Spatial separation of Xist RNA and polycomb proteins revealed by superresolution microscopy
Andrea Cerase1, Daniel Smeets, Y Amy Tang
1Department of Biochemistry, University of Oxford, Oxford OX1 3QU, United Kingdom.
Summary
X chromosome inactivation involves Xist RNA recruiting Polycomb Repressive Complex 2 (PRC2). This study found PRC2 binding sites do not correlate with gene silencing and PRC2 does not colocalize with Xist RNA, challenging direct interaction models.
Area of Science:
- Epigenetics
- Genetics
- Molecular Biology
Background:
- X chromosome inactivation (XCI) equalizes gene dosage in female mammals by silencing one X chromosome.
- X-inactive specific transcript (Xist) RNA is the master regulator, recruiting chromatin modifiers like Polycomb Repressive Complex 2 (PRC2).
- A direct interaction between Xist RNA's 5' A-repeat region and PRC2 proteins (Ezh2, Suz12) has been proposed.
Purpose of the Study:
- To investigate the mechanism of Xist-mediated PRC2 recruitment in XCI.
- To determine the spatial relationship between Xist RNA and PRC2 during XCI.
Main Methods:
- Microarray-based epigenomic mapping of PRC2 binding sites after Xist induction.
- Superresolution 3D structured illumination microscopy to visualize Xist RNA and PRC2 localization.
- Analysis in an ES cell line with an inducible Xist transgene and in normal XX somatic cells.
Main Results:
- Induced Xist expression led to PRC2 binding predominantly in gene-rich regions and gene bodies.
- Newly deposited PRC2 sites did not correlate with Xist-mediated gene silencing.
- 3D microscopy revealed significant spatial separation and lack of colocalization between Xist RNA and PRC2 proteins.
Conclusions:
- The findings challenge the model of direct interaction between Xist RNA and PRC2.
- A reappraisal of the mechanism for PRC2 recruitment in X chromosome inactivation is warranted.
- Xist RNA may recruit PRC2 indirectly, or PRC2 may be recruited by other factors in XCI.
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