EXPERIMENT_TITLE: GSM3911979: hct116_Rad21DegronPolIIhet_HiC_auxinamanitin_br1_tr1; Homo sapiens; Hi-C
EXPERIMENT_DESIGN_LIBRARY_DESCRIPTOR_LIBRARY_CONSTRUCTION_PROTOC: Promoter Capture  HiC-Seq:  we used the same protocol at Montefiori et al., (Elife, 2018 Jul 10; 7)                                                                                                                                                                                                                                                                          BS-seq: Genomic DNA was isolated from 3×106 cells using Qiagen DNeasy blood and tissue kit (Cat #: 69506). The bisulfite conversion was performed using the EZ DNA Methylation-Gold™ Kit (Cat #: D5005) from Zymo Research. Whole-genome bisulfite sequencing libraries were prepared from 50 ng converted DNA using Illumina TruSeq® DNA Methylation Kit (Cat #: EGMK91324), which was performed following the manufacturer's protocol. Promoter Capture HiC-seq: DNA was isolated and prepared for paired-end Illumina sequencing following the genomic DNA protocol. 100 cycles of sequencing data were acquired on the HiSeq 3000 (Illumina).                                                                                             BS-seq: standard RNA-Seq   library   preparation   was performed following  the genomic DNA protocol with 20% PhiX. 50 cycles of sequencing data were acquired on HiSeq 3000 (Illumina)
STUDY_DESCRIPTOR_STUDY_ABSTRACT: Our mechanistic understanding of Mediator derives largely from studies of the 25-subunit yeast complex. Here we combine CRISPR-Cas9 genetic screens, degron assays, in situ Hi-C, and cryo-EM to dissect the function and structure of the 33-subunit mammalian Mediator (mMED). Deletion analyses in B, T and ES cells reveal that depletion of the entire complex blocks PolII recruitment genome-wide, while loss of non-essential subunits, including the Tail module, primarily affects promoters linked to multiple enhancers. Contrary to current models, we find that mMED is not required to tether regulatory DNA, a topological activity controlled predominantly by architectural proteins. Structurally, we show that alterations in the Tail module, particularly at the core-Tail interphase, effect crucial mMED-PolII contacts, providing a rationale for how TFs stabilize the mMED-PolII holoenzyme and promote gene expression. Our studies therefore reveal key insights into how Mediator functionally bridge promoters and enhancers to regulate transcription initiation in higher eukaryotes. Overall design: ChIP-seq, ATACseq, Hi-C,PromterCaputre Hi-C, CRISPR screen and RNAseq from mouse ESC, CD4+ T cell or CH12 cell line and/or human HCT-116 cell line The processed data files generated from multiple samples are indicated in the corresponding sample description field, and linked to the Series records or to the corresponding 'r1' sample records.
SAMPLE_TITLE: hct116_Rad21DegronPolIIhet_HiC_auxinamanitin_br1_tr1
SAMPLE_SAMPLE_ATTRIBUTES_SAMPLE_ATTRIBUTE: [{"TAG": "source_name", "VALUE": "HCT-116 cell line"}, {"TAG": "genotype", "VALUE": "Rad21Degron_PolII+/-"}, {"TAG": "cell type", "VALUE": "colon carcinoma cell line"}]
Pool_Member_@attributes_sample_title: hct116_Rad21DegronPolIIhet_HiC_auxinamanitin_br1_tr1
RUN_SET_RUN_Pool_Member_@attributes_sample_title: hct116_Rad21DegronPolIIhet_HiC_auxinamanitin_br1_tr1
RUN_SET_RUN_SRAFiles_SRAFile: [{"Alternatives": [null, null, null], "@attributes": {"cluster": "public", "filename": "SRR9621905.sralite", "url": "https://sra-downloadb.be-md.ncbi.nlm.nih.gov/sos5/sra-pub-zq-14/SRR009/621/SRR9621905.sralite.1", "size": "15409662444", "date": "2020-06-23 06:52:18", "md5": "362faa0f900ee6ed40bd18e5aaec5e5b", "version": "1", "semantic_name": "SRA Lite", "supertype": "Primary ETL", "sratoolkit": "1"}}, {"Alternatives": null, "@attributes": {"cluster": "public", "filename": "SRR9621905", "url": "https://sra-pub-run-odp.s3.amazonaws.com/sra/SRR9621905/SRR9621905", "size": "26373387696", "date": "2019-07-02 20:12:54", "md5": "c051f7802184f50b1a31b32d648f1506", "version": "1", "semantic_name": "SRA Normalized", "supertype": "Primary ETL", "sratoolkit": "1"}}, {"Alternatives": null, "@attributes": {"cluster": "public", "filename": "hct116_Rad21DegronPolIIhet_HiC_auxinamanitin_br1_tr1_R2.fastq.gz", "size": "20997245073", "date": "2019-07-02 19:49:00", "md5": "cfb130fc8d670d1f9842a8e1bcc031c3", "version": "1", "semantic_name": "fastq", "supertype": "Original", "sratoolkit": "0"}}, {"Alternatives": null, "@attributes": {"cluster": "public", "filename": "hct116_Rad21DegronPolIIhet_HiC_auxinamanitin_br1_tr1_R1.fastq.gz", "size": "18181039354", "date": "2019-07-02 19:36:28", "md5": "b5fa240c6ac2aefe147f684c9acfd4c1", "version": "1", "semantic_name": "fastq", "supertype": "Original", "sratoolkit": "0"}}]