EXPERIMENT_TITLE: GSM6754655: NL_25_NPC_HiC; Homo sapiens; Hi-C
EXPERIMENT_DESIGN_LIBRARY_DESCRIPTOR_LIBRARY_CONSTRUCTION_PROTOC: We prepared Hi-C libraries using the Arima Genomics Hi-C kit (Arima Genomics, A510008) according to the manufacturer's protocol. Briefly, we enzymatically digested genomic DNA within nuclei of crosslinked cell pellets, and created biotinylated ligation junctions between the digested ends at proximity. Then we extracted DNAs and sheared to an average size of ~400 bp using a Covaris S220 sonicator at 140 W peak incident power, 10 % duty factor, and 200 cycles per burst for 55 seconds. We further size selected the sheared DNA to 200-600 bp using AgenCourt Ampure XP beads (Beckman Coulter, A63881) according to manufacturer's protocols. Biotin-tagged ligation junctions via pulldown using streptavidin breads from the Arima Hi-C kit (Arima Genomics, A510008) according to manufacturer's protocol. Streptavidin beads containing Hi-C libraries were stored at -20 °C for no more than 3 days before Illumina sequencing library preparation was performed.  Hi-C libraries were prepped for sequencing by first washing adaptor-ligated Hi-C libraries on streptavidin beads twice in 150 uL of wash buffer at 55 °C and once in 100 ml of elution buffer at room temperature using Hi-C kit (Arima Genomics, A510008). DNA was eluted from streptavidin beads by boiling at 98 °C for 10 min in 15 uL elution buffer. Subsequently the libraries were amplified using NEBNext Ultra II DNA Library Prep Kit for Illumina (NEB, E7645S) with 8 PCR cycles according to the manufacturer's protocol.
STUDY_DESCRIPTOR_STUDY_ABSTRACT: Short tandem repeat (STR) instability causes transcriptional silencing in several repeat expansion disorders. In fragile X syndrome (FXS), mutation-length expansion of a CGG STR represses FMR1 via local DNA methylation. Here, we find Megabase-scale H3K9me3 domains on autosomes and encompassing FMR1 on the X-chromosome in induced pluripotent stem cell (iPSC)-derived neural progenitors, B-cells, and fibroblasts with mutation-length CGG expansion. H3K9me3 domains connect via inter-chromosomal interactions and demarcate severe misfolding of TADs and loops. They harbor long synaptic genes replicating at the end of S phase, replication stress-induced double strand breaks, and STRs prone to stepwise somatic instability. CRISPR engineering of the full-mutation CGG to premutation-length reverses H3K9me3 domains on the X-chromosome and multiple autosomes, refolds TADs, and restores expression. H3K9me3 domains also arise in a subset of normal-length iPSCs with increased STR instability burden. Our results reveal Mb-scale heterochromatinization and trans interactions among chromosomes susceptible to repeat genetic instability. Overall design: Hi-C, H3K9me3 ChIP-seq, CTCF ChIP-seq, RNA-seq, H3K9me3 CUT&RUN, and short and long read genome sequencing were used in multiple cell types to interrogate epigenetic changes in cell lines with normal, premutation, and disease level repeat expansions in FMR1.
SAMPLE_TITLE: NL_25_NPC_HiC
SAMPLE_SAMPLE_ATTRIBUTES_SAMPLE_ATTRIBUTE: [{"TAG": "source_name", "VALUE": "iPSC derived NPC"}, {"TAG": "cell type", "VALUE": "iPSC derived NPC"}]
Pool_Member_@attributes_sample_title: NL_25_NPC_HiC
RUN_SET_RUN_Pool_Member_@attributes_sample_title: NL_25_NPC_HiC
RUN_SET_RUN_SRAFiles_SRAFile: [{"Alternatives": null, "@attributes": {"cluster": "public", "filename": "hic_158_R1_001.fastq.gz", "size": "28228584644", "date": "2022-11-24 02:04:17", "md5": "0ecdd8884cd170c48a2b611c49ee4821", "version": "1", "semantic_name": "fastq", "supertype": "Original", "sratoolkit": "0"}}, {"Alternatives": null, "@attributes": {"cluster": "public", "filename": "hic_158_R2_001.fastq.gz", "size": "28959748322", "date": "2022-11-24 02:06:17", "md5": "b51f97c93cd61dc2646c2e79304dfc8b", "version": "1", "semantic_name": "fastq", "supertype": "Original", "sratoolkit": "0"}}, {"Alternatives": null, "@attributes": {"cluster": "public", "filename": "SRR22398603", "url": "https://sra-pub-run-odp.s3.amazonaws.com/sra/SRR22398603/SRR22398603", "size": "31330506810", "date": "2022-11-24 02:17:31", "md5": "646361261e6e87c19b3aa9689cba37e8", "version": "1", "semantic_name": "SRA Normalized", "supertype": "Primary ETL", "sratoolkit": "1"}}, {"Alternatives": [null, null, null], "@attributes": {"cluster": "public", "filename": "SRR22398603.lite", "url": "https://sra-downloadb.be-md.ncbi.nlm.nih.gov/sos2/sra-pub-zq-20/SRR022/22398/SRR22398603/SRR22398603.lite.1", "size": "24275960586", "date": "2022-11-29 03:25:28", "md5": "3d69ef75bb4bab4712da9278a7c545f8", "version": "1", "semantic_name": "SRA Lite", "supertype": "Primary ETL", "sratoolkit": "1"}}]