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MESP2 variants contribute to conotruncal heart defects by inhibiting cardiac neural crest cell proliferation

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Abstract

Conotruncal heart defects (CTDs) are closely related to defective outflow tract (OFT) development, in which cardiac neural crest cells (CNCCs) play an indispensable role. However, the genetic etiology of CTDs remains unclear. Mesoderm posterior 2 (MESP2) is an important transcription factor regulating early cardiogenesis. Nevertheless, MESP2 variants have not been reported in congenital heart defect (CHD) patients. We first identified four MESP2 variants in 601 sporadic nonsyndromic CTD patients that were not detected in 400 healthy controls using targeted sequencing. Reverse transcription-quantitative PCR (RT-qPCR), immunohistochemistry, and immunofluorescence assays revealed MESP2 expression in the OFT of Carnegie stage (CS) 11, CS13, and CS15 human embryos and embryonic day (E) 8.5, E10, and E11.5 mouse embryos. Functional analyses in HEK 293T cells, HL-1 cells, JoMa1 cells, and primary mouse CNCCs revealed that MESP2 directly regulates the transcriptional activities of downstream CTD-related genes and promotes CNCC proliferation by regulating cell cycle factors. Three MESP2 variants, c.346G>C (p.G116R), c.921C>G (p.Y307X), and c.59A>T (p.Q20L), altered the transcriptional activities of MYOCD, GATA4, NKX2.5, and CFC1 and inhibited CNCC proliferation by upregulating p21cip1 or downregulating Cdk4. Based on our findings, MESP2 variants disrupted MESP2 function by interfering with CNCC proliferation during OFT development, which may contribute to CTDs.

Key messages

  • This study first analyzed MESP2 variants identified in sporadic nonsyndromic CTD patients.

  • MESP2 is expressed in the OFT of different stages of human and mouse embryos.

  • MESP2 regulates the transcriptional activities of downstream CTD-related genes and promotes CNCC proliferation by regulating cell cycle factor p21cip1 or Cdk4.

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Acknowledgments

We thank all the patients and control individuals who participated in the study. We thank Dr. Shubao Chen for providing helpful advice during the preparation of the manuscript. We thank the members of the Medical Laboratory and Department of Pediatric Cardiology, Shanghai Children’s Medical Center, for their indispensable assistance with peripheral blood collection. We thank the members of the Lab of Pediatric Cardiovascular and Department of Pediatric Cardiovascular, Xinhua Hospital, for providing technical assistance. We also thank the members of the Department of Obstetrics and Gynecology, Xinhua Hospital, for their collaborative assistance with human embryo collection.

Funding

This work was supported by grants from the National Natural Science Foundation of China (grant number 81670285, 81974021, and 81974012), the Major International (Regional) Joint Research Project of the National Natural Science Foundation of China (grant number 81720108003), the Shanghai Hospital Development Center (grant number SHDC12015102), and the Shanghai Municipal Health Bureau (grant number ZHYY-ZXYJHZX-1-04).

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Authors and Affiliations

  1. Department of Pediatric Cardiovascular, Xinhua Hospital, School of Medicine, Shanghai Jiaotong University, 1665 Kongjiang Road, Shanghai, 200092, China

    Erge Zhang, Jianping Yang, Yang Liu, Nanchao Hong, Huilin Xie, Sun Chen, Yu Yu & Kun Sun

  2. Medical Laboratory, Shanghai Children’s Medical Center, School of Medicine, Shanghai Jiaotong University, Shanghai, China

    Qihua Fu

  3. Department of Pediatric Cardiology, Shanghai Children’s Medical Center, School of Medicine, Shanghai Jiaotong University, Shanghai, China

    Fen Li

  4. Institute for Developmental and Regenerative Cardiovascular Medicine, Xinhua Hospital, School of Medicine, Shanghai Jiaotong University, Shanghai, China

    Yu Yu

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  1. Erge Zhang
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Correspondence to Yu Yu or Kun Sun.

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The human study was approved by the Ethics Committee of Xinhua Hospital (XHEC-2016-C-2016-303) and Shanghai Children’s Medical Center (SCMC-201015) and performed in accordance with the Declaration of Helsinki. Peripheral blood and embryos were collected from the participants after informed consent was obtained. The animal experiments were approved by the Ethics Committee of Xinhua Hospital (XHEC-2016-F-2016-197). All animal procedures conformed to the NIH Guidelines for the Care and Use of Laboratory Animals.

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Zhang, E., Yang, J., Liu, Y. et al. MESP2 variants contribute to conotruncal heart defects by inhibiting cardiac neural crest cell proliferation. J Mol Med 98, 1035–1048 (2020). https://doi.org/10.1007/s00109-020-01929-4

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