Loss of function and inhibitory effects of human CSX/NKX2.5 homeoprotein mutations associated with congenital heart disease
Hideko Kasahara, … , Christine E. Seidman, Seigo Izumo
Hideko Kasahara, … , Christine E. Seidman, Seigo Izumo
Published January 15, 2000
Citation Information: J Clin Invest. 2000;106(2):299-308. https://doi.org/10.1172/JCI9860.
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Article

Loss of function and inhibitory effects of human CSX/NKX2.5 homeoprotein mutations associated with congenital heart disease

Abstract

CSX/NKX2.5 is an evolutionarily conserved homeodomain-containing (HD-containing) transcription factor that is essential for early cardiac development. Recently, ten different heterozygous CSX/NKX2.5 mutations were found in patients with congenital heart defects that are transmitted in an autosomal dominant fashion. To determine the consequence of these mutations, we analyzed nuclear localization, DNA binding, transcriptional activation, and dimerization of mutant CSX/NKX2.5 proteins. All mutant proteins were translated and located to the nucleus, except one splice-donor site mutant whose protein did not accumulate in the cell. All mutants that had truncation or missense mutations in the HD had severely reduced DNA binding activity and little or no transcriptional activation function. In contrast, mutants with intact HDs exhibit normal DNA binding to the monomeric binding site but had three- to ninefold reduction in DNA binding to the dimeric binding sites. HD missense mutations that preserved homodimerization ability inhibited the activation of atrial natriuretic factor by wild-type CSX/NKX2.5. Although our studies do not characterize the genotype-phenotype relationship of the ten human mutations, they identify specific abnormalities of CSX/NKX2.5 function essential for transactivation of target genes.

Authors

Hideko Kasahara, Bora Lee, Jean-Jacques Schott, D. Woodrow Benson, J.G. Seidman, Christine E. Seidman, Seigo Izumo

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Figure 1

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Diagram of CSX/NKX2.5 cDNA with the location of ten mutation sites ident...
Diagram of CSX/NKX2.5 cDNA with the location of ten mutation sites identified in congenital heart disease. Ten mutation sites (asterisks in Wild) were divided into five groups based on the predicted protein structure: nonsense mutation in the HD (group 1: M149 and M170); missense mutation in the HD (group 2: M178, M188, M189, and M191); premature termination after HD (group 3: M198 and M259); 25Arg-Cys missense mutation (group 4: M25); and mutation at the intron-splicing donor site (group 5: M112). Phenotypes observed in patients are listed on the left. For example, “11/12” indicates that 11 patients show the phenotype among 12 patients examined. These mutation sites were mapped on CSX/NKX2.5 cDNA, which encodes 324 amino acids including 60 amino acids of HD (shaded box). Nuclear localization signal at the NH2-terminus of the HD is indicated (black box). Predicted translated product of M112 mutation in splicing donor site is indicated with a light gray box. AV block, atrioventricular conduction block; ASD, atrial septal defect; VSD, ventricular septal defect; TOF, tetralogy of Fallot; TV, tricuspid valve abnormality; DORV, double outlet right ventricle; NLS, nuclear localization signal; HD, homeodomain.