Evidence that homologous X-chromosome pairing requires transcription and Ctcf protein.
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.
Xu
Na
N
Howard Hughes Medical Institute Department of Molecular Biology, Massachusetts General Hospital Department of Genetics, Harvard Medical School Boston, Massachusetts 02114, USA.
Donohoe
Mary E
ME
Silva
Susana S
SS
Lee
Jeannie T
JT
D000818:Animals; D001483:Base Sequence; D001755:Blastocyst / Q000166:cytology; D000076246:CCCTC-Binding Factor; D002454:Cell Differentiation; D033481:Chromosomes, Mammalian / Q000235:genetics*; D004247:DNA / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000037:antagonists & inhibitors / Q000235:genetics* / Q000378:metabolism; D004622:Embryo, Mammalian / Q000166:cytology / Q000378:metabolism; D053595:Embryonic Stem Cells / Q000166:cytology; D005260:Female; D018628:Gene Dosage; D006801:Humans; D017404:In Situ Hybridization, Fluorescence; D008297:Male; D051379:Mice; D008822:Mice, Transgenic; D008969:Molecular Sequence Data; D011485:Protein Binding; D062085:RNA, Long Noncoding; D034741:RNA, Small Interfering / Q000494:pharmacology; D022661:RNA, Untranslated / Q000235:genetics / Q000378:metabolism; D012097:Repressor Proteins / Q000037:antagonists & inhibitors / Q000235:genetics* / Q000378:metabolism; D015534:Trans-Activators / Q000378:metabolism; D014158:Transcription, Genetic; D014960:X Chromosome / Q000235:genetics*; D049951:X Chromosome Inactivation / Q000502:physiology*; D050986:YY1 Transcription Factor / Q000235:genetics / Q000378:metabolism; D016335:Zinc Fingers
D000076246:CCCTC-Binding Factor; C000619697:CTCF protein, human; C000619698:Ctcf protein, mouse; D004268:DNA-Binding Proteins; D062085:RNA, Long Noncoding; D034741:RNA, Small Interfering; D022661:RNA, Untranslated; D012097:Repressor Proteins; D015534:Trans-Activators; C465658:Tsix transcript, mouse; D050986:YY1 Transcription Factor; C494699:Yy1 protein, mouse; D004247:DNA