ESPE Abstracts (2019) 92 P1-140

Methylation Status of X Inactivation-Escape Genes in Controls and Females with X Chromosome Rearrangements

Sayaka Kawashima1,2, Keiko Matsubara1, Machiko Toki3, Rika Kosaki4, Yukihiro Hasegawa5, Maki Fukami1, Masayo Kagami1


1Department of Molecular Endocrinology, National Research Institute for Child Health and Development, Tokyo, Japan. 2Department of Pediatrics, Tohoku University School of Medicine, Sendai, Japan. 3Department of Pediatrics, Keio University School of Medicine, Tokyo, Japan. 4Division of Medical Genetics, National Center for Child Health and Development, Tokyo, Japan. 5Division of Endocrinology and Metabolism Tokyo Metropolitan Children's Medical Center, Tokyo, Japan


Context: X chromosome inactivation (XCI) is a process in which one of the two X chromosomes in a female is randomly inactivated in order to correct gene dosage between males and females. However, about 15% of genes escape from XCI (termed escapees), and 10% of genes are variably inactivated (variable genes). The mechanism of inactivation and escape remains to be revealed. The promoter regions of escapees are hypomethylated compared to those of the inactivated genes. The objects of this study are to confirm validity to predict escape genes by comparing methylation status of promoters between male and female controls and to reveal the influence on the methylation status of promoters of escapees in patients with X chromosome rearrangements.

Subjects: Four patients (XX) with X chromosome rearrangements and 11 female and 12 male controls.

Methods: We performed array-based methylation analysis on peripheral blood of patients and controls using Infinium MethylationEPIC BeadChip. We examined the methylation levels of the promoter regions located within 1 kb up and downstream of transcription start sites. In controls, we regarded the genes of which promoter regions showed equally hypomethylated in both sexes as "escapees".

The breakpoints of X chromosome rearrangements were determined by whole genome sequencing and Sanger sequencing.

Results: 51 genes were detected as escapee in controls. 74 % of them had previously reported as an escapee or a variable gene. One patient with 46,X,der(X)(pter→p22.13::p11.23→p11.23::p11.23→q23::q22.1→q21.31::q22.2→q24::q23→q24::p22.11→p11.4::p22.11→p22.13::q22.1→q22.1::p22.12→pter) showed hypermethylation in five escapees. These genes could be partially inactivated. These genes were located in a duplicated region, however, most of the escapees at the duplicated region and all the escapees in regions with normal copy number remained hypomethylated in this patient. Three other patients did not have any hypermethylated escapees although the patients had duplications at Xp.

Discussion: Most escapees ascertained in this study were previously reported as escapees or variable genes. X chromosome rearrangements in a single patient affected the methylation levels of the promoters of five escapees. This finding suggest that structural abnormalities on X chromosome can affect the methylation levels of promoter regions in some escapees to result in inactivation.

Conclusion: We successfully determined escapees by comparing methylation status of promoters between male and female controls. Specific X chromosome rearrangements likely affect the methylation status of promoters of some escapees.

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