GATA4 is known to be a causative gene for congenital heart disease, but has also now been associated with disorders of sexual development (DSD). We here report a pathogenic variant of GATA4 in a 46,XY DSD patient with an atrial septal defect, identified by whole‐exome sequencing to be c.487C>T (p.Pro163Ser). This mutation resulted in reduced transcriptional activity of the downstream gene. When we compared this transcriptional activity level with other GATA4 variants, those that had been identified in patients with cardiac defects and DSD showed less activity than those in patients with cardiac defect only. This suggests that the normal development of the heart requires more strict regulation of GATA4 transcription than testicular development. Further, when the different variants were co‐expressed with wild‐type, the transcriptional activities were consistently lower than would be expected from an additive effect, suggesting a dominant‐negative impact of the variant via dimer formation of the GATA4 protein. Since these pathogenic GATA4 variants are occasionally identified in healthy parents, a threshold model of quantitative traits may explain the cardiac defect or DSD phenotypes that they cause.

Objective: Xq chromosome duplication with complex rearrangements is generally acknowledged to be associated with neurodevelopmental disorders, such as Pelizaeus-Merzbacher disease (PMD) and MECP2 duplication syndrome. For couples who required a PGT-M (pre-implantation genetic testing for monogenic disease) for these disorders, junction-specific PCR is useful to directly detect pathogenic variants. Therefore, pre-clinical workup for PGT-M requires the identification of the junction of duplicated segments in PMD and MECP2 duplication syndrome, which is generally difficult. Methods:In this report, we used nanopore long-read sequencing targeting the X chromosome using an adaptive sampling method to identify breakpoint junctions in disease-causing triplications.Results: By long-read sequencing, we successfully identified breakpoint junctions in one PMD case with PLP1 triplication and in another MECP2 triplication case in a single sequencing run. Surprisingly, the duplicated region involving MECP2 was inserted 45 Mb proximal to the original position. This inserted region was confirmed by FISH analysis. With the help of precise mapping of the pathogenic variant, we successfully re-established STR haplotyping for PGT-M and avoided any potential misinterpretation of the pathogenic allele due to recombination. Conclusion:Long-read sequencing with adaptive sampling in a PGT-M pre-clinical workup is a beneficial method for identifying junctions of chromosomal complex structural rearrangements. Key pointsWhat's already known about this topic? � For couples who required a PGT-M (pre-implantation genetic testing for monogenic disease) for diseases with complex chromosomal structural rearrangements, junction-specific PCR is useful to directly detect pathogenic variants. Therefore, pre-clinical workup for 304 -