title: Transcriptional defect of an inherited NKX2-5 haplotype comprising a SNP, a nonsynonymous and a synonymous mutation, associated with human congenital heart disease.
abstract: Germline mutations in cardiac-specific transcription factor genes have been associated with congenital heart disease (CHD) and the homeodomain transcription factor NKX2-5 is an important member of this group. Indeed, more than 40 heterozygous NKX2-5 germline mutations have been observed in individuals with CHD, and these are spread along the coding region, with many shown to impact protein function. In pursuit of understanding causes of CHD, we analyzed n = 49 cardiac biopsies from 28 patients and identified by direct sequencing two nonsynonymous NKX2-5 alterations affecting alanine 119, namely c.356C>A (p.A119E) and c.355G>T, (p.A119S), in patients with AVSD and HLHS, respectively. In functional assays, a significant reduction in transcriptional activities could be determined for the NKX2-5 variants. Importantly, in one family the mother, besides p.A119E, carried a synonymous mutant allele in the homeodomain (c.543G>A, p.Q181), and a synonymous dbSNP (c.63A>G, p.E21) in the transactivation domain of the protein, that were transmitted to the CHD daughter. The presence of these variants in-cis with the p.A119E mutation led to a further reduction in transcriptional activities. Such difference in activity may be in part related to reduced protein expression for the double variant c.356C>A and c.543G>A. We propose changes in mRNA stability and folding, due to a silent mutation and a dbSNP in the NKX2-5 coding region to contribute to the functional defect. Although the clinical significance of the NKX2-5 haplotype identified in the CHD patients remains to be ascertained, we provide evidence of an interaction of a dbSNP, with synonymous and nonsynonymous mutations to negatively impact NKX2-5 transcriptional activity.
authors.affiliation: Fraunhofer Institute of Toxicology and Experimental Medicine, Hannover, Germany.
authors.forename: Stella Marie
authors.initials: SM
authors.lastname: Reamon-Buettner
authors.affiliation: Institute of Natural Sciences, Massey University, Auckland, New Zealand.
authors.forename: Evelyn
authors.initials: E
authors.lastname: Sattlegger
authors.affiliation: Laboratory of Transcriptional Networks, Centre for Integrative Biology, University of Trento, Mattarello, Italy.
authors.forename: Yari
authors.initials: Y
authors.lastname: Ciribilli
authors.affiliation: Laboratory of Transcriptional Networks, Centre for Integrative Biology, University of Trento, Mattarello, Italy.
authors.forename: Alberto
authors.initials: A
authors.lastname: Inga
authors.affiliation: Department of Pediatric Cardiology and Intensive Care Medicine, Hannover Medical School, Hannover, Germany.
authors.forename: Armin
authors.initials: A
authors.lastname: Wessel
authors.affiliation: Centre for Pharmacology and Toxicology, Hannover Medical School, Hannover, Germany.
authors.forename: Jürgen
authors.initials: J
authors.lastname: Borlak
mesh_terms: D000483:Alleles; D019943:Amino Acid Substitution; D005786:Gene Expression Regulation; D018095:Germ-Line Mutation; D006239:Haplotypes; D006330:Heart Defects, Congenital / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D006579:Heterozygote; D000072576:Homeobox Protein Nkx-2.5; D018398:Homeodomain Proteins / Q000235:genetics* / Q000378:metabolism; D006801:Humans; D009690:Nucleic Acid Conformation; D020641:Polymorphism, Single Nucleotide; D017434:Protein Structure, Tertiary; D020871:RNA Stability; D012333:RNA, Messenger / Q000235:genetics* / Q000378:metabolism; D014157:Transcription Factors / Q000235:genetics* / Q000378:metabolism; D014158:Transcription, Genetic
chemical_list: D000072576:Homeobox Protein Nkx-2.5; D018398:Homeodomain Proteins; C095552:NKX2-5 protein, human; D012333:RNA, Messenger; D014157:Transcription Factors