Evaluation of the NK2 homeobox 1 gene (NKX2-1) as a Hirschsprung's disease locus

Ann Hum Genet. 2008 Mar;72(Pt 2):170-7. doi: 10.1111/j.1469-1809.2007.00403.x. Epub 2007 Dec 12.


Hirschsprung's disease (HSCR, colonic aganglionosis) is an oligogenic entity that usually requires mutations in RET and other interacting loci. Decreased levels of RET expression may lead to the manifestation of HSCR. We previously showed that RET transcription was decreased due to alteration of the NKX2-1 binding site by two HSCR-associated RET promoter single nucleotide polymorphisms (SNPs). This prompted us to investigate whether DNA alterations in NKX2-1 could play a role in HSCR by affecting the RET-regulatory properties of the NKX2-1 protein. Our initial study on 86 Chinese HSCR patients revealed a Gly322Ser amino acid substitution in the NKX2-1 protein. In this study, we have examined 102 additional Chinese and 70 Caucasian patients and 194 Chinese and 60 Caucasian unselected, unrelated, subjects as controls. The relevance of the DNA changes detected in NKX2-1 by direct sequencing were evaluated using bioinformatics, reporter and binding-assays, mouse neurosphere culture, immunohistochemistry and immunofluorescence techniques. Met3Leu and Pro48Pro were identified in 2 Caucasian and 1 Chinese patients respectively. In vitro analysis showed that Met3Leu reduced the activity of the RET promoter by 100% in the presence of the wild-type or HSCR-associated RET promoter SNP alleles. The apparent binding affinity of the NKX2-1 mutated protein was not decreased. The Met3Leu mutation may affect the interaction of NKX2-1 with its protein partners. The absence of NKX2-1 expression in mouse but not in human gut suggests that the role of NKX2-1 in gut development differs between the two species. NKX2-1 mutations could contribute to HSCR by affecting RET expression through defective interactions with other transcription factors.

Publication types

  • Comparative Study
  • Research Support, Non-U.S. Gov't
  • Retracted Publication

MeSH terms

  • Animals
  • Asian Continental Ancestry Group / genetics
  • Australia
  • Base Sequence
  • Cell Line, Tumor
  • China
  • Computational Biology
  • Digestive System / embryology
  • Digestive System / metabolism
  • Electrophoretic Mobility Shift Assay
  • European Continental Ancestry Group / genetics
  • Fluorescent Antibody Technique
  • Gene Components
  • Genetic Predisposition to Disease / genetics*
  • Genotype
  • Hirschsprung Disease / genetics*
  • Humans
  • Immunohistochemistry
  • Mice
  • Molecular Sequence Data
  • Mutation / genetics
  • Nuclear Proteins / genetics*
  • Nuclear Proteins / metabolism
  • Proto-Oncogene Proteins c-ret / genetics
  • Proto-Oncogene Proteins c-ret / metabolism*
  • Reverse Transcriptase Polymerase Chain Reaction
  • Sequence Analysis, DNA
  • Thyroid Nuclear Factor 1
  • Transcription Factors / genetics*
  • Transcription Factors / metabolism


  • Nkx2-1 protein, mouse
  • Nuclear Proteins
  • Thyroid Nuclear Factor 1
  • Transcription Factors
  • Proto-Oncogene Proteins c-ret
  • RET protein, human