X inactivation results in silencing of genes on one X chromosome in females. However, a number of genes escape from X inactivation. Persistence of escape genes in humans is largely unexplained, but is important for a normal phenotype. Indeed, women with a single X chromosome have Turner syndrome, due to haplo-insufficiency for escape genes. Thus, tissue-specific expression of sex-chromosomal genes is likely to be determined by a combination of gene copy number and dosage regulation in addition to hormonal influences. The role of escape genes in sex-specific differences and in sex chromosome aneuploidy is poorly understood. In this proposal we address the sex-specific expression and the role of escape genes in different tissues using genome-wide approaches. Based on our previous studies supported by this grant we will construct a new mouse model to evaluate escape from X inactivation in vivo. To determine expression from each X allele we will generate this model from two mouse species in which X inactivation is skewed, and use RNA-sequencing together with SNP identification to quantify allele-specific expression. The extent of X inactivation and escape for protein-coding genes and for non-coding RNA genes will be evaluated in different tissues and developmental stages. Escape genes are devoid of repressive histone modifications associated with X inactivation, such as tri-methylation of histone H3 lysine 27. Our preliminary evidence indicates that the histone demethylase KDM6A is associated with escape genes in early ES cell differentiation. Thus, we will manipulate the dose of this gene to determine how expression influences X-linked gene expression. The higher expression of escape genes in females suggests that these genes have roles in female-specific functions or processes. One example is Xist, an escape gene that produces a non-coding RNA essential for the onset of X inactivation during early development. An important factor in silencing by repressive chromatin modifications is the position of the inactive X in nuclei. We discovered two regions that bind CTCF only on the inactive X and produce non-coding RNAs that escape X inactivation. In fact, one of these genes is more highly expressed from the inactive than the active X. Both regions are associated with the nucleolus, a nuclear compartment that plays a role in maintenance of heterochromatin. We will examine the role of these regions by knockdown of the long non-coding RNAs in ES cells and in a mouse knockout. Our study will help understanding the role of escape genes in normal development, and in diseases associated with sex chromosome aneuploidy.

Public Health Relevance

X inactivation is an important process required to balance gene dosage between males and females. Equally important are those genes that escape X inactivation, which causes sex- specific differences in gene expression levels and play a role in sex chromosome disorders. Our proposal aims at a better understanding of the function of escape genes in relation to female- specific processes such as X inactivation.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
2R01GM046883-19
Application #
8205564
Study Section
Special Emphasis Panel (ZRG1-GGG-N (03))
Program Officer
Carter, Anthony D
Project Start
1992-08-01
Project End
2015-07-31
Budget Start
2011-09-30
Budget End
2012-07-31
Support Year
19
Fiscal Year
2011
Total Cost
$323,505
Indirect Cost
Name
University of Washington
Department
Pathology
Type
Schools of Medicine
DUNS #
605799469
City
Seattle
State
WA
Country
United States
Zip Code
98195
Keown, Christopher L; Berletch, Joel B; Castanon, Rosa et al. (2017) Allele-specific non-CG DNA methylation marks domains of active chromatin in female mouse brain. Proc Natl Acad Sci U S A 114:E2882-E2890
Wei, Gengze; Deng, Xinxian; Agarwal, Saurabh et al. (2016) Patient Mutations of the Intellectual Disability Gene KDM5C Downregulate Netrin G2 and Suppress Neurite Growth in Neuro2a Cells. J Mol Neurosci 60:33-45
Disteche, Christine M (2016) Dosage compensation of the sex chromosomes and autosomes. Semin Cell Dev Biol 56:9-18
Berletch, Joel B; Ma, Wenxiu; Yang, Fan et al. (2015) Identification of genes escaping X inactivation by allelic expression analysis in a novel hybrid mouse model. Data Brief 5:761-9
Berletch, Joel B; Ma, Wenxiu; Yang, Fan et al. (2015) Escape from X inactivation varies in mouse tissues. PLoS Genet 11:e1005079
Yang, Fan; Deng, Xinxian; Ma, Wenxiu et al. (2015) The lncRNA Firre anchors the inactive X chromosome to the nucleolus by binding CTCF and maintains H3K27me3 methylation. Genome Biol 16:52
Deng, Xinxian; Ma, Wenxiu; Ramani, Vijay et al. (2015) Bipartite structure of the inactive mouse X chromosome. Genome Biol 16:152
Ma, Wenxiu; Ay, Ferhat; Lee, Choli et al. (2015) Fine-scale chromatin interaction maps reveal the cis-regulatory landscape of human lincRNA genes. Nat Methods 12:71-8
Disteche, Christine M; Berletch, Joel B (2015) X-chromosome inactivation and escape. J Genet 94:591-9
Deng, Xinxian; Berletch, Joel B; Nguyen, Di K et al. (2014) X chromosome regulation: diverse patterns in development, tissues and disease. Nat Rev Genet 15:367-78

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