Limits...
Developmentally regulated alterations in Polycomb repressive complex 1 proteins on the inactive X chromosome.

Plath K, Talbot D, Hamer KM, Otte AP, Yang TP, Jaenisch R, Panning B - J. Cell Biol. (2004)

Bottom Line: In this study, we demonstrate that mPRC1 proteins localize to the Xi, and that different mPRC1 proteins accumulate on the Xi during initiation and maintenance of X inactivation in embryonic cells.The Xi accumulation of mPRC1 proteins requires Xist RNA and is not solely regulated by the presence of H3-K27 methylation, as not all cells that exhibit this epigenetic mark on the Xi show Xi enrichment of mPRC1 proteins.Our results implicate mPRC1 in X inactivation and suggest that the regulated assembly of PcG protein complexes on the Xi contributes to this multistep process.

View Article: PubMed Central - PubMed

Affiliation: Whitehead Institute for Biomedical Research, Cambridge, MA 02142, USA. plath@wi.mit.edu

ABSTRACT
Polycomb group (PcG) proteins belonging to the polycomb (Pc) repressive complexes 1 and 2 (PRC1 and PRC2) maintain homeotic gene silencing. In Drosophila, PRC2 methylates histone H3 on lysine 27, and this epigenetic mark facilitates recruitment of PRC1. Mouse PRC2 (mPRC2) has been implicated in X inactivation, as mPRC2 proteins transiently accumulate on the inactive X chromosome (Xi) at the onset of X inactivation to methylate histone H3 lysine 27 (H3-K27). In this study, we demonstrate that mPRC1 proteins localize to the Xi, and that different mPRC1 proteins accumulate on the Xi during initiation and maintenance of X inactivation in embryonic cells. The Xi accumulation of mPRC1 proteins requires Xist RNA and is not solely regulated by the presence of H3-K27 methylation, as not all cells that exhibit this epigenetic mark on the Xi show Xi enrichment of mPRC1 proteins. Our results implicate mPRC1 in X inactivation and suggest that the regulated assembly of PcG protein complexes on the Xi contributes to this multistep process.

Show MeSH
Immunolocalization of human PRC1 proteins in 293 cells. (a) 293 cells were stained for H3-3mK27 (third column) in combination with FISH for XIST RNA to detect the Xi (second column). Nuclei were stained with DAPI (first column) and merged image (fourth column) consists of XIST RNA in green and H3-3mK27 in red. 293 cells contain two Xis and virtually all cells showed Xi accumulation of H3-3mK27 (unpublished data). (b) Immunostaining for H3-3mK27 (third column) and hPRC1 proteins (second column) in 293 cells. DAPI staining (first column) was used to mark the nuclei and the merge (fourth column) represents H3-3mK27 in red and hPRC1 proteins in green. The fraction of cells with an H3-3mK27-marked Xi that also exhibited Xi enrichment for each mPRC1 protein assayed is indicated on the right. All percentages are based on counts of >75 cells.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC2172612&req=5

fig3: Immunolocalization of human PRC1 proteins in 293 cells. (a) 293 cells were stained for H3-3mK27 (third column) in combination with FISH for XIST RNA to detect the Xi (second column). Nuclei were stained with DAPI (first column) and merged image (fourth column) consists of XIST RNA in green and H3-3mK27 in red. 293 cells contain two Xis and virtually all cells showed Xi accumulation of H3-3mK27 (unpublished data). (b) Immunostaining for H3-3mK27 (third column) and hPRC1 proteins (second column) in 293 cells. DAPI staining (first column) was used to mark the nuclei and the merge (fourth column) represents H3-3mK27 in red and hPRC1 proteins in green. The fraction of cells with an H3-3mK27-marked Xi that also exhibited Xi enrichment for each mPRC1 protein assayed is indicated on the right. All percentages are based on counts of >75 cells.

Mentions: The percentages of MEFs with Xi enrichment of mPRC1 proteins and the standard errors are based on counts of 100–400 cells in each of three to nine independent experiments similar to that shown in Fig. 1. Percentages of cells with mPRC1 Xi enrichment in differentiating ES cells and undifferentiated ES cells ectopically expressing Xist are taken from Figs. 5 and 6. The percentages of TS and 293 cells with Xi enrichment of PRC1 proteins are based on counts of >75 cells in each of two or more experiments similar to those displayed in Figs. 2 and 3. H3-3mK27 percentages in all cell types are based on counts of at least 100 cells in two or more experiments. In all cases, only interphase cells were counted.


Developmentally regulated alterations in Polycomb repressive complex 1 proteins on the inactive X chromosome.

Plath K, Talbot D, Hamer KM, Otte AP, Yang TP, Jaenisch R, Panning B - J. Cell Biol. (2004)

Immunolocalization of human PRC1 proteins in 293 cells. (a) 293 cells were stained for H3-3mK27 (third column) in combination with FISH for XIST RNA to detect the Xi (second column). Nuclei were stained with DAPI (first column) and merged image (fourth column) consists of XIST RNA in green and H3-3mK27 in red. 293 cells contain two Xis and virtually all cells showed Xi accumulation of H3-3mK27 (unpublished data). (b) Immunostaining for H3-3mK27 (third column) and hPRC1 proteins (second column) in 293 cells. DAPI staining (first column) was used to mark the nuclei and the merge (fourth column) represents H3-3mK27 in red and hPRC1 proteins in green. The fraction of cells with an H3-3mK27-marked Xi that also exhibited Xi enrichment for each mPRC1 protein assayed is indicated on the right. All percentages are based on counts of >75 cells.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2172612&req=5

fig3: Immunolocalization of human PRC1 proteins in 293 cells. (a) 293 cells were stained for H3-3mK27 (third column) in combination with FISH for XIST RNA to detect the Xi (second column). Nuclei were stained with DAPI (first column) and merged image (fourth column) consists of XIST RNA in green and H3-3mK27 in red. 293 cells contain two Xis and virtually all cells showed Xi accumulation of H3-3mK27 (unpublished data). (b) Immunostaining for H3-3mK27 (third column) and hPRC1 proteins (second column) in 293 cells. DAPI staining (first column) was used to mark the nuclei and the merge (fourth column) represents H3-3mK27 in red and hPRC1 proteins in green. The fraction of cells with an H3-3mK27-marked Xi that also exhibited Xi enrichment for each mPRC1 protein assayed is indicated on the right. All percentages are based on counts of >75 cells.
Mentions: The percentages of MEFs with Xi enrichment of mPRC1 proteins and the standard errors are based on counts of 100–400 cells in each of three to nine independent experiments similar to that shown in Fig. 1. Percentages of cells with mPRC1 Xi enrichment in differentiating ES cells and undifferentiated ES cells ectopically expressing Xist are taken from Figs. 5 and 6. The percentages of TS and 293 cells with Xi enrichment of PRC1 proteins are based on counts of >75 cells in each of two or more experiments similar to those displayed in Figs. 2 and 3. H3-3mK27 percentages in all cell types are based on counts of at least 100 cells in two or more experiments. In all cases, only interphase cells were counted.

Bottom Line: In this study, we demonstrate that mPRC1 proteins localize to the Xi, and that different mPRC1 proteins accumulate on the Xi during initiation and maintenance of X inactivation in embryonic cells.The Xi accumulation of mPRC1 proteins requires Xist RNA and is not solely regulated by the presence of H3-K27 methylation, as not all cells that exhibit this epigenetic mark on the Xi show Xi enrichment of mPRC1 proteins.Our results implicate mPRC1 in X inactivation and suggest that the regulated assembly of PcG protein complexes on the Xi contributes to this multistep process.

View Article: PubMed Central - PubMed

Affiliation: Whitehead Institute for Biomedical Research, Cambridge, MA 02142, USA. plath@wi.mit.edu

ABSTRACT
Polycomb group (PcG) proteins belonging to the polycomb (Pc) repressive complexes 1 and 2 (PRC1 and PRC2) maintain homeotic gene silencing. In Drosophila, PRC2 methylates histone H3 on lysine 27, and this epigenetic mark facilitates recruitment of PRC1. Mouse PRC2 (mPRC2) has been implicated in X inactivation, as mPRC2 proteins transiently accumulate on the inactive X chromosome (Xi) at the onset of X inactivation to methylate histone H3 lysine 27 (H3-K27). In this study, we demonstrate that mPRC1 proteins localize to the Xi, and that different mPRC1 proteins accumulate on the Xi during initiation and maintenance of X inactivation in embryonic cells. The Xi accumulation of mPRC1 proteins requires Xist RNA and is not solely regulated by the presence of H3-K27 methylation, as not all cells that exhibit this epigenetic mark on the Xi show Xi enrichment of mPRC1 proteins. Our results implicate mPRC1 in X inactivation and suggest that the regulated assembly of PcG protein complexes on the Xi contributes to this multistep process.

Show MeSH