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Analysis of the DNA topology upon multiple histone H1 variants depletion by Hi-C experiments

ABSTRACT: The combined depletion of histones H1.2 and H1.4 in T47D-MTVL human breast cancer cells has a strong deleterious effect, deregulates gene expression, promotes the appearance of genome-wide accessibility sites, and triggers the interferon response. We have further analyzed the consequences of multiple H1 variants depletion at the topological level by performing Hi-C experiments on T47D-MTVL cancer cells expressing a Dox-inducible shRNA against multiple H1 variants.

ORGANISM(S): Homo sapiens

PROVIDER: GSE172618 | GEO | 2022/03/23

REPOSITORIES: GEO

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Genome-wide profiling of histone H1 variants and H3K9me3 in T47D-MTVL cells upon multiple H1 depletion

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Project description:Human somatic cells may contain up to seven members of the histone H1 family contributing to chromatin compaction and regulation of nuclear processes, apparently with certain subtype specificities. Previous studies in T47D breast cancer cells determined that H1 variants are distributed in a variant-specific manner throughout the genome, although only H1.2 and H1X endogenous variants were mapped. Now, we performed ChIP-seq of five endogenous H1 variants (H1.0, H1.2, H1.4, H1.5, H1X) in T47D cells.

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Project description:Gene expression of T47D-MTVL human breast cancer cells expressing Dox-inducible shRNAs against histone H1.4 (sh120) or multiple H1 variants (sh225), or overexpressing WT or K26A mutant HA-tagged H1.4. T47D-MTVL, breast cancer cell line carrying one stably integrated copy of luciferase reporter gene driven by the MMTV promoter, is stably infected with an inducible system for the expression of shRNAs.Cells stably express RedFP and KRAB repressor fused to Tet regulator.Upon Dox treatment, cells express RedFP and the cloned shRNA. Stable breast cancer-derived cell lines expressing an shRNA against one of each of the histone H1 isoforms in response to doxycycline (Dox) were grown for six days in the presence or absence of Dox, in duplicate, and RNA extracted for microarray hybridization. Cell lines used: inducible shRNA against H1.4 or multiple H1 variants, and random shRNA-expression vector. Stable breast cancer-derived cell lines expressing the histone H1.4 isoform, WT or K26A, HA-tagged, in duplicate, and RNA extracted for microarray hybridization. Cell lines used: overexpressing H1.4 WT or K26A mutant.

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Project description:H1.2 (and control) knock-down cell lines were established from T47D-MTVL cells: Human breast cancer cell line T47D modified to contain one stably integrated copy of Luciferase reporter gene driven by the Mouse Mammary Tumor Virus promoter, named T47D-MTVL (Truss, M., J. Bartsch, A. Schelbert, R. J. Hache, and M. Beato. 1995). Initially, a cell line expressing the Dox-responsive KRAB repressor and RedFP (ptTR-KRAB-Red) was generated. Then, this cell line was infected with viruses for expression of the different H1 variants shRNAs (pLVTHM). The inducible knocked-down cell lines were sorted in a FACSvantageSE (Becton Dickinson) for RedFP-positive and GFP-positive fluorescence after 3 days of Dox treatment. Then cells were amplified in the absence of Dox until an experiment was performed. Plasmids for the lentivirus vector-mediated drug-inducible RNA interference system (pLVTHM, ptTR-KRAB-Red, pCMC-R8.91 and pMD.G) were provided by D. Trono (University of Geneva) (Wiznerowicz M & Trono D (2003) Conditional suppression of cellular genes: lentivirus vector-mediated drug-inducible RNA interference. J Virol 77: 8957-8961). Keywords: Gene expression - T47D-MTVL-control sh/sh H1.2 +/- Dox induction

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Project description:Genome-wide profiling of histone H1 variants and H3K9me3 in T47D-MTVL cells upon multiple H1 depletion

| PRJNA656518 | ENA