463% p=<0,0001 - OE: 2,053% p=0,3260) at an age of 4 months of the animals. Furthermore, the polyubiquitination of misfolded proteins(relative expression WT: 1,0 – KO:2,613 p=<0,0001 -  OE: 1,047 p=0,9806) was lowered and autophagosomal marker P62/Sequestosome1(relative expression WT: 1,0 – KO:4,016 p=0,0044 -  OE: 0,8873 p=0,9878) was decreased  manifesting an important role of HSPA in Protein Quality Control in cardiomyocytes and not other cell types of the heart. We next tested HSPA4 overexpression as a therapeutical strategy in a mouse model of proteotoxic stress with the drug Adriamycin(Doxorubicin). Here we could show that the overexpression of HSPA4 ameliorated the atrophic effects on cell size (MFD Control: 9,567µm – DOX: 8,810µm p=0,0026 OE: 9,160µm p=0,1009) and transcription factor TRIM63(relative mRNA Control: 1,0 – DOX:1,933 p=0,0013 -  OE: 1,191 p=0,6216) as well as ventricular wall thickness(LVAW;d Control: 0,7825mm – DOX: 0,6976mm p= 0,0009 – OE:0,7420mm p=0,1282). Additionally, the autophagic process was improved with lowered P62/Sequestosome1 levels (relative expression Control: 1,0 – DOX: 2,230 p=0,0068 – OE: 1,437 p=0,4065) and LC3BII (relative expression Control: 1,0 – DOX: 4,245 p=0,0005 – OE: 1,727 p=0,5153) Conclusion Protein Quality Control is an essential process in the maintenance in cardiomyocytes, especially under stress due to increased workload of the heart or cardiotoxic chemotherapy. We demonstrated that HSPA4 is a crucial factor in upholding homeostasis of proteins in the heart most likely due to improvement in chaperone function in the cell. We could also show that overexpressing HSPA4 via gene therapy could be beneficial in lowering the cardiotoxic side effects of the common chemotherapeutic doxorubicin, making it a promising new treatment for the future.

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