Integration of Cardiac Actin Mutants Causing Hypertrophic (p.A295S) and Dilated Cardiomyopathy (p.R312H and p.E361G) into Cellular Structures
The human mutant cardiac α-actins p.A295S or p.R312H and p.E361G, correlated with hypertrophic or dilated cardiomyopathy, respectively, were expressed by the <i>baculovirus/Sf21</i> insect cell system and purified to homogeneity. The purified cardiac actins maintained their native state...
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MDPI AG
2021-07-01
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| Series: | Antioxidants |
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| Online Access: | https://www.mdpi.com/2076-3921/10/7/1082 |
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| author | Constanze Erdmann Roua Hassoun Sebastian Schmitt Carlos Kikuti Anne Houdusse Antonina J. Mazur Andreas Mügge Nazha Hamdani Matthias Geyer Kornelia Jaquet Hans Georg Mannherz |
| author_facet | Constanze Erdmann Roua Hassoun Sebastian Schmitt Carlos Kikuti Anne Houdusse Antonina J. Mazur Andreas Mügge Nazha Hamdani Matthias Geyer Kornelia Jaquet Hans Georg Mannherz |
| author_sort | Constanze Erdmann |
| collection | DOAJ |
| description | The human mutant cardiac α-actins p.A295S or p.R312H and p.E361G, correlated with hypertrophic or dilated cardiomyopathy, respectively, were expressed by the <i>baculovirus/Sf21</i> insect cell system and purified to homogeneity. The purified cardiac actins maintained their native state but showed differences in Ca<sup>2+</sup>-sensitivity to stimulate the myosin-subfragment1 ATPase. Here we analyzed the interactions of these c-actins with actin-binding and -modifying proteins implicated in cardiomyocyte differentiation. We demonstrate that Arp2/3 complex and the formin mDia3 stimulated the polymerization rate and extent of the c-actins, albeit to different degrees. In addition, we tested the effect of the MICAL-1 monooxygenase, which modifies the supramolecular actin organization during development and adaptive processes. MICAL-1 oxidized these c-actin variants and induced their de-polymerization, albeit at different rates. Transfection experiments using MDCK cells demonstrated the preferable incorporation of wild type and p.A295S c-actins into their microfilament system but of p.R312H and p.E361G actins into the submembranous actin network. Transduction of neonatal rat cardiomyocytes with adenoviral constructs coding HA-tagged c-actin variants showed their incorporation into microfilaments after one day in culture and thereafter into thin filaments of nascent sarcomeric structures at their plus ends (Z-lines) except the p.E361G mutant, which preferentially incorporated at the minus ends. |
| first_indexed | 2024-03-10T09:48:25Z |
| format | Article |
| id | doaj.art-eb753d4bf5df46adb87a057ae7098888 |
| institution | Directory Open Access Journal |
| issn | 2076-3921 |
| language | English |
| last_indexed | 2024-03-10T09:48:25Z |
| publishDate | 2021-07-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | Antioxidants |
| spelling | doaj.art-eb753d4bf5df46adb87a057ae70988882023-11-22T03:05:06ZengMDPI AGAntioxidants2076-39212021-07-01107108210.3390/antiox10071082Integration of Cardiac Actin Mutants Causing Hypertrophic (p.A295S) and Dilated Cardiomyopathy (p.R312H and p.E361G) into Cellular StructuresConstanze Erdmann0Roua Hassoun1Sebastian Schmitt2Carlos Kikuti3Anne Houdusse4Antonina J. Mazur5Andreas Mügge6Nazha Hamdani7Matthias Geyer8Kornelia Jaquet9Hans Georg Mannherz10Department of Anatomy and Molecular Embryology, Medical Faculty, Ruhr-University Bochum, D-44780 Bochum, GermanyInstitut für Forschung und Lehre (IFL), Molecular and Experimental Cardiology, Ruhr University Bochum, D-44780 Bochum, GermanyInstitute of Structural Biology, University of Bonn, D-53127 Bonn, GermanyInstitut Curie, Structural Motility Team, F-75005 Paris, FranceInstitut Curie, Structural Motility Team, F-75005 Paris, FranceDepartment of Cell Pathology, Faculty of Biotechnology, University of Wroclaw, Pl-50-383 Wroclaw, PolandInstitut für Forschung und Lehre (IFL), Molecular and Experimental Cardiology, Ruhr University Bochum, D-44780 Bochum, GermanyInstitut für Forschung und Lehre (IFL), Molecular and Experimental Cardiology, Ruhr University Bochum, D-44780 Bochum, GermanyInstitute of Structural Biology, University of Bonn, D-53127 Bonn, GermanyInstitut für Forschung und Lehre (IFL), Molecular and Experimental Cardiology, Ruhr University Bochum, D-44780 Bochum, GermanyDepartment of Anatomy and Molecular Embryology, Medical Faculty, Ruhr-University Bochum, D-44780 Bochum, GermanyThe human mutant cardiac α-actins p.A295S or p.R312H and p.E361G, correlated with hypertrophic or dilated cardiomyopathy, respectively, were expressed by the <i>baculovirus/Sf21</i> insect cell system and purified to homogeneity. The purified cardiac actins maintained their native state but showed differences in Ca<sup>2+</sup>-sensitivity to stimulate the myosin-subfragment1 ATPase. Here we analyzed the interactions of these c-actins with actin-binding and -modifying proteins implicated in cardiomyocyte differentiation. We demonstrate that Arp2/3 complex and the formin mDia3 stimulated the polymerization rate and extent of the c-actins, albeit to different degrees. In addition, we tested the effect of the MICAL-1 monooxygenase, which modifies the supramolecular actin organization during development and adaptive processes. MICAL-1 oxidized these c-actin variants and induced their de-polymerization, albeit at different rates. Transfection experiments using MDCK cells demonstrated the preferable incorporation of wild type and p.A295S c-actins into their microfilament system but of p.R312H and p.E361G actins into the submembranous actin network. Transduction of neonatal rat cardiomyocytes with adenoviral constructs coding HA-tagged c-actin variants showed their incorporation into microfilaments after one day in culture and thereafter into thin filaments of nascent sarcomeric structures at their plus ends (Z-lines) except the p.E361G mutant, which preferentially incorporated at the minus ends.https://www.mdpi.com/2076-3921/10/7/1082ATPaseArp2/3 complexcardiac actincardiomyopathiesMICAL |
| spellingShingle | Constanze Erdmann Roua Hassoun Sebastian Schmitt Carlos Kikuti Anne Houdusse Antonina J. Mazur Andreas Mügge Nazha Hamdani Matthias Geyer Kornelia Jaquet Hans Georg Mannherz Integration of Cardiac Actin Mutants Causing Hypertrophic (p.A295S) and Dilated Cardiomyopathy (p.R312H and p.E361G) into Cellular Structures Antioxidants ATPase Arp2/3 complex cardiac actin cardiomyopathies MICAL |
| title | Integration of Cardiac Actin Mutants Causing Hypertrophic (p.A295S) and Dilated Cardiomyopathy (p.R312H and p.E361G) into Cellular Structures |
| title_full | Integration of Cardiac Actin Mutants Causing Hypertrophic (p.A295S) and Dilated Cardiomyopathy (p.R312H and p.E361G) into Cellular Structures |
| title_fullStr | Integration of Cardiac Actin Mutants Causing Hypertrophic (p.A295S) and Dilated Cardiomyopathy (p.R312H and p.E361G) into Cellular Structures |
| title_full_unstemmed | Integration of Cardiac Actin Mutants Causing Hypertrophic (p.A295S) and Dilated Cardiomyopathy (p.R312H and p.E361G) into Cellular Structures |
| title_short | Integration of Cardiac Actin Mutants Causing Hypertrophic (p.A295S) and Dilated Cardiomyopathy (p.R312H and p.E361G) into Cellular Structures |
| title_sort | integration of cardiac actin mutants causing hypertrophic p a295s and dilated cardiomyopathy p r312h and p e361g into cellular structures |
| topic | ATPase Arp2/3 complex cardiac actin cardiomyopathies MICAL |
| url | https://www.mdpi.com/2076-3921/10/7/1082 |
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