Functional Characterization of a Novel Mutation in NKX2-5 Associated With Congenital Heart Disease and Adult-Onset Cardiomyopathy

Abstract

Background The transcription factor NKX2-5 is crucial for heart development and mutations in this gene have been implicated in diverse congenital heart diseases (CHD) and conduction defects (CD) in mouse models and humans. Whether NKX2-5 mutations have a role in adult-onset heart disease is unknown. Methods and Results Mutation screening was performed in 220 probands with adult-onset dilated cardiomypathy (DCM). Six NKX2-5 coding sequence variants were identified, including 3 non-synonymous variants. A novel heterozygous mutation, I184M, located within the NKX2-5 homeodomain (HD), was identified in one family. A subset of family members had CHD, but there was an unexpectedly high prevalence of DCM. Functional analysis of I184M in vitro demonstrated a striking increase in protein expression when transfected into COS-7 cells or HL-1 cardiomyocytes, due to reduced degradation by the ubiquitin-proteasome system (UPS). In functional assays, DNA binding activity of I184M was reduced, resulting in impaired activation of target genes, despite increased expression levels of mutant protein. Conclusions Certain NKX2-5 HD mutations show abnormal protein degradation via the UPS and partially impaired transcriptional activity. We propose that this class of mutation can impair heart development and mature heart function, and contribute to NKX2-5-related cardiomyopathies with graded severity.

A systematic variant screening in familial cases of congenital heart defects demonstrates the usefulness of molecular genetics in this field

“…dilated cardiomyopathy as already observed in two previous reports. 14,15 Family MC177 is interesting because the maternal grandmother had a D-TGV and carried the heterozygous ZIC3 diseasecausing variant, suggesting that this variant could be dominant or that she had an unbalanced X inactivation as already shown in two other familial cases. 16,17 It is much easier in familial cases to infer the putative causal gene than in sporadic cases, because in familial cases the spectrum of phenotypic anomalies is broader by compiling the various anomalies of affected relatives.…”

Prevalence and Spectrum of NKX2-5 Mutations Associated With Sporadic Adult-Onset Dilated Cardiomyopathy

“…38) Costa and colleagues 32) screened NKX2-5 in 220 probands with adult-onset DCM, and found a novel missense mutation (p.I184M) in a proband, as well as two previously reported functional polymorphisms (p.R25C and p. A119S) in 2 other probands, with a mutational prevalence of about 1.36%. Genetic analysis of the mutation carrier's family showed that the mutation was present in all affected living adult family members.…”

“…In humans, NKX2-5 loss-of-function mutations have been previously implicated in familial adult-onset DCM, ASD, and arrhythmias, 32,33) of which the most commonly reported phenotypes are ASD and AVB in 68.4% and 65.7% of the cases, respectively. 38) Costa and colleagues 32) screened NKX2-5 in 220 probands with adult-onset DCM, and found a novel missense mutation (p.I184M) in a proband, as well as two previously reported functional polymorphisms (p.R25C and p. A119S) in 2 other probands, with a mutational prevalence of about 1.36%.…”

“…34) Individuals with idiopathic DCM in the presence of congenital ASD were also included because NKX2-5 mutations have been involved in familial DCM and ASD. 32) Individuals with a positive family history of DCM were excluded from the present study. The study was carried out in accordance with the principles of the Declaration of Helsinki.…”

“…[22][23][24][25][26][27][28][29][30][31] More interestingly, mutations in NKX2-5, another cardiac key transcriptional factor, have been found to be responsible for familial adult-onset DCM, 32,33) which makes it justifiable to evaluate the prevalence and spectrum of NKX2-5 mutations in the patients with sporadic adult-onset DCM.…”