EXPERIMENT_TITLE: GSM3330966: InsituHiC_IMR90_G_Bio1; Homo sapiens; Hi-C
EXPERIMENT_DESIGN_LIBRARY_DESCRIPTOR_LIBRARY_CONSTRUCTION_PROTOC: In situ Hi-C was performed as previously described (Rao et al., 2014, Cell) Sequencing adapters were ligated using NEBNext Ultra Ligation module (New England Biolabs, Inc) on Dynabeads, and DNA was PCR-amplified for 8-14 cycles using NEBNext Q5 Hot Start HiFi PCR master Mix and NEBNext multiplex oligos (New England Biolabs, Inc).
STUDY_DESCRIPTOR_STUDY_ABSTRACT: Cellular senescence is a state of stable cell growth arrest induced by various stimuli such as oncogene expression and telomere shortening, referred to as oncogene-induced senescence (OIS) and replicative senescence (RS), respectively.  Senescence is accompanied not only by global transcriptional alterations but also by 3D genome reorganization.  Here we demonstrate that the human condensin II complex participates in cellular senescence via gene regulation and reorganization of euchromatic A and heterochromatic B chromatin compartments.  OIS and RS are accompanied by A-to-B and B-to-A compartmental transitions, the latter of which occur more frequently and account for 15% of the human genome.  Mechanistically, condensin is enriched at A compartments, especially at gene promoters and typical/super enhancers, and implicated in the B-to-A transitions.  Genes present at B-to-A-switching regions tend to be up-regulated.  The full activation of senescence genes (SASP genes and p53 targets) requires condensin; its depletion impairs senescence markers such as the activity of senescence-associated beta-galactosidase (SA-beta-gal) and senescence-associated heterochromatic foci (SAHF).  This study describes that condensin reinforces euchromatic A compartments and is required for optimal expression of senescence genes, thereby contributing to the establishment and maintenance of the senescent state. Overall design: in situ HI-C with human senescence cells. ChIP-seq and RNA-seq with human senescence cells. Normal diploid IMR90 human fibroblasts were cultured in DMEM medium supplemented with 10% FBS, 100 U/ml penicillin, 100 ug/ml streptomycin, 0.15% sodium bicarbonate, 2 mM L-glutamine, 1 mM sodium pyruvate, and 1 x MEM non-essential amino acids.  Growing cells in population doublings (PD) between 26 and 32 were used for experiments.  pBABE-puro plasmids carrying H-rasV12 were used for retrovirus packaging.  H-rasV12-expressing cells were cultured in medium containing 1 µg/ml puromycin for 7 days to obtain OIS cells. For  CAP-H2 knockdown, pTRIPZ plasmids containing NCAPH2 shRNAs (#1, V3THS_326285; #2, V3THS_326286; #3, V3THS_326290, Dharmacon: CAPH2_KD1, 2, 3) and empty vector (control) were used.  Doxycycline (1ug/ml) was added to culture medium every 48 hours after lentiviral transfection for shRNA expression.  OIS cells were further infected with lentivirus encoding one of the three shRNA constructs (KD1, 2, and 3) against NCAPH2 and harvested 3 days after the lentivirus infection.
SAMPLE_TITLE: InsituHiC_IMR90_G_Bio1
SAMPLE_SAMPLE_ATTRIBUTES_SAMPLE_ATTRIBUTE: [{"TAG": "source_name", "VALUE": "IMR90 cells"}, {"TAG": "passage", "VALUE": "28-32"}, {"TAG": "cell type", "VALUE": "Lung fibroblasts"}, {"TAG": "karyotype", "VALUE": "Normal diploid"}]
Pool_Member_@attributes_sample_title: InsituHiC_IMR90_G_Bio1
RUN_SET_RUN_Pool_Member_@attributes_sample_title: InsituHiC_IMR90_G_Bio1
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