TY - JOUR
T1 - S100A4 as a target of the E3-ligase Asb2β and its effect on engineered heart tissue
AU - Braumann, Simon
AU - Thottakara, Tilo
AU - Stücker, Sabrina
AU - Reischmann-Düsener, Silke
AU - Krämer, Elisabeth
AU - Groß, Julia
AU - Hirt, Marc N.
AU - Doroudgar, Shirin
AU - Carrier, Lucie
AU - Friedrich, Felix W.
N1 - Funding Information:
This work was part of the Graduiertenkolleg Individualized Cardiovascular Medicine of the CVRC (Cardiovascular Research Center) and supported by the DZHK (German Centre for Cardiovascular Research). SD and JG were supported by an Excellence Grant from the German Centre for Cardiovascular Research (DZHK), and Department of Cardiology, Angiology, and Pneumology, University Hospital Heidelberg. We thank Gisèle Bonne (Sorbonne Université, INSERM UMRS_974, Paris) for providing us with animal samples and we thank Ingke Braren from the Vector Core Facility of the UKE for production of the AAV6.
Publisher Copyright:
© 2018 Braumann, Thottakara, Stücker, Reischmann-Düsener, Krämer, Groß, Hirt, Doroudgar, Carrier and Friedrich.
PY - 2018/9/19
Y1 - 2018/9/19
N2 - Background: S100A4 has recently emerged as an important player in cardiac disease, affecting phenotype development in animal models of myocardial infarction and pathological cardiac hypertrophy, albeit it is unclear whether S100A4 exerts a detrimental or beneficial function. The goal of the current study was to analyze S100A4 expression in models of cardiac pathology, investigate its degradation by the ubiquitin-proteasome system (UPS), and furthermore examine the functional effects of S100A4 levels in a 3D model of engineered heart tissue (EHT). Methods and Results: S100A4 mRNA and protein levels were analyzed in different models of cardiac pathology via quantitative RT-PCR and Western blot, showing a higher S100A4 steady-state protein concentration in hearts of Mybpc3-knock-in (KI) hypertrophic cardiomyopathy (HCM) mice. COS-7 cells co-transfected with plasmids encoding mutant (MUT) Asb2β lacking the E3 ligase activity in combination with V5-tagged S100A4 plasmid presented higher S100A4-V5 protein steady-state concentrations than cells co-transfected with the Asb2β wild type (WT) plasmid. This effect was blunted by treatment with the specific proteasome inhibitor epoxomicin. Adeno-associated virus serotype 6 (AAV6)-mediated S100A4 overexpression in a 3D model of EHT did not affect contractile parameters. Immunofluorescence analysis showed a cytosolic and partly nuclear expression pattern of S100A4. Gene expression analysis in EHTs overexpressing S100A4-V5 showed markedly lower steady-state concentrations of genes involved in cardiac fibrosis and pathological cardiac hypertrophy. Conclusion: We showed that S100A4 protein level is higher in cardiac tissue of Mybpc3-KI HCM mice probably as a result of a lower degradation by the E3 ligase Asb2β. While an overexpression of S100A4 did not alter contractile parameters in EHTs, downstream gene expression analysis points toward modulation of signaling cascades involved in fibrosis and hypertrophy.
AB - Background: S100A4 has recently emerged as an important player in cardiac disease, affecting phenotype development in animal models of myocardial infarction and pathological cardiac hypertrophy, albeit it is unclear whether S100A4 exerts a detrimental or beneficial function. The goal of the current study was to analyze S100A4 expression in models of cardiac pathology, investigate its degradation by the ubiquitin-proteasome system (UPS), and furthermore examine the functional effects of S100A4 levels in a 3D model of engineered heart tissue (EHT). Methods and Results: S100A4 mRNA and protein levels were analyzed in different models of cardiac pathology via quantitative RT-PCR and Western blot, showing a higher S100A4 steady-state protein concentration in hearts of Mybpc3-knock-in (KI) hypertrophic cardiomyopathy (HCM) mice. COS-7 cells co-transfected with plasmids encoding mutant (MUT) Asb2β lacking the E3 ligase activity in combination with V5-tagged S100A4 plasmid presented higher S100A4-V5 protein steady-state concentrations than cells co-transfected with the Asb2β wild type (WT) plasmid. This effect was blunted by treatment with the specific proteasome inhibitor epoxomicin. Adeno-associated virus serotype 6 (AAV6)-mediated S100A4 overexpression in a 3D model of EHT did not affect contractile parameters. Immunofluorescence analysis showed a cytosolic and partly nuclear expression pattern of S100A4. Gene expression analysis in EHTs overexpressing S100A4-V5 showed markedly lower steady-state concentrations of genes involved in cardiac fibrosis and pathological cardiac hypertrophy. Conclusion: We showed that S100A4 protein level is higher in cardiac tissue of Mybpc3-KI HCM mice probably as a result of a lower degradation by the E3 ligase Asb2β. While an overexpression of S100A4 did not alter contractile parameters in EHTs, downstream gene expression analysis points toward modulation of signaling cascades involved in fibrosis and hypertrophy.
KW - Engineered heart tissue
KW - Fibrosis
KW - Hypertrophic cardiomyopathy
KW - S100A4
KW - Ubiquitin proteasome system
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U2 - 10.3389/fphys.2018.01292
DO - 10.3389/fphys.2018.01292
M3 - Article
AN - SCOPUS:85055157573
SN - 1664-042X
VL - 9
JO - Frontiers in Physiology
JF - Frontiers in Physiology
IS - SEP
M1 - 1292
ER -