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Tetralogy of Fallot: variants of MYH6 gene promoter and cellular functional analyses

Pediatric Research (2023)Cite this article

Abstract

Background

Tetralogy of Fallot (TOF) is a common form of congenital heart disease. The MYH6 gene has important effects on cardiovascular growth and development.

Methods

In 608 subjects, including 315 TOF patients, we investigated the MYH6 gene promoter variants and verified the effect on gene expression by using cellular functional experiments with three cell lines (HEK-293, HL-1, and H9C2 cells) and bioinformatics analysis.

Results

In the MYH6 gene promoter, 12 variants were identified from 608 subjects. Five variants were found only in patients with TOF and two of them (g.3384G>T and g.4518T>C) were novel. Electrophoretic mobility shift assay with three cell lines (HEK-293, HL-1, and H9C2) showed significant changes in the transcription factors bound by the promoter variants compared to the wild-type. Dual luciferase reporter showed that four of the five variants reduced the transcriptional activity of the MYH6 gene promoter (pā€‰<ā€‰0.05).

Conclusions

This study is the first to test the cellular function of variants in the promoter region of the MYH6 gene in patients with TOF, which provides new insights into the genetic basis of TOF and provides a basis for further study of the mechanism of TOF formation.

Impact

  • DNA from 608 human subjects was sequenced for MYH6 gene promoter region variants with five variants found only in TOF patients and two were novel.

  • EMSA and dual luciferase reporter experiments in three cell lines found these variants pathological.

  • Prediction by JASPAR database indicated that these variants alter the transcription factor binding sites.

  • The study, for the first time, confirmed that there are variants at the MYH6 gene promoter region and these variants alter the cellular function.

  • The variants found in this study suggest the possible pathological role in the formation of TOF.

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Fig. 1: The experiment flowchart.
Fig. 2: Locations and sequencing chromatograms of MYH6 gene promoter variants.
Fig. 3: Results of dual luciferase reporter gene assay and echocardiography of the patients with the variants.
Fig. 4: Schema describing the role of MYH6 gene promoter variants.

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Data availability

The datasets generated during and analyzed during the current study are available from https://ngdc.cncb.ac.cn/omix/view/OMIX001938 and the corresponding author upon request.

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Acknowledgements

We thank the patients and their family members for cooperation. The assistance of surgeons and Nursing staff at the Division of Pediatric Cardiac Surgery, Department of Cardiovascular Surgery is gratefully acknowledged.

Funding

This work was funded by the National Natural Science Foundation of China [82170353]; Tianjin Science and Technology Commission [22ZYQYSY00020];the Non-profit Central Research Institute Fund of Chinese Academy of Medical Sciences [2020-PT310-007]; TEDA International Cardiovascular Hospital [2021-ZX-002]; and Tianjin Key Medical Discipline (Specialty) Construction Project (TJYXZDXK-019A).

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Authors and Affiliations

  1. The Institute of Cardiovascular Diseases & Department of Cardiovascular Surgery, TEDA International Cardiovascular Hospital, Tianjin University & Chinese Academy of Medical Sciences, Tianjin, 300457, China

    Ji-Yang Zuo,Ā Huan-Xin Chen,Ā Qin Yang,Ā Zhi-Gang LiuĀ &Ā Guo-Wei He

  2. Tianjin Key Laboratory of Molecular Regulation of Cardiovascular Diseases and Translational Medicine, Tianjin, 300457, China

    Ji-Yang Zuo,Ā Huan-Xin Chen,Ā Qin Yang,Ā Zhi-Gang LiuĀ &Ā Guo-Wei He

  3. Clinical School of Cardiovascular Disease, Tianjin Medical University, Tianjin, China

    Ji-Yang Zuo

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Contributions

G.-W.H. obtained the funding for this project. J.-Y.Z. and H.-X.C. performed experiments. J.-Y.Z., H.-X.C., Z.-G.L. and G.-W.H. analyzed experimental data. J.-Y.Z., H.-X.C., Z.-G.L. and G.-W.H. collected the human blood samples. Q.Y. discussed the findings and participated in the project. J.-Y.Z. and G.-W.H. wrote the manuscript. G.-W.H. supervised the whole project. All authors read and approved the final manuscript.

Corresponding authors

Correspondence to Zhi-Gang Liu or Guo-Wei He.

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The authors declare no competing interests.

Ethics approval

This study involving human participants was reviewed and approved by the ethics committee of TEDA International Cardiovascular Hospital, China (No. 0715-4, 2021, August 2, 2021).

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Patients signed informed consent regarding publishing their data and photographs.

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Zuo, JY., Chen, HX., Yang, Q. et al. Tetralogy of Fallot: variants of MYH6 gene promoter and cellular functional analyses. Pediatr Res (2023). https://doi.org/10.1038/s41390-023-02955-x

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