Live cell imaging of the nascent inactive X chromosome during the early differentiation process of naive ES cells towards epiblast stem cells.
Random X-chromosome inactivation ensures dosage compensation in mammals through the transcriptional silencing of one of the two X chromosomes present in each female cell. Silencing is initiated in the differentiating epiblast of the mouse female embryos through coating of the nascent inactive X chro...
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doaj-3632c931d13f4fa19d431e19e9fd8b632020-11-25T02:48:44ZengPublic Library of Science (PLoS)PLoS ONE1932-62032014-01-01912e11610910.1371/journal.pone.0116109Live cell imaging of the nascent inactive X chromosome during the early differentiation process of naive ES cells towards epiblast stem cells.Aurélia GuyochinSylvain MaennerErin Tsi-Jia ChuAsma HentatiMikael AttiaPhilip AvnerPhilippe ClercRandom X-chromosome inactivation ensures dosage compensation in mammals through the transcriptional silencing of one of the two X chromosomes present in each female cell. Silencing is initiated in the differentiating epiblast of the mouse female embryos through coating of the nascent inactive X chromosome by the non-coding RNA Xist, which subsequently recruits the Polycomb Complex PRC2 leading to histone H3-K27 methylation. Here we examined in mouse ES cells the early steps of the transition from naive ES cells towards epiblast stem cells as a model for inducing X chromosome inactivation in vitro. We show that these conditions efficiently induce random XCI. Importantly, in a transient phase of this differentiation pathway, both X chromosomes are coated with Xist RNA in up to 15% of the XX cells. In an attempt to determine the dynamics of this process, we designed a strategy aimed at visualizing the nascent inactive X-chromosome in live cells. We generated transgenic female XX ES cells expressing the PRC2 component Ezh2 fused to the fluorescent protein Venus. The fluorescent fusion protein was expressed at sub-physiological levels and located in nuclei of ES cells. Upon differentiation of ES cell towards epiblast stem cell fate, Venus-fluorescent territories appearing in interphase nuclei were identified as nascent inactive X chromosomes by their association with Xist RNA. Imaging of Ezh2-Venus for up to 24 hours during the differentiation process showed survival of some cells with two fluorescent domains and a surprising dynamics of the fluorescent territories across cell division and in the course of the differentiation process. Our data reveal a strategy for visualizing the nascent inactive X chromosome and suggests the possibility for a large plasticity of the nascent inactive X chromosome.http://europepmc.org/articles/PMC4278889?pdf=render |
collection |
DOAJ |
language |
English |
format |
Article |
sources |
DOAJ |
author |
Aurélia Guyochin Sylvain Maenner Erin Tsi-Jia Chu Asma Hentati Mikael Attia Philip Avner Philippe Clerc |
spellingShingle |
Aurélia Guyochin Sylvain Maenner Erin Tsi-Jia Chu Asma Hentati Mikael Attia Philip Avner Philippe Clerc Live cell imaging of the nascent inactive X chromosome during the early differentiation process of naive ES cells towards epiblast stem cells. PLoS ONE |
author_facet |
Aurélia Guyochin Sylvain Maenner Erin Tsi-Jia Chu Asma Hentati Mikael Attia Philip Avner Philippe Clerc |
author_sort |
Aurélia Guyochin |
title |
Live cell imaging of the nascent inactive X chromosome during the early differentiation process of naive ES cells towards epiblast stem cells. |
title_short |
Live cell imaging of the nascent inactive X chromosome during the early differentiation process of naive ES cells towards epiblast stem cells. |
title_full |
Live cell imaging of the nascent inactive X chromosome during the early differentiation process of naive ES cells towards epiblast stem cells. |
title_fullStr |
Live cell imaging of the nascent inactive X chromosome during the early differentiation process of naive ES cells towards epiblast stem cells. |
title_full_unstemmed |
Live cell imaging of the nascent inactive X chromosome during the early differentiation process of naive ES cells towards epiblast stem cells. |
title_sort |
live cell imaging of the nascent inactive x chromosome during the early differentiation process of naive es cells towards epiblast stem cells. |
publisher |
Public Library of Science (PLoS) |
series |
PLoS ONE |
issn |
1932-6203 |
publishDate |
2014-01-01 |
description |
Random X-chromosome inactivation ensures dosage compensation in mammals through the transcriptional silencing of one of the two X chromosomes present in each female cell. Silencing is initiated in the differentiating epiblast of the mouse female embryos through coating of the nascent inactive X chromosome by the non-coding RNA Xist, which subsequently recruits the Polycomb Complex PRC2 leading to histone H3-K27 methylation. Here we examined in mouse ES cells the early steps of the transition from naive ES cells towards epiblast stem cells as a model for inducing X chromosome inactivation in vitro. We show that these conditions efficiently induce random XCI. Importantly, in a transient phase of this differentiation pathway, both X chromosomes are coated with Xist RNA in up to 15% of the XX cells. In an attempt to determine the dynamics of this process, we designed a strategy aimed at visualizing the nascent inactive X-chromosome in live cells. We generated transgenic female XX ES cells expressing the PRC2 component Ezh2 fused to the fluorescent protein Venus. The fluorescent fusion protein was expressed at sub-physiological levels and located in nuclei of ES cells. Upon differentiation of ES cell towards epiblast stem cell fate, Venus-fluorescent territories appearing in interphase nuclei were identified as nascent inactive X chromosomes by their association with Xist RNA. Imaging of Ezh2-Venus for up to 24 hours during the differentiation process showed survival of some cells with two fluorescent domains and a surprising dynamics of the fluorescent territories across cell division and in the course of the differentiation process. Our data reveal a strategy for visualizing the nascent inactive X chromosome and suggests the possibility for a large plasticity of the nascent inactive X chromosome. |
url |
http://europepmc.org/articles/PMC4278889?pdf=render |
work_keys_str_mv |
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