RIF1 and KAP1 differentially regulate the choice of inactive versus active X chromosomes

Abstract

The onset of random X chromosome inactivation in mouse requires the switch from a symmetric to an asymmetric state, where the identities of the future inactive and active X chromosomes are assigned. This process is known as X chromosome choice. Here, we show that RIF1 and KAP1 are two fundamental factors for the definition of this transcriptional asymmetry. We found that at the onset of differentiation of mouse embryonic stem cells (mESCs), biallelic up-regulation of the long non-coding RNA Tsix weakens the symmetric association of RIF1 with the Xist promoter. The Xist allele maintaining the association with RIF1 goes on to upregulate Xist RNA expression in a RIF1-dependent manner. Conversely, the promoter that loses RIF1 gains binding of KAP1, and KAP1 is required for the increase in Tsix levels preceding the choice. We propose that the mutual exclusion of Tsix and RIF1, and of RIF1 and KAP1, at the Xist promoters establish a self-sustaining loop that transforms an initially stochastic event into a stably inherited asymmetric X-chromosome state.

Mechanisms of Choice in X-Chromosome Inactivation

“…A very similar role has been proposed for YY1 [82], and KAP1 binds to the allele with no Xist expression, due to the absence of RIF1 [81]. The asymmetric binding of RIF1 (and KAP1) seems to be downstream of fluctuations of antisense transcription across the Xist locus [81], while YY1 binding is due to differential DNA methylation of Xist alleles at the onset of XCI [82]. This has also been reported downstream of antisense transcription across the Xist locus [83][84][85].…”

“…Certain trans-acting factors have been proposed to bind asymmetrically to the Xist promoter: YY1, RIF1 and KAP1. At the onset of XCI, RIF1 is associated with the X chromosome upregulating Xist and is critical for upregulation [81]. A very similar role has been proposed for YY1 [82], and KAP1 binds to the allele with no Xist expression, due to the absence of RIF1 [81].…”

“…At the onset of XCI, RIF1 is associated with the X chromosome upregulating Xist and is critical for upregulation [81]. A very similar role has been proposed for YY1 [82], and KAP1 binds to the allele with no Xist expression, due to the absence of RIF1 [81]. The asymmetric binding of RIF1 (and KAP1) seems to be downstream of fluctuations of antisense transcription across the Xist locus [81], while YY1 binding is due to differential DNA methylation of Xist alleles at the onset of XCI [82].…”

Distal and proximal cis-regulatory elements sense X chromosome dosage and developmental state at the Xist locus

“…At the same time when the Xist promoter is activated on the future Xi, we observe active repression at the Xa through deposition of H3K9me3. This might be mediated by TRIM28/KAP1, which has been reported to bind the region on the Xa (Enervald et al, 2021) and recruits H3K9-specific histone methyl-transferases (Ecco et al, 2017). How KAP1 is targeted to the region however remains an open question.…”

RIF1 regulates replication origin activity and early replication timing in B cells

“…We also performed Repli-seq in a more physiological system, namely, primary activated splenic B cells from Rif1 +/+ , RIF1 heterozygous (Rif1 −/+ ) and Rif1 −/− mice 33 (Fig. S1C).…”