A tyrosine-rich domain within homeodomain transcription factor Nkx2-5 is an essential element in the early cardiac transcriptional regulatory machinery

Development. 2006 Apr;133(7):1311-22. doi: 10.1242/dev.02305. Epub 2006 Mar 1.

Abstract

Homeodomain factor Nkx2-5 is a central component of the transcription factor network that guides cardiac development; in humans, mutations in NKX2.5 lead to congenital heart disease (CHD). We have genetically defined a novel conserved tyrosine-rich domain (YRD) within Nkx2-5 that has co-evolved with its homeodomain. Mutation of the YRD did not affect DNA binding and only slightly diminished transcriptional activity of Nkx2-5 in a context-specific manner in vitro. However, the YRD was absolutely essential for the function of Nkx2-5 in cardiogenesis during ES cell differentiation and in the developing embryo. Furthermore, heterozygous mutation of all nine tyrosines to alanine created an allele with a strong dominant-negative-like activity in vivo: ES cell<-->embryo chimaeras bearing the heterozygous mutation died before term with cardiac malformations similar to the more severe anomalies seen in NKX2.5 mutant families. These studies suggest a functional interdependence between the NK2 class homeodomain and YRD in cardiac development and evolution, and establish a new model for analysis of Nkx2-5 function in CHD.

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Animals, Newborn
  • Blotting, Western
  • Cell Line
  • Cells, Cultured
  • Cephalopoda
  • Conserved Sequence
  • Electrophoretic Mobility Shift Assay
  • Embryo, Mammalian
  • Embryo, Nonmammalian
  • Gene Expression Regulation, Developmental
  • Gene Targeting
  • Genes, Reporter
  • Glutathione Transferase / metabolism
  • Green Fluorescent Proteins / metabolism
  • Heterozygote
  • Homeobox Protein Nkx-2.5
  • Homeodomain Proteins / chemistry
  • Homeodomain Proteins / genetics*
  • Homeodomain Proteins / metabolism*
  • In Situ Hybridization
  • Luciferases / metabolism
  • Mice
  • Molecular Sequence Data
  • Mutation
  • Myocardium / cytology
  • Myocardium / metabolism*
  • Myocardium / pathology
  • Myocytes, Cardiac / cytology
  • Myocytes, Cardiac / metabolism
  • Phylogeny
  • Protein Structure, Tertiary
  • Recombinant Fusion Proteins / metabolism
  • Reverse Transcriptase Polymerase Chain Reaction
  • Sequence Homology, Amino Acid
  • Transcription Factors / chemistry
  • Transcription Factors / genetics*
  • Transcription Factors / metabolism*
  • Transcription, Genetic*
  • Transcriptional Activation
  • Tyrosine / chemistry*

Substances

  • Homeobox Protein Nkx-2.5
  • Homeodomain Proteins
  • Nkx2-5 protein, mouse
  • Recombinant Fusion Proteins
  • Transcription Factors
  • Green Fluorescent Proteins
  • Tyrosine
  • Luciferases
  • Glutathione Transferase