Familial cases of point mutations in the XIST promoter reveal a correlation between CTCF binding and pre-emptive choices of X chromosome inactivation

Abstract

The choice mechanisms that determine the future inactive X chromosome in somatic cells of female mammals involve the regulated expression of the XIST gene. A familial C(-43)G mutation in the XIST promoter results in skewing of X chromosome inactivation (XCI) towards the inactive X chromosome of heterozygous females, whereas a C(-43)A mutation found primarily in the active X chromosome results in the opposite skewing pattern. Both mutations point to the existence of a factor that might be responsible for the skewed patterns. Here we identify this factor as CTCF, a conserved protein with a 11 Zn-finger (ZF) domain that can mediate multiple sequence-specificity and interactions between DNA-bound CTCF molecules. We show that mouse and human Xist/XIST promoters contain one homologous CTCF-binding sequence with the matching dG-contacts, which in the human XIST include the -43 position within the DNase I footprint of CTCF. While the C(-43)A mutation abrogates CTCF binding, the C(-43)G mutation results in a dramatic increase in CTCF-binding efficiency by altering ZF-usage mode required for recognition of the altered dG-contacts of the mutant site. Thus, the skewing effect of the two -43C mutations correlates with their effects on CTCF binding. Finally, CTCF interacts with the XIST/Xist promoter only in female human and mouse cells. The interpretation that this reflected a preferential interaction with the promoter of the active Xist allele was confirmed in mouse fetal placenta. These observations are in keeping with the possibility that the choice of X chromosome inactivation reflects stabilization of a higher order chromatin conformation impinging on the CTCF-XIST promoter complex.

Genetic Control of X Chromosome Inactivation in Mice: Definition of the Xce Candidate Interval

“…In fact, in humans a rare variant found within the XIST promoter has been shown to cause significant skewing of the X inactivation pattern (Plenge et al 1997). This bias in determining the X inactivation pattern is thought to be the result of a dramatic increase in CTCF binding to the XIST promoter on the variant allele (Pugacheva et al 2005).…”

A Boundary Element BetweenTsixandXistBinds the Chromatin Insulator Ctcf and Contributes to Initiation of X-Chromosome Inactivation

“…We speculate that RS14 may organize chromatin architecture at the Xic. Apart from DRS14, Ctcf also binds to enhancer elements within Xite, to the 59 end of Tsix around the DXPas34 repeat, and to the Xist promoter (Chao et al 2002;Pugacheva et al 2005;Xu et al 2007). Interestingly, all of these Ctcf-binding sites occur at the bases of known looping interactions (Tsai et al 2008).…”

Familial skewed X-chromosome inactivation linked to a component of the cohesin complex, SA2

“…45 Mutations of the CTCF binding site within the XIST promoter result in skewed XCI choice. 46,47 XCI is initiated by transient colocalisation of the X-chromosomes, 48 and CTCF is required for such pairing. 49 The enhancer-blocking function of CTCF has been implicated in XCI.…”