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Time-lapse super-resolution imaging reveals key principles for cis-limited Xist RNA localization [ChrRNA-seq]

ABSTRACT: Time-lapse super-resolution imaging reveals key principles for cis-limited Xist RNA localization [ChrRNA-seq]

PROVIDER: PRJNA646963 | ENA |

REPOSITORIES: ENA

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			Action	DRS
	SRR12248800_1.fastq.gz	Fastqsanger.gz
	SRR12248800_2.fastq.gz	Fastqsanger.gz
	SRR12248801_1.fastq.gz	Fastqsanger.gz
	SRR12248801_2.fastq.gz	Fastqsanger.gz
	SRR12248802_1.fastq.gz	Fastqsanger.gz

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Time-lapse super-resolution imaging reveals key principles for cis-limited Xist RNA localization [ChrRNA-seq]

Project description:Xist RNA, which directs the process of X chromosome inactivation in mammals, localizes in cis over the chromosome from which it is transcribed, recruiting chromatin modifiers to silence underlying genes. To analyze cis-limited Xist RNA localization we have developed time-resolved super-resolution imaging of individual Xist RNA molecules in single cells. Using this approach, we quantify fundamental parameters underpinning Xist RNA dynamics, demonstrating a feedback mechanism linking Xist RNA synthesis and degradation, and an unusual coupling behavior that physically links temporally separated Xist RNA molecules. Additionally, we show that the protein SPEN, a key factor for Xist-mediated gene-silencing, has a separable function in Xist RNA localization, stability and in coupling behavior. Our results provide important insights towards understanding the unique and unusual behavior of Xist RNA.

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Time-lapse super-resolution imaging reveals key principles for cis-limited Xist RNA localization (RNA-SPLIT)

Project description:Time-lapse super-resolution imaging reveals key principles for cis-limited Xist RNA localization (RNA-SPLIT)

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Time-lapse super-resolution imaging reveals key principles for cis-limited Xist RNA localization (RNA-SPLIT)

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Project description:X chromosome inactivation (XCI) is mediated by the non-coding RNA Xist which directs chromatin modification and gene silencing in cis. The RNA binding protein SPEN and associated corepressors have a central role in Xist-mediated gene silencing. Other silencing factors, notably the Polycomb system, have been reported to function downstream of SPEN. Making use of a SPEN separation-of-function mutation we show that SPEN and Polycomb pathways in fact function in parallel to establish gene silencing. Additionally, we find that differentiation-dependent recruitment of the chromosomal protein SmcHD1 is required for silencing many X-linked genes.

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Project description:Purpose: Compare the transcriptome of homogeneous XIST+ and XIST- hES cell populations. Methods: We isolated homogeneous XIST+ and XIST- cell populations. The XIST+ cells correspond to cells with a XIST cloud and one ATRX pinpoint. The XIST- cells correspond to cells with no XIST cloud and one ATRX pinpoint. Results: We took advantage of the clonal pattern of X-chromosome inactivation in H9 cells and analyzed the data in an allelic manner. By comparing the RNA-Seq data with known H9 SNPs, we identified genomic positions which were relaxed from XCI in XIST- cells compared to XIST+ cells. Conclusions: Genic as well as unannotated transcripts are massively relaxed from XCI in H9 cells when XIST expression is lost, however, this reactivation is only partial and a large region around the centromere is protected from relaxation of silencing. Total RNA (rRNA depleted) profiles of XIST+ and XIST- human embryonic stem cells

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