EXPERIMENT_TITLE: GSM4287006: HiC_T47DTamR+sapitinib_rep1; Homo sapiens; Hi-C
EXPERIMENT_DESIGN_LIBRARY_DESCRIPTOR_LIBRARY_CONSTRUCTION_PROTOC: In situ Hi-C experiments were performed as described previously (Rao et al, 2015; Zhou et al, 2020). The breast cancer cells were crosslinked with 1% formaldehyde and lysed with ice cold lysis buffer (10mM Tris-Hcl pH 8.0, 10mM NaCl, 0.2 % NP-40, 1mM DTT) to collect nuclei. The pelleted nuclei were digested with 200 units of HINDIII (NEB, R3104L) at 37°C overnight. The restriction 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) was 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: Recent studies suggested that crosstalk between ERa and EGFR/HER2 pathways plays a critical role in mediating endocrine therapy resistance. Several targeting EGFR/HER2 signaling inhibitors including FDA-approved lapatinib and gefitinib as well as a novel dual tyrosine kinase inhibitor (TKI) sapitnib showed greater inhibitory efficacies. However, how a 3D chromatin landscape of the response to the inhibition to EGFR/HER2 pathway remains to be elucidated. In this study, we conducted in situ Hi-C and RNA-seq in two ERa+ breast cancer cell systems, tamoxifen-sensitive MCF7 and T47D and tamoxifen-resistant MCF7TR and T47DTR before and after the treatment of sapitnib. We identified differential response of topologically associated domains (TADs), looping genes and expressed genes. Interestingly, we found that many differential TADs and looping genes are reversible, indicating that EGFR/HER2 signaling may play a role in reshaping and rewiring the high order genome organization. We further examined and recapitulated the reversible looping genes in 3D spheroids of breast cancer cells. Our data provides a rich resource for further evaluating chromatin structural response to anti-EGFR/HER2 targeted therapies in endocrine-resistant breast cancer. Overall design: Hi-C experiment of 3 cell types, 2 biological replicates for each cell type;
SAMPLE_TITLE: HiC_T47DTamR+sapitinib_rep1
SAMPLE_SAMPLE_ATTRIBUTES_SAMPLE_ATTRIBUTE: [{"TAG": "source_name", "VALUE": "HiC_T47DTamR+sapitinib"}, {"TAG": "cell line background", "VALUE": "T47D"}, {"TAG": "cell type", "VALUE": "breast cancer cell line"}, {"TAG": "phenotype", "VALUE": "tamoxifen resistant"}, {"TAG": "restriction enzyme", "VALUE": "HindIII"}]
Pool_Member_@attributes_sample_title: HiC_T47DTamR+sapitinib_rep1
RUN_SET_RUN_Pool_Member_@attributes_sample_title: HiC_T47DTamR+sapitinib_rep1
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