Evidence that homologous X-chromosome pairing requires transcription and Ctcf protein

Abstract

X-chromosome inactivation (XCI) ensures the equality of X-chromosome dosages in male and female mammals by silencing one X in the female. To achieve the mutually exclusive designation of active X (Xa) and inactive X (Xi), the process necessitates that two Xs communicate in trans through homologous pairing. Pairing depends on a 15-kb region within the genes Tsix and Xite. Here, we dissect molecular requirements and find that pairing can be recapitulated by 1- to 2-kb subfragments of Tsix or Xite with little sequence similarity. However, a common denominator among them is the presence of the protein Ctcf, a chromatin insulator that we find to be essential for pairing. By contrast, the Ctcf-interacting partner, Yy1 (ref. 8), is not required. Pairing also depends on transcription. Transcriptional inhibition prevents new pair formation but does not perturb existing pairs. The kinetics suggest a pairing half-life of <1 h. We propose that pairing requires Ctcf binding and co-transcriptional activity of Tsix and Xite.

Single molecule detection of direct, homologous, DNA/DNA pairing

“…Recombinational double-strand break repair and programmed homologous recombination during meiosis all involve complex series of biochemical reactions in which single-stranded DNA (ssDNA) plays a prominent role. There also exist homologous pairing reactions that seem to involve interactions between chromosomal regions whose DNAs are chemically intact double-stranded DNA (dsDNA) (1)(2)(3)(4)(5)(6)(7)(8)(9)(10)(11)(12)(13)(14)(15). In some instances, whole chromosomes pair via multiple interactions all along their lengths (1)(2)(3)(4)(5)(6)(7)(8)(9) or via any region present in duplicate copies (10).…”

“…In some instances, whole chromosomes pair via multiple interactions all along their lengths (1)(2)(3)(4)(5)(6)(7)(8)(9) or via any region present in duplicate copies (10). In other cases, pairing occurs preferentially or exclusively in particular localized regions (''pairing sites''), which tend to involve repeated sequences, specific proteins (for establishment and/or maintenance of pairing) and/or heterochromatic regions (characterized by a paucity of genes and a less ''open'' chromatin structure) (1,(11)(12)(13)(14)(15).…”

“…In some instances, whole chromosomes pair via multiple interactions all along their lengths (1-9) or via any region present in duplicate copies (10). In other cases, pairing occurs preferentially or exclusively in particular localized regions (''pairing sites''), which tend to involve repeated sequences, specific proteins (for establishment and/or maintenance of pairing) and/or heterochromatic regions (characterized by a paucity of genes and a less ''open'' chromatin structure) (1,(11)(12)(13)(14)(15).In contrast to recombination-related processes that are known to involve protein-mediated Watson-Crick basepairing interactions between a ssDNA and a ssDNA or dsDNA partner, the fundamental basis for ''recombination-independent'' pairing remains mysterious. The most obvious possibility is direct DNA/DNA interactions.…”

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

“…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. 41 As a boundary element, cohesin-CTCF could establish the unique chromatin structure of the XIST gene, which is in a euchromatic state on the principally heterochromatic Xi.…”

Mechanistic concepts in X inactivation underlying dosage compensation in mammals

“…The mechanism of Xic-Xic pairing is poorly understood and it remains unclear how two X chromosomes move and position themselves toward each other in the short developmental interval before Xist upregulation (Figure 2a). Recently, CCCTC-binding factor (CTCF) has been suggested as a crucial factor for pairing (Xu et al, 2007). As CTCF is also expressed before and after…”