EXPERIMENT_TITLE: GSM5044163: HiC_MCF7TR_3D_rep2; Homo sapiens; Hi-C
EXPERIMENT_DESIGN_LIBRARY_DESCRIPTOR_LIBRARY_CONSTRUCTION_PROTOC: Hi-C was performed as previously described in Zhou. Y et. In brief, spheroids were crosslinked with 1% formaldehyde and lysed with cold lysis buffer to collect nuclei. The pelleted nuclei were digested with 200 units of HindIII (NEB, R3104L) at 37 °C for overnight. The HindIII digested fragment overhangs were filled with biotin-labelled dATP (Life Technologies, 19524-016) in a Klenow end-filling reaction. Four hundred units of T4 DNA Ligase (NEB, M0202) were added for ligation and samples were incubated for 4 h at room temperature with slow rotation. The ligation products were purified, and the chromatin was sheared to a size of 300-500bp using Covaris sonicator (Covaris Woburn, MA). Dynabeads MyOne Streptavidin T1 beads (Life technologies, 65601) were used to pull down the Biotin-labelled DNA. The end repair, dA tailing was performed and ligated with Illumina TruSeq adapters to form the final Hi-C ligation products. Each Hi-C library was amplified with 12 cycles of PCR using Illumina primers. The Hi-C library was purified and then sequenced with Illumina HiSeq3000.
STUDY_DESCRIPTOR_STUDY_ABSTRACT: Organoids or spheroids have emerged as a physiologically relevant in vitro preclinical model to study patient-specific diseases. A recent study used spheroids of MCF10 cells to model breast cancer progression and identified targetable alterations more similar to those in vivo. Thus, it is practical and essential to explore and characterize the spheroids of the commonly used human breast cancer (BC) cells. This study conducted Hi-C analyses in three-dimensional (3D) spheroids of MCF10A, MCF7 and MCF7TR cells and compared TADs and looping genes with those in 2D monolayers. We found that chromatin domains and looping genes' strength have drastically changed during the 3D culture growth, although we identified very similar numbers of TADs and looping genes. We further identified novel 3D growth-specific looping genes within Hippo relevant pathways, of which two genes showed potential prognostic values in measuring the outcome of the endocrine treatment. We finally confirmed a few selected Hippo relevant pathways genes with enhanced looping in breast cancer patient tissues' organoid. Hence, our work has provided significant insights into our understanding of 3D-growth-specific chromatin architecture in tamoxifen-resistant breast cancer. Overall design: Hi-C experiment of 3 cell types in 3D spheroids, 2 biological replicates for each cell type.
SAMPLE_TITLE: HiC_MCF7TR_3D_rep2
SAMPLE_SAMPLE_ATTRIBUTES_SAMPLE_ATTRIBUTE: [{"TAG": "source_name", "VALUE": "Breast cancer cells"}, {"TAG": "cell line", "VALUE": "MCF7TR"}, {"TAG": "cell type", "VALUE": "tamoxifen-resistant breast cancer cell line"}, {"TAG": "culture type", "VALUE": "3D spheroid"}, {"TAG": "restriction enzyme", "VALUE": "HindIII"}]
Pool_Member_@attributes_sample_title: HiC_MCF7TR_3D_rep2
RUN_SET_RUN_Pool_Member_@attributes_sample_title: HiC_MCF7TR_3D_rep2
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