Combinatorial genetic replenishments in myocardial and outflow tract tissues restore heart function in tnnt2 mutant zebrafish
Lian Liu,
Fei Fei,
Ranran Zhang,
Fang Wu,
Qian Yang,
Feng Wang,
Shaoyang Sun,
Hui Zhao,
Qiang Li,
Lei Wang,
Youhua Wang,
Yonghao Gui,
Xu Wang
Affiliations
Lian Liu
Department of Cardiology, Children's Hospital of Fudan University, Shanghai 201102, China
Fei Fei
Cancer Metabolism Laboratory, Cancer Institute, Fudan University Shanghai Cancer Center, Shanghai 200032, China
Ranran Zhang
Department of Pediatrics, the Affiliated Hospital of Qingdao University, Qingdao, Shangdong 266003, China
Fang Wu
Department of Cardiology, Children's Hospital of Fudan University, Shanghai 201102, China
Qian Yang
Department of Cardiology, Children's Hospital of Fudan University, Shanghai 201102, China
Feng Wang
Department of Cardiology, Children's Hospital of Fudan University, Shanghai 201102, China
Shaoyang Sun
Key Laboratory of Metabolism and Molecular Medicine, Ministry of Education, Department of Biochemistry and Molecular Biology, School of Basic Medical Sciences, Fudan University, Shanghai 230002, China
Hui Zhao
Longhua Hospital, Shanghai University of Traditional Chinese Medicine, Shanghai 200032, China
Qiang Li
Translational Medical Center for Development and Disease, Shanghai Key Laboratory of Birth Defect, Institute of Pediatrics, Children's Hospital of Fudan University, Shanghai 201102, China
Lei Wang
Key Laboratory of Metabolism and Molecular Medicine, Ministry of Education, Department of Biochemistry and Molecular Biology, School of Basic Medical Sciences, Fudan University, Shanghai 230002, China
Youhua Wang
Longhua Hospital, Shanghai University of Traditional Chinese Medicine, Shanghai 200032, China
Yonghao Gui
Department of Cardiology, Children's Hospital of Fudan University, Shanghai 201102, China
Xu Wang
Cancer Metabolism Laboratory, Cancer Institute, Fudan University Shanghai Cancer Center, Shanghai 200032, China
Cardiac muscle troponin T (Tnnt2) mediates muscle contraction in response to calcium ion dynamics, and Tnnt2 mutations are associated with multiple types of cardiomyopathy. Here, we employed a zebrafish model to investigate the genetic replenishment strategies of using conditional and inducible promoters to rescue the deficiencies in the heart. tnnt2a mutations were induced in zebrafish via the CRISPR/Cas9 technique, and the mutants displayed heart arrest and dilated cardiomyopathy-like phenotypes. We first utilized the classic myocardial promoter of the myl7 and TetOn inducible system to restore tnnt2a expression in myocardial tissue in tnnt2a mutant zebrafish. However, this attempt failed to recover normal heart function and circulation, although heart pumping was partially restored. Further analyses via both RNA-seq and immunofluorescence indicated that Tnnt2a, which was also expressed in a novel group of myl7-negative smooth muscle cells on the outflow tract (OFT), was indispensably responsible for the normal mechanical dynamics of OFT. Lastly, tnnt2 expression induced by OFT cells in addition to the myocardial cells successfully rescued heart function and circulation in tnnt2a mutant zebrafish. Together, our results reveal the significance of OFT expression of Tnnt2 for cardiac function and demonstrate zebrafish larva as a powerful and convenient in vivo platform for studying cardiomyopathy and the relevant therapeutic strategies.
