Deleterious genetic variants comprise one cause of cardiac conotruncal defects (CTDs). Genes associated with CTDs are gradually being identified. In the present study, we aimed to explore the profile of genetic variants of CTD-associated genes in Chinese patients with non-syndromic CTDs.
Thirty-nine CTD-related genes were selected after reviewing published articles in NCBI, HGMD, OMIM, and HPO. In total, 605 patients with non-syndromic CTDs and 300 healthy controls, all of Han ethnicity, were recruited. High-throughput targeted sequencing was used to detect genetic variants in the protein-coding regions of genes. We performed rigorous variant-level filtrations to identify potentially damaging variants (Dvars) using prediction programs including CADD, SIFT, PolyPhen-2, and MutationTaster.
Dvars were detected in 66.7% (26/39) of the targeted CTD-associated genes. In total, 11.07% (67/605) of patients with non-syndromic CTDs were found to carry one or more Dvars in targeted CTD-associated genes. Dvars in FOXH1, TBX2, NFATC1, FOXC2, and FOXC1 were common in the CTD cohort (1.5% [9/605], 1.2% [7/605], 1.2% [7/605], 1% [6/605], and 0.5% [3/605], respectively).
Targeted exon sequencing is a cost-effective approach for the genetic diagnosis of CTDs. Our findings contribute to an understanding of the genetic architecture of non-syndromic CTDs.

The embryonic process of forming a complex structure such as the heart remains poorly understood. Here, we show that Six2 marks a dynamic subset of second heart field progenitors. Six2-positive (Six2+) progenitors are rapidly recruited and assigned, and their descendants are allocated successively to regions of the heart from the right ventricle (RV) to the pulmonary trunk. Global ablation of Six2+ progenitors resulted in RV hypoplasia and pulmonary atresia. An early stage-specific ablation of a small subset of Six2+ progenitors did not cause any apparent structural defect at birth but rather resulted in adult-onset cardiac hypertrophy and dysfunction. Furthermore, Six2 expression depends in part on Shh signaling, and Shh deletion resulted in severe deficiency of Six2+ progenitors. Collectively, these findings unveil the chronological features of cardiogenesis, in which the mammalian heart is built sequentially by temporally distinct populations of cardiac progenitors, and provide insights into late-onset congenital heart disease.

Congenital diseases caused by abnormal development of the cranial neural crest usually present craniofacial malformations and heart defects while the precise mechanism is not fully understood. Here, we show that the zebrafish eif3ba mutant caused by pseudo-typed retrovirus insertion exhibited a similar phenotype due to the hypogenesis of cranial neural crest cells (NCCs). The derivatives of cranial NCCs, including the NCC-derived cell population of pharyngeal arches, craniofacial cartilage, pigment cells and the myocardium derived from cardiac NCCs, were affected in this mutant. The expression of several neural crest marker genes, including crestin, dlx2a and nrp2b, was specifically reduced in the cranial regions of the eif3ba mutant. Through fluorescence-tracing of the cranial NCC migration marker nrp2b, we observed reduced intensity of NCC-derived cells in the heart. In addition, p53 was markedly up-regulated in the eif3ba mutant embryos, which correlated with pronounced apoptosis in the cranial area as shown by TUNEL staining. These findings suggest a novel function of eif3ba during embryonic development and a novel level of regulation in the process of cranial NCC development, in addition to providing a potential animal model to mimic congenital diseases due to cranial NCC defects. Furthermore, we report the identification of a novel transgenic fish line Et(gata2a:EGFP)pku418 to trace the migration of cranial NCCs (including cardiac NCCs); this may serve as an invaluable tool for investigating the development and dynamics of cranial NCCs during zebrafish embryogenesis.