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ABSTRACT: RNA-seq of SPEN gene deletion in primary mouse hepatocytes

PROVIDER: PRJNA887130 | ENA |

REPOSITORIES: ENA

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			Action	DRS
	SRR21808323_1.fastq.gz	Fastqsanger.gz
	SRR21808323_2.fastq.gz	Fastqsanger.gz
	SRR21808324_1.fastq.gz	Fastqsanger.gz
	SRR21808324_2.fastq.gz	Fastqsanger.gz
	SRR21808325_1.fastq.gz	Fastqsanger.gz

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SPEN is a molecular integrator that bridges Xist with transcriptional and epigenetic control of X-chromosome inactivation

Project description:Xist represents a paradigm for long non-coding RNA function in epigenetic regulation, although how it mediates X-chromosome inactivation (XCI) remains largely unexplained. Multiple Xist-RNA binding proteins have recently been identified, including SPEN/SHARP, whose knockdown has been associated with deficient XCI at multiple loci. Here we demonstrate that SPEN is a key orchestrator of XCI in vivo and unravel its mechanism of action. We show that SPEN is essential for initiating gene silencing on the X chromosome in preimplantation mouse embryos and embryonic stem cells. On the other hand, SPEN is dispensable for maintenance of XCI in neural progenitor cells, although it significantly dampens expression of genes that escape from XCI. During initiation of XCI, we show by live-cell imaging and CUT&RUN approaches that SPEN is immediately recruited to the X chromosome upon Xist up-regulation, where it is targeted to enhancers and promoters of actively transcribed genes. SPEN rapidly disengages from chromatin once silencing is accomplished, implying a need for active transcription to tether it to chromatin. We define SPEN’s SPOC (SPEN paralog and ortholog C-terminal) domain as a major effector of SPEN’s gene silencing function, and show that artificial tethering of SPOC to Xist RNA is sufficient to mediate X-linked gene silencing. We identify SPOC’s protein partners which include NCOR/SMRT, the m6A RNA methylation machinery, the NuRD complex, RNA polymerase II and factors involved in regulation of transcription initiation and elongation. We propose that SPEN acts as a molecular integrator for initiation of XCI, bridging Xist RNA with the transcription machinery as well as nucleosome remodelers and histone deacetylases, at active enhancers and promoters.

2021-04-19 | PXD015699 | Pride

H2AK119ub1 native ChIP-seq upon Xist indution

Project description:Native ChIP-seq for H2AK119ub1 upon Xist induction in iXist-ChrX mouse embryonic stem cells. Comparison of WT with Spen RRM deletion and SPOC domain mutants for two timepoints of Xist induction, 3h and 24h.

2021-07-01 | GSE154549 | GEO

Study of SPEN knockdown in MCF10A cells

Project description:In this experiment, we knocked down the expression of Split Ends (SPEN), a gene that we found was involved in the regulation of primary cilia formation, in MCF10A cells, to evaluate its effects on transcription. Split Ends (SPEN) is a transcriptional coregulator that have formerly identified as a tumour suppressor gene in ER-positive breast cancers. We expect the knockdown of SPEN in MCF10A cells to be accompanied with the down-regulation of genes involved in ciliogenesis as we observed that SPEN knockdown decreases primary cilia formation in those cells. We chose MCF10A cells because they harbour lots of primary cilia and therefore represent a good model to study the effect of SPEN on primary cilia formation.

2017-08-15 | E-MTAB-4974 | biostudies-arrayexpress

The effect of TRIB3 overexpression on mouse hepatocytes

Project description:To investigate the effect of TRIB3 overexpression on regulation of lipid metabolism in hepatocytes, we isolated mouse primary hepatocytes from AAV-GFP or AAV-Trib3 mice. We then performed gene expression profiling analysis using data obtained from RNA-seq of two groups of mouse primary hepatocytes from AAV-GFP or AAV-Trib3 mice.

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In vitro differentiation of hepatoblasts into hepatocytes

Project description:We differentiated primary mouse E14.5 Dlk1+ hepatoblasts in vitro into hepatocyte-like cells that recapitulated morphological features of hepatocytes. To assess the level of differentiation, we performed RNA-seq analysis and compared gene expression profiles of hepatobalsts, in vitro differentiated hepatocytes and mature hepatocytes isolated from adult mouse livers as a control.

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Project description:mouse primary hepatocytes transcriptome

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Ability of SPEN to bind to chromatin in testicular cells

Project description:Since SPEN co-localizes with transcriptionally active chromatin and it is considered as a nuclear matrix platform that organizes and integrates transcriptional responses, we aimed to find out how it is involved in gene expression regulation in spermatogenic cells. We assumed that SPEN could be directly involved in down-regulation of gene expression during recovery from heat shock, when general repression of the transcription was observed. Nascent SPEN (~400 kDa) is processed to several fragments. We tested antibodies to N-terminal, internal, or C-terminal SPEN sequences in ChIP experiments on mouse testicular cells either untreated or heat shocked. Only antibodies against N-terminal or internal SPEN epitopes were effective while C-terminal SPEN did not bind to chromatin. However sequencing of N-terminal or internal SPEN ChIP samples did not reveal any strong SPEN binding to the chromatin and the enrichment was seen primarily in genomic structural variants. This suggests that SPEN could be loosely associated with DNA via other proteins.

2017-10-20 | GSE99856 | GEO

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