Comparison of the effects of a truncating and a missense MYBPC3 mutation on contractile parameters of engineered heart tissue
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Comparison of the effects of a truncating and a missense MYBPC3 mutation on contractile parameters of engineered heart tissue. / Wijnker, Paul; Friedrich, Felix W; Dutsch, Alexander; Reischmann, Silke; Eder, Alexandra; Mannhardt, Ingra; Mearini, Giulia; Eschenhagen, Thomas; van der Velden, Jolanda; Carrier, Lucie.
In: J MOL CELL CARDIOL, Vol. 97, 2016, p. 82-92.Research output: SCORING: Contribution to journal › SCORING: Journal article › Research › peer-review
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TY - JOUR
T1 - Comparison of the effects of a truncating and a missense MYBPC3 mutation on contractile parameters of engineered heart tissue
AU - Wijnker, Paul
AU - Friedrich, Felix W
AU - Dutsch, Alexander
AU - Reischmann, Silke
AU - Eder, Alexandra
AU - Mannhardt, Ingra
AU - Mearini, Giulia
AU - Eschenhagen, Thomas
AU - van der Velden, Jolanda
AU - Carrier, Lucie
PY - 2016
Y1 - 2016
N2 - Hypertrophic cardiomyopathy (HCM) is a cardiac genetic disease characterized by left ventricular hypertrophy, diastolic dysfunction and myocardial disarray. The most frequently mutated gene is MYBPC3, encoding cardiac myosin-binding protein-C (cMyBP-C). We compared the pathomechanisms of a truncating mutation (c.2373_2374insG) and a missense mutation (c.1591G>C) in MYBPC3 in engineered heart tissue (EHT). EHTs enable to study the direct effects of mutants without interference of secondary disease-related changes. EHTs were generated from Mybpc3-targeted knock-out (KO) and wild-type (WT) mouse cardiac cells. MYBPC3 WT and mutants were expressed in KO EHTs via adeno-associated virus. KO EHTs displayed higher maximal force and sensitivity to external [Ca2+] than WT EHTs. Expression of WT-Mybpc3 at MOI-100 resulted in ~73% cMyBP-C level but did not prevent the KO phenotype, whereas MOI-300 resulted in ≥95% cMyBP-C level and prevented the KO phenotype. Expression of the truncating or missense mutation (MOI-300) or their combination with WT (MOI-150 each), mimicking the homozygous or heterozygous disease state, respectively, failed to restore force to WT level. Immunofluorescence analysis revealed correct incorporation of WT and missense, but not of truncated cMyBP-C in the sarcomere. In conclusion, this study provides evidence in KO EHTs that i) haploinsufficiency affects EHT contractile function if WTcMyBP-C protein levels are ≤ 73%, ii) missense or truncatingmutations, but not WT do not fully restore the disease phenotype and have different pathogenic mechanisms, e.g. sarcomere poisoning for the missense mutation, iii) the direct impact of (newly identified) MYBPC3 gene variants can be evaluated.
AB - Hypertrophic cardiomyopathy (HCM) is a cardiac genetic disease characterized by left ventricular hypertrophy, diastolic dysfunction and myocardial disarray. The most frequently mutated gene is MYBPC3, encoding cardiac myosin-binding protein-C (cMyBP-C). We compared the pathomechanisms of a truncating mutation (c.2373_2374insG) and a missense mutation (c.1591G>C) in MYBPC3 in engineered heart tissue (EHT). EHTs enable to study the direct effects of mutants without interference of secondary disease-related changes. EHTs were generated from Mybpc3-targeted knock-out (KO) and wild-type (WT) mouse cardiac cells. MYBPC3 WT and mutants were expressed in KO EHTs via adeno-associated virus. KO EHTs displayed higher maximal force and sensitivity to external [Ca2+] than WT EHTs. Expression of WT-Mybpc3 at MOI-100 resulted in ~73% cMyBP-C level but did not prevent the KO phenotype, whereas MOI-300 resulted in ≥95% cMyBP-C level and prevented the KO phenotype. Expression of the truncating or missense mutation (MOI-300) or their combination with WT (MOI-150 each), mimicking the homozygous or heterozygous disease state, respectively, failed to restore force to WT level. Immunofluorescence analysis revealed correct incorporation of WT and missense, but not of truncated cMyBP-C in the sarcomere. In conclusion, this study provides evidence in KO EHTs that i) haploinsufficiency affects EHT contractile function if WTcMyBP-C protein levels are ≤ 73%, ii) missense or truncatingmutations, but not WT do not fully restore the disease phenotype and have different pathogenic mechanisms, e.g. sarcomere poisoning for the missense mutation, iii) the direct impact of (newly identified) MYBPC3 gene variants can be evaluated.
U2 - 10.1016/j.yjmcc.2016.03.003
DO - 10.1016/j.yjmcc.2016.03.003
M3 - SCORING: Journal article
VL - 97
SP - 82
EP - 92
JO - J MOL CELL CARDIOL
JF - J MOL CELL CARDIOL
SN - 0022-2828
ER -