A system to analyze the initiation of random X-chromosome inactivation using time-lapse imaging of single cells

In female eutherian mammal development, X-chromosome inactivation (XCI) of one of the two X chromosomes is initiated early. Understanding the relationship between the initiation of XCI and cell fate is critical for understanding early female development and requires a system that can monitor XCI in...

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Bibliographic Details
Published in:Scientific reports 2024-09, Vol.14 (1), p.20327-12, Article 20327
Main Authors: Koshiguchi, Manami, Yonezawa, Nao, Hatano, Yu, Suenaga, Hikaru, Yamagata, Kazuo, Kobayashi, Shin
Format: Article
Language:English
Subjects:
631/136/2086
631/1647/2210
631/337/176/1433
Animals
Cell culture
Cell differentiation
Cell Differentiation - genetics
Cell disruption
Cell fate
Cell Line
Chromosomes
Embryo cells
Embryogenesis
Embryonic Stem Cells - cytology
Embryonic Stem Cells - metabolism
Female
Females
Genes, Reporter
Humanities and Social Sciences
Inactivation
Karyotypes
Mice
multidisciplinary
Neomycin
Puromycin
Science
Science (multidisciplinary)
Single-Cell Analysis - methods
Stem cells
Time-Lapse Imaging - methods
X Chromosome - genetics
X Chromosome Inactivation - genetics
X chromosomes
X-chromosome inactivation
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Summary:In female eutherian mammal development, X-chromosome inactivation (XCI) of one of the two X chromosomes is initiated early. Understanding the relationship between the initiation of XCI and cell fate is critical for understanding early female development and requires a system that can monitor XCI in single living cells. Traditional embryonic stem cells (ESCs) used for XCI studies often lose X chromosomes spontaneously during culture and differentiation, making accurate monitoring difficult. Additionally, most XCI assessment methods necessitate cell disruption, hindering cell fate tracking. We developed the Momiji (version 2) ESC line to address these difficulties, enabling real-time monitoring of X-chromosome activity via fluorescence. We inserted green and red fluorescent reporter genes and neomycin and puromycin resistance genes into the two X chromosomes of PGK12.1 ESCs, creating a female ESC line that retains two X chromosomes more faithfully during differentiation. Momiji (version 2) ESCs exhibit a more stable XX karyotype than other ESC lines, including the parental PGK12.1 line. This new tool offers valuable insights into the relationship between XCI and cell fate, improving our understanding of early female development.
ISSN:2045-2322
2045-2322
DOI:10.1038/s41598-024-71105-y