Introduction
Atrial septal defect (ASD) is a common cardiovascular malformation, accounting for 10% of congenital heart defects (CHD), which is one of the major birth defects in the world. ASD is often reported in sporadic form; however, the reported familial cases have more research value [
1‐
3]. ASD may be isolated or associated with other CHDs, such as pulmonary valve stenosis (PVS), ventricular septal defect (VSD) and conduction defects, one of the study found that GATA4 genetic variations are associated with ASD, TOF and VSD in South Indian patients. In silico analysis provides further evidence that some of the observed mutations are pathogenic [
4]. In addition, persistent left to right blood shunt may result in atrial dysfunctions and atrial arrhythmias, in the absence of surgical or catheter-based repair [
5]. Therefore, CHD is still a serious threat to human, so the early prenatal screening and diagnosis for this type of birth defect are urgently required.
Although the CHD etiology is too complicated to be well characterized because of the complexity of heart development, numerous intrinsic factors [
6] (genetic factors) and extrinsic threats [
7] (environmental) were identified as contribution to CHD. Many candidate genes such as
GATA4,
TBX5,
NKX2.5,
BMP4 and
HAND1 have been proven to be responsible for heart development and diseases [
8].
GATA4 is one of the most widely investigated genes in CHD, with over 100 known mutation sites, which are related to the structural heart defect such as ASD, VSD, and PVS [
9]. Recently, the phenotypic genetics of familial ASD has been widely investigated, and transcription factors as an important mediator in cardiac development are still the focus of attention [
10‐
12]. Intrinsic factors have been identified as a major contributor to the pathogenesis of family ASD with the development of sequencing technology [
13], and mutations in the
GATA4 gene have been identified as a pathogenic factor of familial ASD [
3,
14].
The zinc-finger transcription factors were encoded by GATA binding protein 4 (
GATA4), which is essential for heart development [
15,
16] and considered to be a gene regulating embryogenesis and myocardial differentiation and function, and bound the GATA motif which is present in the promoters of many genes [
17].
GATA4 has 442 amino acids, including the N-terminus zinc fingers (NZf), the C-terminus zinc fingers (CZf) and the nuclear localization signal (NLS) [
2]. More importantly, the 271–322 amino acid fragment in the DNA binding domain has been reported and proved to be the smallest functional NLS region, which is vital to the process of cardiac development [
18]. In the current study, we checked out a clinically characterized family with a diagnosis of ASD. We found an obvious autosomal-dominant inheritance with reduced penetrance in this family. In addition, after performing surgical confirming and surgical repairs on patients, we conducted a clinical and genetic analysis and identified for the first time a novel pathogenic mutation of
GATA4 in the NLS region (NM_002052: exon5: c.T929C: p.M310T) by whole-exome sequencing of the patient in the family, which was confirmed by Sanger sequencing. Taken together, our study strongly suggests that the dominant family ASD involved in this study may be caused by
GATA4 gene deficiency.
Discussion
ASD is the third most common type of congenital heart disease, of which about 65–70% are secundum defects [
25]. In these patients, changes in cardiac structure are triggered by increased blood flow resulting from a left-to-right shunt due to intracardiac defect. Therefore, early detection and surgical treatment are the main strategies. Thanks to the development of technique, including the introduction of cardiopulmonary bypass and accompanying large-scale heart surgery skills improvement, the mortality of ASD has been dramatically reduced [
26]. However, due to the complexity of heart development, the exact causes of ASD, especially for the complex overlapping phenotype of ASD, still need more ongoing research efforts though we know that knowledge of heart development and function is the absolute necessity for better survival of human. Nowadays, the discovery of genetic causes of ASD has been being accelerated by many new technologies including single nucleotide polymorphism arrays, next-generation sequencing (NGS), and copy number variant platforms [
27]. Among all new genetic technologies, the application of NGS in various aspects of heart biology has resulted in discoveries, generating novel insights into this field of study [
28]. In the present study, WES combined ROH was employed to find out the suspicious disease-causing gene in one consanguineous family. This technique has been considered as a rapid and cost-effective tool for screening the new variants or genes for rare Mendelian unknown disorders [
29,
30]. It helps genetic diagnostics for clinical cases with a mutational spectrum of known and unknown diseases. Some filtering strategies are needed for excluding variants that are implausible to cause disease because sometimes it is difficult to identify between pathogenic and benign mutations in the WES results [
31]. With the analysis of WES result in this study, we quickly determined the most possible pathogenic mutation in this family is
GATA4 p.M310T and confirmed the result by Sanger sequencing.
GATA4 gene belongs to a GATA family, which is consisted of 6 structure-conserved transcription factors.
GATA4 gene, which is expressed in the cardiac system and endodermal derivatives [
32], is a highly conserved transcriptional factor with seven exons. The
GATA4 protein is comprised of TAD, NZF and NLS [
2,
33]. Many shreds of evidence showed that
GATA4 plays significant roles in many stages of heart development, including looping morphogenesis, septation, ventricular myocardium proliferation, and heart contraction [
34]. For this reason,
GATA4 was considered as a regular candidate for CHD genetic screening. Many mutations in the
GATA4 coding region have been identified as the genotype of CHD patients and not all of them were predicted by bioinformatics tools, such as PROVEAN and SIFT, as the pathogenic genes [
34]. NLS was considered as a crucial role in ASD epidemiology because 5 family cases were reported in this region, include S52F, G296S, 1074 (delC), 1075 (delG) and M310V [
2,
3,
35]. Especially in our present study, we determined a different protein mutation in the same protein location (M310V) as reported [
2]. What’s more, in the mouse mutation study, M310V transgenic mice had shown a higher incidence of CHD than wild-type control mice [
36], which indicated codon 310 in the
GATA4 gene is a CHD-related pathogenic coding region.
Here, we report for the first time an M310T mutation in the NLS region, which is necessary and sufficient for
GATA4 transcription factor activity and cardiac development. Using the Swiss model tool, we identified the NLS region as the potential impact region of the mutation on protein structure (Fig.
4). The changes in this region may lead to a decrease of transcriptional activity, thus affecting the transcriptional activation process during development. What's more, the region affected by M310T mutation is also immediately adjacent to CZf region, which is crucial for DNA binding and cofactor interaction [
36,
37]. In addition, Garg reported [
14] that the G296S mutation disrupts the DNA-binding and transactivation activity of
GATA4 and destroys the synergy in transcriptional activation between
GATA4 and its cofactor
TBX5, resulting in heart anomalies such as pulmonary stenosis, atrioventricular septal defect, and ASD. Therefore,
GATA4 mutation in NLS region may also affect the expression of other transcription factors (
TBX5 or
NKX2.5) [
14,
35], which are crucial in the development of heart, resulting in the observation of ASD in three affected members of this family.
ASD families mostly present the same subtype structural defects without arrhythmia. In this study, all affected family members presented similar defects with distinct differences and all 3 patients were detected different tachyarrhythmia. Although some reported arrhythmias and conduction disorders may be associated with atrial septal defects [
38], the reason is unknown and some transcriptional factors genes mutations were related to the ASD family with arrhythmias [
39], one possible evidence shown by computationally Mattapally et al. [
40], where they established that
NKX2.5 cooperativity with
GATA4 facilitates its activating and repressing functions [
41‐
43]. The interaction between
NKX2.5 (TN domain) and
GATA4 might also be important for the function as a repressor of ion channels and its downstream target genes. Therefore, they speculate that mutation present in TN domain of
NKX2.5 gene will result loss of
NKX2.5 and
GATA4 interaction, thus will lead to loss of several activator and repressor function of this complex.
GATA4 p.M310T in future studies we also need show any effect activator and repressor function have trachyarrythemia. Further investigation is needed for the different phenotypes with the same genotype in the ASD family with arrhythmias.
CHD is a heavy load for the young family and the whole society. In China, the prevalence is still high although many CHD fetuses were aborted when the prenatal screening predicted highly possible of CHD. In the past two decades, cardiac development and genetic studies had provided much detailed information and identified many critical genes in the development of the heart. Although many mutants of these genes had been screened out, more biochemical methods and vivo models should be adopted for confirming. A limitation of this study is that only genetic screening methods had been employed but no furthermore study, such as vivo models, were used. Further studies will be conducted in the future to study the pathogenic mechanism of ASD families with arrhythmias and the reason for different phenotypes with the same genotype.
Publisher's Note
Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.