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Metadata Commons Identifier

HMC000128

Project Title

Impact of Flanking Chromosomal Sequences on Localization and Silencing by the ncRNA XIST

Project Description

Background: X-chromosome inactivation is a striking example of epigenetic silencing in which expression of the long non-coding RNA XIST initiates the heterochromatinization and silencing of one of the pair of X chromosomes in mammalian females. To understand how the RNA can establish silencing across millions of basepairs of DNA we have modelled the process by inducing expression of XIST from nine different locations in human HT1080 cells.Results: Localization of XIST, depletion of Cot-1 RNA, perinuclear localization, and ubiquitination of H2A occurs at all sites examined, while recruitment of H3K9me3 was not observed. Recruitment of the heterochromatic features SMCHD1, macroH2A, H3K27me3, and H4K20me1 occurs independently of each other in an integration site-dependent manner. Silencing of flanking reporter genes occurs at all sites, but the spread of silencing to flanking endogenous human genes is variable in extent of silencing as well as extent of spread, with silencing able to skip regions. The spread of H3K27me3 and loss of H3K27ac correlates with the pre-existing levels of the modifications, and overall the extent of silencing correlates with the ability to recruit additional heterochromatic features.Conclusions: The non-coding RNA XIST functions as a cis-acting silencer when expressed from nine different locations throughout the genome. A hierarchy among the features of heterochromatin reveals the importance of interaction with the local chromatin neighborhood for optimal spread of silencing, as well as the independent yet cooperative nature of the establishment of heterochromatin by the non-coding XIST RNAOmics: We performed RNA-seq and ChIP-seq on clones of human cell lines carrying an inducible XIST transgene on 1p, 8p, or 12q to study the effects of allelic silencing in cis. Hi-C dataset from <a href="https://pubmed.ncbi.nlm.nih.gov/22495300/">Jesse R Dixon et al</a> were integrated into the data analysis.

Project Funder(s)

Canadian Institutes of Health Research (CIHR)

Project Institution(s)

University of British Columbia

Project Investigator(s)

Christine Yang, Danny Leung, Carolyn Brown

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Keywords

XIST, Long non-coding RNA, Dosage compensation, X-chromosome inactivation, Nucleolar localization, Facultative heterochromatin, SMCHD1, macroH2A, H3K27me3, H4K20me1

Publication Link

https://pubmed.ncbi.nlm.nih.gov/26429547

Study Completed

Cohort Name

Human cell lines carrying an inducible XIST transgene

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Disease/Condition Studied

transgene impact, adult fibrosarcoma, X chromosome inactivation

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