CTCF-mediated insulation and chromatin environment modulate Car5b escape from X inactivation
Abstract Background Genes that escape X-chromosome inactivation (XCI) in female somatic cells vary in number and levels of escape among mammalian species and tissues, potentially contributing to species- and tissue-specific sex differences. CTCF, a master chromatin conformation regulator, is enriche...
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2025-03-01
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| Online Access: | https://doi.org/10.1186/s12915-025-02137-7 |
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| author | He Fang Ana R. Tronco Giancarlo Bonora Truong Nguyen Jitendra Thakur Joel B. Berletch Galina N. Filippova Steven Henikoff Jay Shendure William S. Noble Zhijun Duan Christine M. Disteche Xinxian Deng |
| author_facet | He Fang Ana R. Tronco Giancarlo Bonora Truong Nguyen Jitendra Thakur Joel B. Berletch Galina N. Filippova Steven Henikoff Jay Shendure William S. Noble Zhijun Duan Christine M. Disteche Xinxian Deng |
| author_sort | He Fang |
| collection | DOAJ |
| description | Abstract Background Genes that escape X-chromosome inactivation (XCI) in female somatic cells vary in number and levels of escape among mammalian species and tissues, potentially contributing to species- and tissue-specific sex differences. CTCF, a master chromatin conformation regulator, is enriched at escape regions and may play an important role in regulating escape, but the molecular mechanisms remain elusive. Results CTCF binding profiles and epigenetic features were systematically examined at escape genes (escapees) using mouse allelic systems with skewed XCI to distinguish the inactive X (Xi) and active X (Xa) chromosomes. We found that six constitutive and two facultative escapees are located inside 30-800 kb domains marked by convergent arrays of CTCF binding sites, consistent with the formation of chromatin loops. Facultative escapees show clear differences in CTCF binding depending on their XCI status in specific cell types/tissues. In addition, sets of strong and in some cases divergent CTCF binding sites located at the boundary between an escapee and its adjacent neighbors subject to XCI would also help insulate domains. Indeed, deletion but not inversion of a CTCF binding site at the boundary between the facultative escapee Car5b and its silent neighbor Siah1b results in a dramatic reduction of Car5b escape. This is associated with reduced CTCF and cohesin binding, which indicates loss of looping and insulation and is supported by 3C combined with Hi-C analysis. In addition, enrichment in the repressive mark H3K27me3 invades the Car5b domain in deleted cells, consistent with loss of expression from the Xi. In contrast, cells with an inversion of the CTCF binding site retain CTCF and cohesin binding, as well as looping, in line with persistence of escape. Interestingly, the levels of escape increase in cells with deletion of either Dxz4, which disrupts the Xi-specific compact 3D structure, or Firre, which results in lower H3K27me3 enrichment on the Xi, indicating that the structural and epigenetic features of the Xi constrain escape from XCI in wild type conditions. Conclusions Taken together, our findings support the idea that escape from XCI in female somatic cells is modulated by both the topological insulation of domains via CTCF binding and the surrounding heterochromatin environment. |
| format | Article |
| id | doaj-art-cffddc7ea7954b4fb57c66f5479a0aeb |
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| publishDate | 2025-03-01 |
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| spelling | doaj-art-cffddc7ea7954b4fb57c66f5479a0aeb2025-08-20T03:05:44ZengBMCBMC Biology1741-70072025-03-0123112010.1186/s12915-025-02137-7CTCF-mediated insulation and chromatin environment modulate Car5b escape from X inactivationHe Fang0Ana R. Tronco1Giancarlo Bonora2Truong Nguyen3Jitendra Thakur4Joel B. Berletch5Galina N. Filippova6Steven Henikoff7Jay Shendure8William S. Noble9Zhijun Duan10Christine M. Disteche11Xinxian Deng12Department of Laboratory Medicine and Pathology, University of WashingtonDepartment of Laboratory Medicine and Pathology, University of WashingtonDepartment of Genome Sciences, University of WashingtonDepartment of Laboratory Medicine and Pathology, University of WashingtonBasic Sciences Division, Fred Hutchinson Cancer Research CenterDepartment of Laboratory Medicine and Pathology, University of WashingtonDepartment of Laboratory Medicine and Pathology, University of WashingtonBasic Sciences Division, Fred Hutchinson Cancer Research CenterDepartment of Genome Sciences, University of WashingtonDepartment of Genome Sciences, University of WashingtonInstitute for Stem Cell and Regenerative Medicine, University of WashingtonDepartment of Laboratory Medicine and Pathology, University of WashingtonDepartment of Laboratory Medicine and Pathology, University of WashingtonAbstract Background Genes that escape X-chromosome inactivation (XCI) in female somatic cells vary in number and levels of escape among mammalian species and tissues, potentially contributing to species- and tissue-specific sex differences. CTCF, a master chromatin conformation regulator, is enriched at escape regions and may play an important role in regulating escape, but the molecular mechanisms remain elusive. Results CTCF binding profiles and epigenetic features were systematically examined at escape genes (escapees) using mouse allelic systems with skewed XCI to distinguish the inactive X (Xi) and active X (Xa) chromosomes. We found that six constitutive and two facultative escapees are located inside 30-800 kb domains marked by convergent arrays of CTCF binding sites, consistent with the formation of chromatin loops. Facultative escapees show clear differences in CTCF binding depending on their XCI status in specific cell types/tissues. In addition, sets of strong and in some cases divergent CTCF binding sites located at the boundary between an escapee and its adjacent neighbors subject to XCI would also help insulate domains. Indeed, deletion but not inversion of a CTCF binding site at the boundary between the facultative escapee Car5b and its silent neighbor Siah1b results in a dramatic reduction of Car5b escape. This is associated with reduced CTCF and cohesin binding, which indicates loss of looping and insulation and is supported by 3C combined with Hi-C analysis. In addition, enrichment in the repressive mark H3K27me3 invades the Car5b domain in deleted cells, consistent with loss of expression from the Xi. In contrast, cells with an inversion of the CTCF binding site retain CTCF and cohesin binding, as well as looping, in line with persistence of escape. Interestingly, the levels of escape increase in cells with deletion of either Dxz4, which disrupts the Xi-specific compact 3D structure, or Firre, which results in lower H3K27me3 enrichment on the Xi, indicating that the structural and epigenetic features of the Xi constrain escape from XCI in wild type conditions. Conclusions Taken together, our findings support the idea that escape from XCI in female somatic cells is modulated by both the topological insulation of domains via CTCF binding and the surrounding heterochromatin environment.https://doi.org/10.1186/s12915-025-02137-7CTCFChromatin loopingInsulationX-chromosome inactivationEscape from X-chromosome inactivation |
| spellingShingle | He Fang Ana R. Tronco Giancarlo Bonora Truong Nguyen Jitendra Thakur Joel B. Berletch Galina N. Filippova Steven Henikoff Jay Shendure William S. Noble Zhijun Duan Christine M. Disteche Xinxian Deng CTCF-mediated insulation and chromatin environment modulate Car5b escape from X inactivation BMC Biology CTCF Chromatin looping Insulation X-chromosome inactivation Escape from X-chromosome inactivation |
| title | CTCF-mediated insulation and chromatin environment modulate Car5b escape from X inactivation |
| title_full | CTCF-mediated insulation and chromatin environment modulate Car5b escape from X inactivation |
| title_fullStr | CTCF-mediated insulation and chromatin environment modulate Car5b escape from X inactivation |
| title_full_unstemmed | CTCF-mediated insulation and chromatin environment modulate Car5b escape from X inactivation |
| title_short | CTCF-mediated insulation and chromatin environment modulate Car5b escape from X inactivation |
| title_sort | ctcf mediated insulation and chromatin environment modulate car5b escape from x inactivation |
| topic | CTCF Chromatin looping Insulation X-chromosome inactivation Escape from X-chromosome inactivation |
| url | https://doi.org/10.1186/s12915-025-02137-7 |
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