Abstract

We carried out padlock capture, a high-resolution RNA allelotyping method, to study X chromosome inactivation (XCI). We examined the gene reactivation pattern along the inactive X (Xi), after Xist (X-inactive specific transcript), a prototype long non-coding RNA essential for establishing X chromosome inactivation (XCI) in early embryos, is conditionally deleted from Xi in somatic cells (Xi∆Xist). We also monitored the behaviors of X-linked non-coding transcripts before and after XCI. In each mutant cell line, gene reactivation occurs to ~6% genes along Xi∆Xist in a recognizable pattern. Genes with upstream regions enriched for SINEs are prone to be reactivated. SINE is a class of retrotransposon transcribed by RNA polymerase III (Pol III). Intriguingly, a significant fraction of Pol III transcription from non-coding regions is not subjected to Xist-mediated transcriptional silencing. Pol III inhibition affects gene reactivation status along Xi∆Xist, alters chromatin configuration and interferes with the establishment XCI during in vitro differentiation of ES cells. These results suggest that Pol III transcription is involved in chromatin structure re-organization during the onset of XCI and functions as a general mechanism regulating chromatin configuration in mammalian cells.

Highlights

  • Allele- expressed from Xi in somatic cells ? Are non-coding transcripts subjected to Xist-mediated transcriptional silencing ? If so, are they reactivated along Xi∆Xist ? Again, a high-resolution RNA allelotyping method is required to answer these questions

  • The transcription across the non-coding SNP (12344545), which was repressed by the polymerase III (Pol III) inhibitor (Fig. 4B), is located upstream of a gene Med[14], which was reactivated in HR7 cells (Fig. 4F)

  • Pol III inhibition generated more chromatin interactions especially around the boundaries of the two topologically associating domains. These results show that Pol III inhibition affects chromatin configuration

Read more

Summary

Introduction

Allele- expressed from Xi in somatic cells ? Are non-coding transcripts subjected to Xist-mediated transcriptional silencing ? If so, are they reactivated along Xi∆Xist ? Again, a high-resolution RNA allelotyping method is required to answer these questions. Completely different from X-linked genes, which are exclusively expressed from Xcast in the control fibroblasts (Xi129XaCast), the allelotype of non-coding transcription showed a high degree of allele bias against either Xcast or X129 (Fig. 3A).

Results
Conclusion
Full Text
Published version (Free)

Talk to us

Join us for a 30 min session where you can share your feedback and ask us any queries you have

Schedule a call