Novel Point Mutations of CITED2 Gene Are Associated with Non-familial Congenital Heart Disease (CHD) in Sporadic Pediatric Patients

  • Sima Dianatpour
  • Mehri KhatamiEmail author
  • Mohammad Mehdi Heidari
  • Mehdi Hadadzadeh


CITED2 is a cardiac transcription factor that plays a critical role in cardiac development. Gene mutations in CITED2 lead to a series of cardiac malformations and congenital heart defects (CHD). Congenital heart disease generally refers to defects in the heart’s structure or function and often seen in many forms such as ventricular septal defects (VSDs), atrial septal defects (ASDs), and tetralogy of Fallot (TOF). However, the mechanisms involved in these mutations are poorly understood. The aim of the present study was to evaluate the mutations of the CITED2 gene in pediatric patients with congenital heart defects. We studied the potential impact of sequence variations of the CITED2 gene in a cohort of 150 patients with non-familial CHD and 98 control individuals by polymerase chain reaction-single-stranded conformation polymorphism (PCR-SSCP) and subsequently direct sequencing. We identified seven novel CITED2 nucleotide changes. Four of these alterations were found in the coding region (c.716insG, c.389A>G, c.450G>C and c.512-538del27) and were only seen in our patients, and not detected in the control group. These mutations are leading to changes in the amino acid sequence in the position of p.Gly236fs, p.Asn125Ser, p.Gln145His, and p.Ser170-Gly178del, respectively. Other variations are located in the 5′UTR region of the gene (c.-43C>T, c.-64C>T and c.-90A>G). CITED2 gene mutations in control subjects were not observed. Our Bioinformatics assay results showed that these novel mutations alter the RNA folding, protein structure, and, therefore, probable effect on the protein function and may play a significant role in the development of congenital heart diseases.


Congenital heart disease CITED2 Mutation PCR-SSCP 



We thank all the patients for providing blood samples for scientific research, including the Afshar cardiac hospital patients (Yazd, Iran). The study was approved by the Yazd University Human Research Ethics Committee.

Author Contributions

M.K. and M.M.H. conceived the study, participated in its design, and drafted the initial manuscript. M.M. and S.D. collected the data. M.K., M.M.H., Y.A.A, and S.D. performed statistics and interpretation of data and drafted the last version of the manuscript. M.K., M.M.H., and M.H. coordinated data interpretation, participated in the design of the study, and helped to draft the manuscript. All authors read and approved the final manuscript.

Compliance with Ethical Standards

Conflict of Interest

The authors declare that they have no conflicts of interest.


  1. 1.
    Hoffman, J. I., & Kaplan, S. (2002). The incidence of congenital heart disease. Journal of the American College of Cardiology, 39(12), 1890–1900.CrossRefGoogle Scholar
  2. 2.
    Blue, G. M., Kirk, E. P., Sholler, G. F., Harvey, R. P., & Winlaw, D. S. (2012). Congenital heart disease: current knowledge about causes and inheritance. The Medical Journal of Australia, 197(3), 155–159.CrossRefGoogle Scholar
  3. 3.
    Russell, M. W., Chung, W. K., Kaltman, J. R., & Miller, T. A. (2018). Advances in the understanding of the genetic determinants of congenital heart disease and their impact on clinical outcomes. Journal of the American Heart Association, 7(6).Google Scholar
  4. 4.
    Bentham, J., & Bhattacharya, S. (2008). Genetic mechanisms controlling cardiovascular development. Annals of the New York Academy of Sciences, 1123(1), 10–19.CrossRefGoogle Scholar
  5. 5.
    Ferencz, C., & Boughman, J. A. (1993). Congenital heart disease in adolescents and adults. Teratology, genetics, and recurrence risks. Cardiology Clinics, 11(4), 557–567.CrossRefGoogle Scholar
  6. 6.
    Clark, K. L., Yutzey, K. E., & Benson, D. W. (2006). Transcription factors and congenital heart defects. Annual Review of Physiology, 68(1), 97–121.CrossRefGoogle Scholar
  7. 7.
    Li, C., Peng, Y., Zhou, B., Bai, W., & Rao, L. (2017). Association of LIM domain 7 Gene polymorphisms and plasma levels of LIM domain 7 with Dilated cardiomyopathy in a chinese population. Applied Biochemistry and Biotechnology, 182(3), 885–897.CrossRefGoogle Scholar
  8. 8.
    Wu, H., Qiao, M., Peng, X., Wu, J., Liu, G., Sun, H., Li, L., & Mei, S. (2013). Molecular characterization, expression patterns, and association analysis with carcass traits of porcine USF1 gene. Applied Biochemistry and Biotechnology, 170(6), 1310–1319.CrossRefGoogle Scholar
  9. 9.
    Khatami, M., Mazidi, M., Taher, S., Heidari, M. M., & Hadadzadeh, M. (2018). Novel point mutations in the NKX2.5 gene in pediatric patients with non-familial congenital heart disease. Medicina (Kaunas, Lithuania), 54.Google Scholar
  10. 10.
    Schott, J. J., Benson, D. W., Basson, C. T., Pease, W., Silberbach, G. M., Moak, J. P., et al. (1998). Congenital heart disease caused by mutations in the transcription factor NKX2-5. Science, 281(5373), 108–111.CrossRefGoogle Scholar
  11. 11.
    Khatami, M., Heidari, M. M., Kazeminasab, F., & Zare Bidaki, R. (2018). Identification of a novel non-sense mutation in TBX5 gene in pediatric patients with congenital heart defects. Journal of Cardiovascular and Thoracic Research, 10(1), 41–45.CrossRefGoogle Scholar
  12. 12.
    Kodo, K., Nishizawa, T., Furutani, M., Arai, S., Ishihara, K., Oda, M., Makino, S., Fukuda, K., Takahashi, T., Matsuoka, R., Nakanishi, T., & Yamagishi, H. (2012). Genetic analysis of essential cardiac transcription factors in 256 patients with non-syndromic congenital heart defects. Circulation Journal, 76(7), 1703–1711.CrossRefGoogle Scholar
  13. 13.
    Xiong, F., Li, Q., Zhang, C., Chen, Y., Li, P., Wei, X., Li, Q., Zhou, W., Li, L., Shang, X., & Xu, X. (2013). Analyses of GATA4, NKX2.5, and TFAP2B genes in subjects from southern China with sporadic congenital heart disease. Cardiovascular Pathology, 22(2), 141–145.CrossRefGoogle Scholar
  14. 14.
    Liu, X. Y., Wang, J., Yang, Y. Q., Zhang, Y. Y., Chen, X. Z., Zhang, W., Wang, X. Z., Zheng, J. H., & Chen, Y. H. (2011). Novel NKX2-5 mutations in patients with familial atrial septal defects. Pediatric Cardiology, 32(2), 193–201.CrossRefGoogle Scholar
  15. 15.
    Yin, Z., Haynie, J., Yang, X., Han, B., Kiatchoosakun, S., Restivo, J., Yuan, S., Prabhakar, N. R., Herrup, K., Conlon, R. A., Hoit, B. D., Watanabe, M., & Yang, Y. C. (2002). The essential role of Cited2, a negative regulator for HIF-1alpha, in heart development and neurulation. Proceedings of the National Academy of Sciences of the United States of America, 99(16), 10488–10493.CrossRefGoogle Scholar
  16. 16.
    Liu, Y., Wang, F., Wu, Y., Tan, S., Wen, Q., Wang, J., Zhu, X., Wang, X., Li, C., Ma, X., & Pan, H. (2014). Variations of CITED2 are associated with congenital heart disease (CHD) in Chinese population. PLoS One, 9(5), e98157.CrossRefGoogle Scholar
  17. 17.
    Li, Q., Pan, H., Guan, L., Su, D., & Ma, X. (2012). CITED2 mutation links congenital heart defects to dysregulation of the cardiac gene VEGF and PITX2C expression. Biochemical and Biophysical Research Communications, 423(4), 895–899.CrossRefGoogle Scholar
  18. 18.
    Xu, M., Wu, X., Li, Y., Yang, X., Hu, J., Zheng, M., & Tian, J. (2014). CITED2 mutation and methylation in children with congenital heart disease. Journal of Biomedical Science, 21(1), 7.CrossRefGoogle Scholar
  19. 19.
    Freedman, S. J., Sun, Z. Y., Kung, A. L., France, D. S., Wagner, G., & Eck, M. J. (2003). Structural basis for negative regulation of hypoxia-inducible factor-1alpha by CITED2. Nature Structural Biology, 10(7), 504–512.CrossRefGoogle Scholar
  20. 20.
    Xu, B., Doughman, Y., Turakhia, M., Jiang, W., Landsettle, C. E., Agani, F. H., Semenza, G. L., Watanabe, M., & Yang, Y. C. (2007). Partial rescue of defects in Cited2-deficient embryos by HIF-1alpha heterozygosity. Developmental Biology, 301(1), 130–140.CrossRefGoogle Scholar
  21. 21.
    Bamforth, S. D., Braganca, J., Eloranta, J. J., Murdoch, J. N., Marques, F. I., Kranc, K. R., et al. (2001). Cardiac malformations, adrenal agenesis, neural crest defects and exencephaly in mice lacking Cited2, a new Tfap2 co-activator. Nature Genetics, 29(4), 469–474.CrossRefGoogle Scholar
  22. 22.
    Sperling, S., Grimm, C. H., Dunkel, I., Mebus, S., Sperling, H. P., Ebner, A., Galli, R., Lehrach, H., Fusch, C., Berger, F., & Hammer, S. (2005). Identification and functional analysis of CITED2 mutations in patients with congenital heart defects. Human Mutation, 26(6), 575–582.CrossRefGoogle Scholar
  23. 23.
    Yang, X. F., Wu, X. Y., Li, M., Li, Y. G., Dai, J. T., Bai, Y. H., et al. (2010). Mutation analysis of Cited2 in patients with congenital heart disease. Zhonghua Er Ke Za Zhi, 48, 293–296.Google Scholar
  24. 24.
    Chen, C. M., Bentham, J., Cosgrove, C., Braganca, J., Cuenda, A., Bamforth, S. D., Schneider, J. E., Watkins, H., Keavney, B., Davies, B., & Bhattacharya, S. (2012). Functional significance of SRJ domain mutations in CITED2. PLoS One, 7(10), e46256.CrossRefGoogle Scholar
  25. 25.
    Lopes Floro, K., Artap, S. T., Preis, J. I., Fatkin, D., Chapman, G., Furtado, M. B., Harvey, R. P., Hamada, H., Sparrow, D. B., & Dunwoodie, S. L. (2011). Loss of Cited2 causes congenital heart disease by perturbing left-right patterning of the body axis. Human Molecular Genetics, 20(6), 1097–1110.CrossRefGoogle Scholar
  26. 26.
    Bamforth, S. D., Braganca, J., Farthing, C. R., Schneider, J. E., Broadbent, C., Michell, A. C., et al. (2004). Cited2 controls left-right patterning and heart development through a Nodal-Pitx2c pathway. Nature Genetics, 36(11), 1189–1196.CrossRefGoogle Scholar
  27. 27.
    Weninger, W. J., Lopes Floro, K., Bennett, M. B., Withington, S. L., Preis, J. I., Barbera, J. P., et al. (2005). Cited2 is required both for heart morphogenesis and establishment of the left-right axis in mouse development. Development, 132(6), 1337–1348.CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2019

Authors and Affiliations

  1. 1.Department of Biology, Faculty of ScienceYazd UniversityYazdIran
  2. 2.Department of Cardiac Surgery, Afshar HospitalShahid Sadoughi University of Medical SciencesYazdIran

Personalised recommendations