Identification and functional validation of variants in the promoter region of HAND1 gene in sporadic tetralogy of Fallot.

Background: Tetralogy of Fallot (TOF) is a common congenital heart disease (CHD) but the impact of the variants of the HAND1 gene promoter region has not been explored.

Methods: DNA from blood samples of 612 subjects (300 sporadic TOF patients and 312 healthy controls) was sequenced to identify variants in the HAND1 gene promoter region that were further tested by cellular function experiments including dual-luciferase reporter gene assays, electrophoretic mobility shift analysis (EMSA), and bioinformatics analysis using JASPAR, a transcription factor binding site database.

Results: Eight variants in HAND1 gene promoter region were identified with 3 only found in TOF patients including one novel g.3639 G > T. All 3 variants significantly reduced the transcriptional activity of HAND1 gene promoter in an in vitro reporter assay (p < 0.05). JASPAR analysis indicated that these variants may alter the binding sites of transcription factors, potentially associated with TOF formation.

Conclusions: In the promoter region of HAND1gene of TOF patients, 3 variants were found only in the patients including one found for the first time. These variants decreased transcription factor activity. Therefore, the present study implies the role of HAND1 gene in the pathogenesis of TOF and further provides new insights into the genetic basis of TOF formation.

Impact: Sequencing of DNA from 612 human subjects (300 TOF patients and 312 healthy controls) identified 8 variants in the promoter region of HAND1 gene with 3 found only in TOF including 1 newly identified. Reduced transcriptional activity at these 3 variants was confirmed by dual-luciferase reporter gene assay and EMSA assay. Predictions from the JASPAR database suggest altered binding sites of transcription factors by the variants. We for the first time demonstrate variants in the HAND1 promoter and that cause cellular dysfunction. The variants identified may have a pathological role in the formation of TOF.