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Series GSE268545

Query DataSets for GSE268545

Status

Public on Nov 28, 2024

Title

ALKBH1-mediated DNA N6-methyladenine Modification Regulates H3K9me3-dependent Heterochromatin in Neural Tube Development [Cut & Tag]

Organism

Mus musculus

Experiment type

Genome binding/occupancy profiling by high throughput sequencing

Summary

Neural Tube Defects (NTDs) are a class of severe congenital developmental defects caused by abnormal closure of the neural tube during early embryonic development, including conditions such as anencephaly, spina bifida, and encephalocele. Currently, NTDs are considered to result from the combined effects of genetic and environmental factors, particularly maternal folate intake during pregnancy, leading to widespread epigenetic dysregulation. However, the role of N6-methyladenine (N6-mA) in embryonic neural development remains largely unknown. Here, we found significant upregulation of Alkbh1 and concomitant significant downregulation of overall 6mA in brain tissue of a mouse model of NTDs. Further investigation through MeDIP, CUT&TAG, and single-cell sequencing revealed crosstalk between N6-mA and H3K9me3-marked heterochromatin, primarily impacting the involvement of embryonic glial cells in the formation of the hindbrain during embryonic neural tube development. These findings collectively suggest a potential epigenetic role of N6-mA in mammalian brain development, potentially exhibiting cell type specificity.

Overall design

1.Establishment of NTD Mouse Model: MTX is a folate antagonist. Under conditions of low folate diet in pregnant mice, we intraperitoneally injected MTX (1.5 mg/kg) at gestational day 7.5. We began observing the closure status of the neural tube in fetal mice during the neural tube closure window at E9.5-10.5. Normal mice appeared plump and round, with fully closed neural tubes, and were categorized into the normal group. Embryos from mice on low folate diet and injected with MTX exhibited varying degrees of developmental delay, smaller embryos, and primarily showed phenotypes of hindbrain malformations and spina bifida, and were categorized into the NTD group. CUT&Tag was performed on normal and NTD fetal brains to investigate the genomic localization of H3K9me3 and identify genomic regions enriched with H3K9me3. 2.Constructing a low-folate cell line：After resuscitating normal mESCs, Sv129 cells were randomly split into FA4 and FA0. The FA0 group uses folate-free mESCs culture medium.Folate (Sigma) was added to the medium of FA0 as medium of the FA4 group(final concentration: 4 mg/L). Next, two groups were added correspond medium and were cultured at 37°C with 5% CO2 in a humidified incubator and passaged every 2–3 days. The culture medium was changed daily. CUT&Tag was performed on FA4 and FA0 groups to investigate the genomic localization of H3K9me3 and identify genomic regions enriched with H3K9me3.

Contributor(s)

He X, Shen J, Wang S

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Submission date

May 29, 2024

Last update date

Nov 28, 2024

Contact name

Xuejia He

Organization name

Capital Institute of Pediatrics