title: Sample_68_RORg_MHCII


title: MB49_POM1_2

status: Public on Sep 06 2022

type: SRA

city: Wu Han

first: Junyi

last: Hu

address: No. 1277 Liberation Avenue

department: Urology

country: China

institute: Union Hospital, Tongji Medical College, Huazhong University of Science and Technology

library_source: transcriptomic

data_processing:
Single-cell RNA sequencing data was collected on a Hiseq X Ten Illumina instrument.
Alignment using Cell Ranger (5.0.0) from 10x Genomics.
Genome_build: mm10 (GRCm38)
Supplementary_files_format_and_content: *.tar: Tar archives include barcodes.tsv.gz, features.tsv.gz, and matrix.mtx.gz files.

molecule: total RNA

organism: Mus musculus

source_name: bladder cancer

growth_protocol: MB49 Subcutaneous tumors.

extract_protocol:
After single cell suspension, CD45+ cells were sorted.
Barcoded scRNA-seq libraries were prepared using the Chromium Single Cell 3’ v3 Reagent Kit.

treatment_protocol: 5 mg/kg of the NTPDase inhibitor sodium polyoxotungstate (POM-1/CD39i, purchased from Santa Cruz Biotechnology, Inc., Dallas, Texas, USA) once per day from day 6 to day 22.

tag: tissue

value: MB49 Subcutaneous tumors

tag: strain

tag: genotype

value: wild type

tag: marker

value: CD45+ sorted

tag: treatment

value: POM-1 treated


title: Sample_67_RORg_MHCII

city: New York

first: Chrysothemis

last: Brown

state: NY

department: Immunology Program, Sloan Kettering Institute

country: USA

institute: Memorial Sloan Kettering Cancer Center

description:
RORgt+ MHCII+ from 3-week old mice
smart-seq2 protocol

data_processing:
base-calling was performed using HiSeq Control Software HD v3.4.0.38
samples were demultiplexed using bcl2FASTQ v2.20.0.422
FASTQ files from demultiplexed samples were aligned to GRCm38 (mm10) using STAR v2.7.7a with ‘--twopassMode Basic --outFilterMultimapNmax 1 --quantMode TranscriptomeSAM’
The alignments of each sample were name-sorted using Samtools v1.11
The count matrix for all cells was created by counting alignments using featureCounts (subread v2.0.1)
Low-quality cells/genes were filtered and downstream analyses (normalization, PCA, etc.) was performed using Seurat v3.2.3.
Cells were clustered using PhenoGraph and UMAP embedding was computed using Seurat v3.2.3.
Genome_build: mm10
Supplementary_files_format_and_content: the count matrix (tab-delimited table) includes raw counts for all cells (smart-seq2 samples).
Supplementary_files_format_and_content: the cell metadata (csv table) includes clusters, annotations, etc. for all cells (corresponding to rows in the count matrix).

source_name: Mouse MLN RORgt+ MHCII+

extract_protocol:
RORgt+MHCII+ cells were enriched from a pool of MLN from 3-week-old RorcVenus-CreERT2 mice. Cells were depleted of Lineage (TCRb, TCRgd, CD19, B220, NK1.1)+ cells via staining with biotinylated antibodies followed by magnetic bead negative selection. Live, Lin(CD3, TCRb, TCRgd CD19, B220, NK1.1)–CD64–Ly6C–MHCII+RORgt(Venus)+ cells were then sorted into single wells. Cells were also stained for CD90, CD11c and CD11b for acquiring index sorting information on cell surface expression. Aire+ mTECs were enriched from a pool of thymi from 3-week old mice via staining with biotinylated antibodies against CD45 followed by magnetic bead negative selection. CD45–Epcam+MHCII+Aire(GFP)+ cells were sorted into single wells. Aire+ DCs were enriched from a pool of MLN from the same 3-week old mice. Cells were depleted of Lin+ cells as described above and live, Lin(CD3, TCRb, TCRgd CD19, B220, NK1.1)–CD90–CD64–Ly6C–CD11c+MHCII+Aire(GFP)+ cells were then sorted into single wells.
Single cells were sorted into Buffer RLT (Qiagen). Cell lysates were immediately sealed and spun down before transferring to dry ice and storing at -80 ºC. RNA was purified using the Agencourt RNAClean XP beads (Beckman Coulter) at a 2.2X ratio. First-strand cDNA synthesis was achieved using Maxima H Minus Reverse Transcriptase (ThermoFisher) according to the manufacturer’s protocol using oligo dT primers, with the addition of a custom template-switch oligo in a 1mM final concentration. cDNA was amplified for 24 cycles using KAPA HiFi HotStart ReadyMix (Kapa Biosystems KK2601). After PicoGreen quantification, 0.1-0.2ng of cDNA was used to prepare libraries with the Nextera XT DNA Library Preparation Kit (Illumina) in a total volume of 6.25µL with 12 cycles of PCR. Indexed libraries were pooled by volume and cleaned by aMPure XP beads (Beckman Coulter) at a 1X ratio. Pools were sequenced on a HiSeq 4000 in a PE50 or PE100 run using the HiSeq 3000/4000 SBS Kit (Illumina).

value: mesenteric lymph nodes (MLN)

tag: age

value: three weeks

tag: cell type

value: RORgt+ MHCII+


title: THP-1 ORF RCL1 rep1

city: Montréal

first: John

middle: D

last: Rioux

state: QC

address: 5000 Bélanger St.

department: Research Center

country: Canada

institute: Montreal Heart Institute

description: THP-1 stable cell line overexpressing RCL1 gene / replicate 1

data_processing:
Illumina HiSeq2000 using HiSeq Control Software
Primary QC analysis was performed using FastQC (v0.11.5; https://www.bioinformatics.babraham.ac.uk/projects/fastqc)
Sequence filtering parameters set by Trimmomatic (v.0.36 ) Primer sequences were filtered out with provided TruSeq sequence data and a quality sliding window of Q over 20 for a 40-nucleotide window; only sequences longer than 70 nucleotides were kept in the filtered FASTQ files. Another FastQC run was performed on all filtered FASTQ files to validate Trimmomatic results.
Filtered FASTQ files were aligned to the human genome with STAR (v. 2.5.3a) using indexes built with STAR from H. sapiens GRCh Build 38 files (reference genome and GTF annotation files) provided by Illumina's iGenome repository.
Default values were selected except for the –quantMode flag that was set to TranscriptomeSAM for downstream quantification. Counts per gene for each sequenced sample were obtained using RSEM (v. 1.2.6)
Gene counts were transformed to FPKM as an approximation of gene relative abundance. Genes were filtered to keep only those with FPKM > 0.3 and count>8 in at least two samples.
Small RNAs and LOC genes without annotations were removed. Quality control of samples was performed, excluding strong outlier samples based on principal component analyses
The log-transformed FPKM were then normalized between samples with cyclicloess (normalizeCyclicloess from sva Bioconductor R package).
RNA-seq and cell culture batches effects were removed using ComBat (sva Bioconductor library v3.18.0).
Genome_build: GRCh 38
Supplementary_files_format_and_content: THP1_overexpression_processed_20210506.txt; tab delimited text file; normalized FPKM table

organism: Homo sapiens

source_name: THP-1 cells

growth_protocol: THP-1 were maintained at 37°C with 5% atmospheric CO2 in RPMI 1600 supplemented with 10% FBS fetal bovine serum, 1% penicillin/streptomycin (Thermo Fisher Scientific)  and 0.05 mM beta-mercaptoethanol  (Invitrogen). THP-1 were grown for an average of 7 days (with a range of 4-16 days) to select successfully transduced cells and reach 1x106 cells/ml (3-5 x106 cells in total) before RNA extraction.

extract_protocol:
Total RNA was extracted from stably transduced THP-1 cultures using the RNeasy Plus Mini kit (Qiagen) according to manufacturer’s protocol. The RNA samples were quantified, and quality controlled using an Agilent RNA 6000 Nano kit (Agilent) on an Agilent Bioanalyzer 2100 system. Samples with RNA Integrity Number (RIN) below 8 were discarded.
RNA samples were transformed into barcoded DNA libraries using TruSeq Stranded mRNA library preparation kits (Illumina), which were then paired-end sequenced, generating 2x100bp reads, using an Illumina HiSeq2000 sequencer

treatment_protocol: THP-1 cells were transduced in triplicate via spinoculation by open reading frames (ORFs) for 42 genes cloned into a modified GATEWAY® compatible polycistronic lentiviral expression vector, pLVX-EF1a-IRES-PURO/eGFP

value: Monocytic leukemia

tag: overexpressed gene

tag: sampleID

tag: plasmid

tag: infection date

tag: days in culture

tag: sequencing batch

value: A


title: MB49_untreated_2

value: untreated


title: THP-1 ORF PTGIR rep3

description: THP-1 stable cell line overexpressing PTGIR gene / replicate 3

value: PTGIR


title: MB49_POM1_1


title: Sample_73_RORg_MHCII


title: THP-1 ORF PTGIR rep2

description: THP-1 stable cell line overexpressing PTGIR gene / replicate 2


title: Sample_65_RORg_MHCII


title: LHCN-P3F1-002

city: Memphis

first: Hongjian

last: Jin

state: TN

address: 262 Danny Thomas Place

department: Center for Applied Bioinformatics

institute: St Jude Children's Research Hospital

data_processing:
Total stranded RNA sequencing data were processed by the internal AutoMapper pipeline.First the raw reads were first trimmed (Trim-Galore version 0.60), mapped to human genome assembly (hg19) (STAR v2.7).
The gene level expression values were quantified (RSEM v1.31) based on GENCODE annotation (v19) .
Assembly: GRCh37(hg19)
Supplementary files format and content: gene-level RSEM raw count, TPM, FPKM text files

source_name: cell line

extract_protocol:
Cell culture: LHCN-M2-GFP and LHCN-M2-PAX3-FOXO1 cells were cultured in medium as reported 1 [four parts Dulbecco's modified Eagle's medium (11965-084, Gibco) to one part medium 199 (11150-059, Gibco)] supplemented with 15% fetal bovine serum, 0.02 M HEPES (15630, Gibco), Basic FGF 10 ng/ml (HRP-0011, BioPioneer), 2.5ng/ml HGF (GF116, Chemicon Int’l), 0.055 µg/ml dexamethasone (D-2915, Sigma-Aldrich), 0.03 µg/ml zinc sulphate (ZnSO4) (Z4750, Sigma-Aldrich) and 1.4 µg/ml vitamin B12 (V2876, Sigma-Aldrich), 60 U/ml penicillin/streptomycin (15140122, Gibco) on dishes coated with 0.1% porcine skin gelatin (G1890-500G, Sigma-Aldrich). All cells were maintained at 37 °C in an atmosphere of 5% CO2.
Total RNA was extracted from xenograft tissues by RNeasy Mini Kit (cat. # 74034) from QIAGEN.
RNA libraries were prepared for sequencing using standard Illumina protocols at the St. Jude Hartwell Center.

tag: cell line

value: PAX3-FOXO1 overexpression


title: THP-1 ORF PTGIR rep1

description: THP-1 stable cell line overexpressing PTGIR gene / replicate 1


title: LHCN-P3F1-001


title: Sample_66_RORg_MHCII


title: MB49_untreated_1


title: THP-1 ORF PRKCB rep2

description: THP-1 stable cell line overexpressing PRKCB gene / replicate 2

value: PRKCB


title: Sample_63_RORg_MHCII


title: Input_3T3_Runx2BioFL_OM_day0

city: Bunkyo-ku

first: Hironori

last: Hojo

state: Tokyo

address: 7-3-1 Hongo

department: Center for Disease Biology and Integrative Medicine

country: Japan

institute: The University of Tokyo

library_source: genomic

data_processing:
DNA-sequence information was aligned to the unmasked mouse genome by bowtie
Peak calling was performed by two-sample analysis on CisGenome software (Ji et al., 2008) with a P-value cutoff of 10-5 comparing with the negative control
In the analysis with ATAC_Col2, ATAC_Col10 and ATAC_Sp7 data sets, to identify the cell-type specific and shared regions, we selected ATAC-seq peaks with the ATAC-seq normalized peak intensity based on the following criteria: raw fe > 30, fold change > 5, and p-value < 0.05, for cell-type specific regions. The differential accessible regions were called as below. C2: C2-Cho-distinct peaks; C10: C10-Cho-distinct peaks; S7: S7-Ob-distinct peaks; C2C10: peaks with high intensity in both C2-Cho and C10-Cho but not in S7-Ob; C10S7: peaks with high intensity in both C10-Cho and S7-Ob but not in C2-Cho; C2S7: peaks with high intensity in both C2-Cho and S7-Ob but not in C10-Cho. These peaks were listed in "ATAC_C2_C10_Sp7_peaks.xlsx”
In the analysis with ATAC_POB_Runx2_cHetKO and ATAC_POB_Runx2_cHomKO data sets, to identify the differential binding regions between the genotypes, DiffBind was applied with the following parameters "summits = 150" in the step of "dba.count" to resize peaks. To obtain statistically different chromatin-accessible regions, we selected the peaks based on FDR < 0.05. The differential accessible regions were listed in “ATAC_Runx2KO_peaks. xlsx”
In the analysis with ATAC_3T3cells data sets, to identify Runx2-responsive and osteoblast induction-responsive regions, DiffBind was applied with the following parameters "summits = 250" in the step of "dba.count" to resize peaks. Pairwise comparison was performed among ATAC-seq profiles of day 0 GFP expression samples, day 0 Runx2 expression samples, day 3 GFP expression samples, day 3 Runx2 expression samples, and no expression samples (None). The Runx2-responsive and the osteoblast induction-responsive regions were selected as ATAC-seq peaks whose intensity was statistically upregulated by both Runx2 expression and osteogenic induction on day 3, based on the following criteria: FDR < 0.05, fold change > 4. The Runx2-responsive regions were listed in “ATAC_3T3_Runx2_responsive_regions.xlsx”.
Genome_build: mm9
Supplementary_files_format_and_content: peak files

molecule: genomic DNA

source_name: 3T3 cells

extract_protocol:
Rib chondrocytes were manually dissected from the neonatal mouse ribs. The ribs were initially digested in 2 mg/mL Pronase (53702, EMD Millipore Corporation) for 30 min at 37 °C, washed three times with phosphate buffered saline (PBS), and then digested in 0.2% collagenase D (1088858, Roche Applied Science) for 3 h at 37 °C. Calvarial osteoblasts were manually dissected from osteoblast-enriched partial calvarias consisting of the frontal bone and a rostral part of the parietal bone. The partial calvarias were digested in five changes of 0.1% collagenase D and 0.2% dispase for 10 min each at 37 °C.   Cultured 3T3 cells were digested in trypsin-EDTA solution 1X (Wako, 206-17291) for 5 to 15 min at  37 °C.
ChIP-seq libraries were constructed using a ThruPLEX®-FD Prep Kit (R40012, Rubicon Genomics) according to the manufacturer′s instructions. ATAC-seq libraries were constructed as previously described (Buenrostro et al., 2013).

treatment_protocol: For in vitro cell culture, osteogenic medium contains 10% FBS/DMEM supplemented with 50 μg/ml ascorbic acid phosphate, 10 mM β-glycerophosphate, and 100 ng/ml rhBMP‑2

value: 3T3 cells overexpressing Runx2BioFL

value: 3T3 cells


title: Input_3T3_GFP_OM_day0


title: THP-1 ORF PRKCB rep3

description: THP-1 stable cell line overexpressing PRKCB gene / replicate 3


title: Sample_64_RORg_MHCII


title: LHCN-Ctrl-002

value: control


title: THP-1 ORF RCL1 rep3

description: THP-1 stable cell line overexpressing RCL1 gene / replicate 3


title: Sample_62_RORg_MHCII


title: Sample_61_RORg_MHCII


title: LHCN-Ctrl-001


title: Input_Runx2-FLAG_3T3_day(-2)


title: RH30_H189A_7C09_2

data_processing:
Total stranded RNA sequencing data were processed by the internal St Jude Cloud WARDEN pipeline.The raw reads were firs trimmed (Trim-Galore version 0.4.4),  mapped to human genome assembly (hg19) (STAR v2.5.3a).
The gene level expression values were quantified (HTseq v0.9.1) based on GENCODE annotation (v19) .
Assembly: GRCh37(hg19)
Supplementary files format and content: gene-level  raw count txt file

growth_protocol: RH30 and RH30_H189A  cells were cultured in RPMI medium, supplemented with 100 U/mL penicillin/streptomycin, and 10% FBS. CHLA15, CHLA20, NB1643 and NB1691 were culture with DMEM medium, supplemented with 2 mM L-glutamine and 10% FBS.

extract_protocol:
Total RNA was extracted from cells by RNeasy Mini Kit (cat. # 74104) from QIAGEN.
RNA libraries were prepared for sequencing using standard Illumina protocols at the St. Jude Hartwell Center.

treatment_protocol: RH30 and RH30_H189A cells were culuter for 24h to extract RNA.

value: KDM4B-H189A mutation clone 7C09


title: THP-1 ORF PRKCB rep1

description: THP-1 stable cell line overexpressing PRKCB gene / replicate 1


title: THP-1 ORF RCL1 rep2

description: THP-1 stable cell line overexpressing RCL1 gene / replicate 2


title: ChIP_Runx2-FLAG_3T3cells_Runx2BioFL_OM_day3

value: 3T3 cells overexpressing Runx2BioFL and cultured with osteogenic medium for 3 days


title: RH30_H189A_7C09_1


title: RH30_Parental_2

value: KDM4B wt


title: Sample_60_RORg_MHCII


title: THP-1 ORF PHTF1 rep2

description: THP-1 stable cell line overexpressing PHTF1 gene / replicate 2


title: ChIP_Runx2-FLAG_3T3cells_GFP_OM_day0

value: 3T3 cells overexpressing GFP


title: THP-1 ORF NOD2 rep3

description: THP-1 stable cell line overexpressing NOD2 gene / replicate 3


title: ATAC-seq_3T3cells_day3_Runx2-BioFL_rep2


title: THP-1 ORF PHTF1 rep1

description: THP-1 stable cell line overexpressing PHTF1 gene / replicate 1


title: Sample_56_RORg_MHCII


title: Sample_57_RORg_MHCII


title: Sample_59_RORg_MHCII


title: RH30-GSKJ4-H3K27ac-01

data_processing:
Library strategy: CUT&Tag
Raw reads were first trimmed low-quality ends using trim-galore (0.4.4; default parameter) and then aligned to the human reference genomes (hg19) using BWA (version 0.7.12; default parameter) and duplicated reads were then marked/removed with Picard (version 1.65). Only properly paired uniquely mapped reads were extracted by samtools (version 1.3.1 parameters used were -q 1 -f 2 -F 1804) for downstream analysis.
MACS2 (version 2.1.1 20160309) was used to call narrow peaks  using 'macs2 callpeak  -t cut_tag_file -f BEDPE -q 0.05 --keep-dup all' .  For calling broad peaks, we used SICER (version 1.1) with parameters of redundancy threshold 1, window size 200bp, effective genome fraction 0.86, gap size 600bp and E 0.00001.
genomeCoverageBed (bedtools 2.25.0) was used to produce genome-wide coverage in BEDGRAPH file and then converted it to bigwig file by bedGraphToBigWig.
Assembly: hg19
Supplementary files format and content: bigwig

growth_protocol: RH30 cells were cultured with RPMI 1640 media supplemented with 100 U/mlpenicillin/streptomycin and 10% fetal bovine serum. Cells were maintained at 37 °C in an atmosphere of 5% CO2.

extract_protocol:
CUT&Tag DNAs from RH30 cells with various treatments were prepared by following the protocol as described previously (Kaya-Okur et al. 2019 and https://www.protocols.io/view/bench-top-cut-amp-tag-bcuhiwt6?step=1 ) with minor modifications. RH30 with vairous treated approximately 500,000 cells were washed with wash buffer (20 mM HEPES pH 7.5; 150 mM NaCl; 0.5 mM Spermidine; 1× Protease inhibitor cocktail). Nuclei were isolated with cold NE1 buffer (20 mM HEPES–KOH, pH 7.9; 10 mM KCl 0.1%; Triton X-100; 20% Glycerol, 0.5 mM Spermidine; 1x Protease Inhibitor) for 10 min on ice. Nuclei were collected by 600 x g centrifuge and resuspended in 1ml washing buffer containing with 10 µL of activated concanavalin A-coated beads (Bangs laboratories, BP531) at RT for 10 min.  Bead-bound nuclei were collected with placing tube on magnet stand and removing clear liquid. The nuclei bound with bead were resuspended in 50 µL Dig-150 buffer (20 mM HEPES pH 7.5; 150 mM NaCl; 0.5 mM Spermidine; 1× Protease inhibitor cocktail; 0.05% Digitonin; 2 mM EDTA) and incubated with a 1:50 dilution of H3K27ac (Abcam, ab4729) for compounds treated cells and H3K9me3 (Abcam, ab76916) for shRNA transfected cells overnight at 4 °C. The unbound primary antibody was removed by placing the tube on the magnet stand and withdrawing the liquid. The primary antibody bound nuclei bead was mixed with Dig-150 buffer 100uL containing guinea pig anti-Rabbit IgG antibody (Antibodies, ABIN101961) 1:100 dilution for 1 hour at RT. Beads bound nuclei were washed using the magnet stand 3× for 5 min in 1 mL Dig-150 buffer to remove unbound antibodies.  A 1:100 dilution of pA-Tn5 adapter complex was prepared in Dig-300 buffer (20 mM HEPES, pH 7.5, 300 mM NaCl, 0.5 mM Spermidine, 0.05% Digitonin, 1× Protease inhibitor cocktail).  After removing the liquid on the magnet stand, 100 µL mixture of pA-Tn5 and Dig- 300 buffer was added to the nuclei bound beads with gentle vortex and incubated at RT for 1 h. After 3× 5 min in 1 mL Dig-300 buffer to remove unbound pA-Tn5 protein, nuclei were resuspended in 250 µL Tagmentation buffer (10 mM MgCl2 in Dig-300 buffer) and incubated at 37 °C for 1 h. EDTA 10 µL of 0.5 M, 3 µL of 10% SDS and 2.5 µL of 20 mg/mL Proteinase K were added to stop tagmentation and incubated at 55 °C for 1 hour. DNA was then precipitated by phenol/chloroform/isoamylalcohol followed by ethanol precipitation with glycogen and then dissolved in water.
Sequencing libraries were prepared using NEBNext HiFi 2× PCR Master Mix (NEB, M0541L) according to the manufacturer's instructions. The PCR products were cleaned up with SPRIselect beads and quantified using Qubit dsDNA HS assay kit (Agilent Technologies). The libraries were sequenced on a HiSeq2500 with paired-end 50-bp reads (Illumina).

treatment_protocol: RH30 cells were either treated with compounds (DMSO 0.1% or C70  5µM or GSKJ4  2.5µM) for 48 h or transfected with shRNAs (shRNA control or KDM4B sh814) for 72 hours.

value: GSKJ4 2.5 microM

tag: chip antibody

value: H3K27ac,  Rabbit, Abcam,4729


title: THP-1 ORF NOD2 rep1

description: THP-1 stable cell line overexpressing NOD2 gene / replicate 1


title: Sample_58_RORg_MHCII

value: mesenteric lymph nodes (MLN)

value: three weeks


title: RH30-GSKJ4-H3K27ac-02


title: THP-1 ORF PHTF1 rep3

description: THP-1 stable cell line overexpressing PHTF1 gene / replicate 3

value: Monocytic leukemia


title: RH30-C70-H3K27ac-02

value: C70 5 microM


title: ATAC-seq_3T3cells_day3_Runx2-BioFL_rep1


title: Sample_55_RORg_MHCII

value: RORgt+ MHCII+


title: THP-1 ORF NOD2 rep2

description: THP-1 stable cell line overexpressing NOD2 gene / replicate 2

value: A


title: ATAC-seq_3T3cells_day3_GFP_rep1

value: 3T3 cells overexpressing GFP and cultured with osteogenic medium for 3 days


title: ATAC-seq_3T3cells_day3_GFP_rep2


title: Sample_54_RORg_MHCII


title: THP-1 ORF KIAA1919 rep2

description: THP-1 stable cell line overexpressing KIAA1919 gene / replicate 2


title: RH30-DMSO-H3K27ac-02

value: DMSO 0.1percent


title: RH30-C70-H3K27ac-01


title: THP-1 ORF KIAA1919 rep3

description: THP-1 stable cell line overexpressing KIAA1919 gene / replicate 3


title: ChIP_Runx2-FLAG_3T3cells_GFP_OM_day3


title: RH30_Parental_1


title: ChIP_Runx2-FLAG_3T3cells_Runx2BioFL_OM_day0

value: 3T3 cells overexpressing Runx2BioFL


title: Sample_53_RORg_MHCII

description:
RORgt+ MHCII+ from 3-week old mice
smart-seq2 protocol


title: ATAC-seq_3T3cells_day0_Runx2-BioFL_rep2

value: 3T3 cells


title: THP-1 ORF KIAA1919 rep1

description: THP-1 stable cell line overexpressing KIAA1919 gene / replicate 1


title: Sample_52_RORg_MHCII


title: RH30-DMSO-H3K27ac-01


title: ATAC-seq_3T3cells_day0_Runx2-BioFL_rep1


title: THP-1 ORF IRF5 rep3

description: THP-1 stable cell line overexpressing IRF5 gene / replicate 3


title: THP-1 ORF IRF5 rep2

description: THP-1 stable cell line overexpressing IRF5 gene / replicate 2


title: RH30-sh814-H3K9me3-2

value: H3K9me3, Rabbit, Abcam,76916


title: Sample_50_RORg_MHCII


title: Sample_51_RORg_MHCII


title: ATAC-seq_3T3cells_day0_GFP_rep2


title: THP-1 ORF IRF5 rep1

description: THP-1 stable cell line overexpressing IRF5 gene / replicate 1


title: ATAC-seq_3T3cells_day0_GFP_rep1


title: RH30-sh814-H3K9me3-1


title: RH30-shCtrl-H3K9me3-1

value: shRNA-control


title: RH30-shCtrl-H3K9me3-2


title: THP-1 ORF IFIH1 rep3

description: THP-1 stable cell line overexpressing IFIH1 gene / replicate 3


title: ATAC-seq_3T3cells_day(-2)_rep1


title: THP-1 ORF IFIH1 rep2

description: THP-1 stable cell line overexpressing IFIH1 gene / replicate 2


title: Sample_49_Aire_DC

description:
Aire(GFP)+ Dendritic Cell from 3-week old mice
smart-seq2 protocol

source_name: Mouse MLN  Aire+ DC

value: Aire(GFP)+ Dendritic Cell


title: Sample_48_Aire_DC


title: RH30-si814-input-2

data_processing:
ChIP-seq raw reads were first trimmed low-quality ends using trim-galore (0.6.6; default parameter) and then aligned to the human reference genomes (hg19) using BWA (version 0.7.12; default parameter) and duplicated reads were then marked with Picard (version 1.65), with only nonduplicated reads kept by samtools (version 1.3.1, parameter ‘‘-q 1 -F 1024’’. We utilized genomeCoverageBed (bedtools 2.25.0) to obtain genome-wide coverage in BEDGRAPH file format and then converted it to bigwig file format by bedGraphToBigWig. The bigwig files were scaled to 15 million reads to allow comparison across samples.
H3K36me3 enriched domains were called by SICER (version 1.1, with parameters of redundancy threshold 1, window size 200 bp, effective genome fraction 0.86, gap size 600 bp, FDR 0.00001)
genomeCoverageBed (bedtools 2.25.0) was used to produce genome-wide coverage in BEDGRAPH file and then converted it to bigwig file by bedGraphToBigWig.
Assembly: hg19(GRCh37)
Supplementary files format and content: bigwig,peak

growth_protocol: RH30 cells were cultured in RPMI medium, supplemented with 100 U/mL penicillin/streptomycin, and 10% FBS.

extract_protocol:
RH30 cells 10 millions with shRNAs transfected for 72h were cross-linked with fresh prepare 1% formaldehyde (final concentration) for 10 min and quenched with 125 mM glycine for 5 min at room temperature. Cells were washed once with Dulbecco's Phosphate-Buffered Saline (DPBS). Cell lysis buffer (5 mM PIPES pH 8.0, 85 mM KCl, 0.5% NP-40 with 1 tablet protease inhibitor cocktail) were added to plate after aspirate DPBS and add 5-8 ml cold (4°C). The cells were scraped with a cell scraper and transfer into 50-ml conical tubes on ice.  Cell pellets were collected by centrifuging at 2,000 rpm for 5 minutes at 4°C. The pellets were resuspended in cell lysis buffer and incubated on ice for 10 min. Cells were passed through a 20-gauge needle 20 times. The nuclei were collected by centrifuging at 2,000 rpm at 4°C for 5 minutes. The nuclear pellets were resuspended and lysed with 250 μl RIPA buffer (1X PBS, 1% NP-40, 0.5% sodium deoxycholate, 0.1% SDS with 1 tablet protease inhibitor cocktail). The lysates were sonicated for at least 30 min (30 sec on / 30 sec off) using Bioruptor Pico (Diagenode, Inc., Denville, NJ) and centrifuged at 14,000 rpm in a microfuge for 15 minutes at 4°C. The superannuants were collected, and sonicated DNA fragments were examined by electrophoresis on a 1% agarose gel. The sonicated samples with enrichment of fragments in the 100 to 500 bp range will be used for the chromatin immunoprecipitation (Chip) after remove 50 μL from each sample as input. The primary antibody (H3K36me3, RevMab,31-1051-00) was coupled with 25 μL magnetic beads (Dynabeads M-280 Sheep Anti-Rabbit IgG, Invitrogen 11203D) overnight in the cold room at 4°C. The primary antibody coupled magnetic beads were washed three times with PBS/BSA (1XPBS / 5 mg/ml BSA (fraction V)) at 4°C, then incubated with sonicated DNA chromatin samples overnight in the cold room at 4°C. The beads containing immuno-bound chromatin were collected by placing the microfuge tube on a magnet rack. The beads were extensively washed with LiCl wash buffer (100 mM Tris pH 7.5 / 500 mM LiCl / 1% NP-40 / 1% sodium deoxycholate) 5 times and TE Buffer (10 mM Tris-HCl pH 7.5 / 0.1 mM Na2EDTA) once. Bound chromatin was eluted by IP Elution Buffer (1 % SDS / 0.1 M NaHCO3) and reverse-crosslinked at 65°C overnight. DNAs were purified using Min-Elute PCR purification kit (Qiagen, Valencia, CA) after the treatment of RNase A and proteinase K.
5-10 ng of DNA was using to prepare libraries by the NEBNext ChIP-Seq Library Prep Reagent Set for Illumina with NEBNext Q5 Hot Start HiFi PCR Master Mix according to the manufacturer’s instructions (New England Biolabs). Completed ChIP-seq libraries were analyzed for insert size distribution on a 2100 BioAnalyzer High Sensitivity kit (Agilent) or Caliper LabChip GX DNA High Sensitivity Reagent Kit (Perkin Elmer). All libraries were quantified using the Quant-iT PicoGreen dsDNA assay (Life Technologies), Kapa Library Quantification kit (Kapa Biosystems).

treatment_protocol: RH30 cells were transfect with shRNAs (shRNA-control or KDM4B shRNA-814) for 72h.

tag: shRNA


title: ATAC-seq_3T3cells_day(-2)_rep2


title: ATAC-seq_osteoblasts_Runx2_cHom_KO_rep2

source_name: E17.5 calvaria osteoblasts

value: Calvarial osteoblasts at E17.5


title: ATAC-seq_osteoblasts_Runx2_cHom_KO_rep1


title: THP-1 ORF IFIH1 rep1

description: THP-1 stable cell line overexpressing IFIH1 gene / replicate 1


title: RH30-si814-input-1


title: Sample_46_Aire_DC


title: RH30-siCtrl-input-2


title: THP-1 ORF GPR18 rep3

description: THP-1 stable cell line overexpressing GPR18 gene / replicate 3


title: Sample_45_Aire_DC


title: ATAC-seq_osteoblasts_Runx2_cHet_KO_rep2


title: THP-1 ORF GPR18 rep2

description: THP-1 stable cell line overexpressing GPR18 gene / replicate 2


title: ATAC-seq_osteoblasts_Runx2_cHet_KO_rep1

description: ATAC_Runx2KO_peaks. xlsx


title: Sample_44_Aire_DC


title: Sample_43_Aire_DC


title: RH30-siCtrl-input-1


title: RH30-sh814-H3K36me3-2


title: ChIP-seq_Jun_Ob

source_name: P1 calvaria osteoblasts

value: Calvarial osteoblasts at P1

value: wild-type


title: THP-1 ORF GPR18 rep1

description: THP-1 stable cell line overexpressing GPR18 gene / replicate 1


title: Sample_47_Aire_DC


title: THP-1 ORF AUH rep2

description: THP-1 stable cell line overexpressing AUH gene / replicate 2

value: AUH


title: Sample_41_Aire_DC

value: mesenteric lymph nodes (MLN)


title: RH30-shCtrl-H3K36me3-2


title: THP-1 ORF AUH rep1

description: THP-1 stable cell line overexpressing AUH gene / replicate 1


title: THP-1 ORF AUH rep3

description: THP-1 stable cell line overexpressing AUH gene / replicate 3


title: Sample_39_Aire_DC

value: three weeks


title: THP-1 ORF FAM49B rep3

description: THP-1 stable cell line overexpressing FAM49B gene / replicate 3

value: Monocytic leukemia


title: Sample_40_Aire_DC


title: THP-1 ORF FAM49B rep2

description: THP-1 stable cell line overexpressing FAM49B gene / replicate 2


title: THP-1 ORF FAM49B rep1

description: THP-1 stable cell line overexpressing FAM49B gene / replicate 1


title: Sample_37_Aire_DC


title: ChIP-seq_Runx2-FLAG_Ob_rep2

value: Runx2-BioFL/+ knock-in


title: Sample_38_Aire_DC


title: Input_Runx2-FLAG_Ob_2


title: ChIP-seq_Runx2-FLAG_Cho_rep2

source_name: P1 rib chondrocytes

value: Rib chondrocytes at P1


title: THP-1 ORF RNFT1 rep3

description: THP-1 stable cell line overexpressing RNFT1 gene / replicate 3

value: A


title: Sample_42_Aire_DC


title: RH30-sh814-H3K36me3-1


title: IgGcontrol_Foxa2_Cho


title: THP-1 ORF RNFT1 rep2

description: THP-1 stable cell line overexpressing RNFT1 gene / replicate 2


title: ATAC-seq_Sp7+_osteoblasts_rep3

value: Sp7-EGFP-positive calvarial osteoblasts at P1

value: Sp7-EGFP tg


title: Sample_35_Aire_DC


title: THP-1 ORF RNFT1 rep1

description: THP-1 stable cell line overexpressing RNFT1 gene / replicate 1


title: ChIP-seq_Foxa2_Cho


title: Sample_36_Aire_DC


title: ATAC-seq_Col2+_chondrocytes_rep2

value: Col2a1-ECFP-positive rib chondrocytes at P1

value: ICR

value: Col2a1-ECFP tg


title: ATAC-seq_Col2+_chondrocytes_rep3


title: RH30-shCtrl-H3K36me3-1

value: shRNA-control


title: THP-1 ORF CHSY3 rep3

description: THP-1 stable cell line overexpressing CHSY3 gene / replicate 3


title: Sample_33_Aire_DC

value: Aire(GFP)+ Dendritic Cell


title: ATAC-seq_Col2+_chondrocytes_rep1


title: THP-1 ORF CHSY3 rep2

description: THP-1 stable cell line overexpressing CHSY3 gene / replicate 2


title: THP-1 ORF CHSY3 rep1

description: THP-1 stable cell line overexpressing CHSY3 gene / replicate 1


title: Input_Runx2-FLAG_Cho_2


title: SEM3, 10, Cas9

city: Padova

first: Oriana

last: Romano

address: Via U. Bassi 58/B

department: Molecular Medicine

country: Italy

institute: University of Padova

data_processing:
Read quality was verified using FastQC (v. 0.11.9). Raw reads were trimmed for adapters and low-quality tails (quality < Q20) with BBDuk (v. 38.92); moreover, the first 10 nucleotides were force-trimmed for low quality. Reads shorter than 35 bp after trimming were removed.
Reads were subsequently aligned to the human reference genome (hg38) using STAR (v. 2.7.9a). Raw gene counts were obtained in R-4.1.1 using the featureCounts function of the Rsubread R package (v. 2.6.4) and the GENCODE 38 basic gene annotation for hg38 reference genome.
Gene counts were normalized to counts per million mapped reads (CPM) and to fragments per kilobase of exon per million mapped reads (FPKM) using the edgeR R package (v. 3.34.1); only genes with a CPM greater than 1 in at least 3 samples were retained for differential analysis.
Differential gene expression analysis was performed using the glmQLFTest function of the edgeR R package, using donor as a blocking variable.
Genome_build: hg38
Supplementary_files_format_and_content: CPM obtained using edgeR

molecule: polyA RNA

source_name: hematopoietic stem/progenitor cells

growth_protocol: CD34+ cells were thawed and cultured at a concentration of 5x10^5 cells/ml in the “HSPC medium” containing StemSpan (STEMCELL Technologies) supplemented with penicillin/streptomycin (Gibco), 250 nM StemRegenin1 (STEMCELL Technologies), and the following recombinant human cytokines (PeproTech): human stem cell factor (SCF) (300 ng/ml), Flt-3L (300 ng/ml), thrombopoietin (TPO) (100 ng/ml), and interleukin-3 (IL-3) (60 ng/ml).

extract_protocol:
Total RNA was isolated using the RNeasy Kit (QIAGEN) including a DNAse treatment step
RNA-seq libraries were prepared starting from 30 ng of total RNA using the Universal Plus mRNA-Seq kit (Nugen) as recommended by the manufacturer. Briefly, mRNA was captured with polyA+ magnetic beads from total RNA. mRNA was chemically fragmented. Single strand and second strand cDNA were produced and then ligated to Illumina compatible adaptors with UDI. To produce oriented RNA-seq libraries, a final step of strand selection was performed. The NuQuant system (Nugen) was used to quantify the RNA-seq libraries.

treatment_protocol: CD34+ HSPCs were transfected with 3.0 μg of CBE or ABE encoding mRNA, and a synthetic sgRNA at a final concentration of 2.3 μM. We used the P3 Primary Cell 4D-Nucleofector X Kit S or L (Lonza) and the CA137 program (Nucleofector 4D). CD34+ HSPCs were transfected with RNP complexes (containing Cas9-GFP protein and a synthetic sgRNA) using the P3 Primary Cell 4D-Nucleofector X Kit S (Lonza) and the CA137 program (Nucleofector 4D) in the presence of a transfection enhancer (IDT). Cells transfected with TE buffer served as negative controls.

value: hematopoietic stem/progenitor cells

tag: developmental stage

value: adult


title: RNA-seq_osteoblasts_Runx2_cHom_KO_rep2

data_processing:
Trimmed the sequences by Trimomatic-0.33
Aligned the sequence to the mouse genome reference sequence mm9 byby Tophat-2.1.0
Quantification of gene expression and differentially expressed gene analysis were performed using Cufflinks/Cuffdiff 2.2.1
The data from Cufflinks/Cuffdiff 2.2.1 were output as fragments per kilobase of transcript per million mapped reads (FPKM) values
For comparison analysis among Col2a1-ECFP chondrocytes, Col10a1-mCherry chondrocytes with calvaria osteoblasts sorted from Sp7-GFP mice at P1, public data set for RNA-seq data of Sp7-GFP osteoblast (GSM1976262, GSM1976263, GSM1976264) was used for Cufflinks/Cuffdiff analysis
Genome_build: mm9
Supplementary_files_format_and_content: tab-delimited text files include FPKM values

extract_protocol:
Rib chondrocytes were manually dissected from the neonatal mouse ribs. The ribs were initially digested in 2 mg/mL Pronase (53702, EMD Millipore Corporation) for 30 min at 37 °C, washed three times with phosphate buffered saline (PBS), and then digested in 0.2% collagenase D (1088858, Roche Applied Science) for 3 h at 37 °C. Calvarial osteoblasts were manually dissected from osteoblast-enriched partial calvarias consisting of the frontal bone and a rostral part of the parietal bone. The partial calvarias were digested in five changes of 0.1% collagenase D and 0.2% dispase for 10 min each at 37 °C.
For RNA-seq with rib chondrocytes sorted from Col2a1-ECFP and Col10a1-mCherry mice at P1, 100 ng of total RNA was subjected to analysis. RNA libraries were constructed using a Tru-Seq Illumina kit (RS-122-2001, Illumina) with some modifications. For RNA-seq for with Runx2_cHet_KO cells and Runx2_cHom_KO cells from calvaria, 1.7 ng of total RNA was subjected to the analysis. RNA libraries were constructed using a SMARTer Stranded Total RNA-Seq Kit v2 Pico Input Mammalian (634411, Takara).

value: Calvarial osteoblasts at E17.5


title: Sample_32_Aire_DC

description:
Aire(GFP)+ Dendritic Cell from 3-week old mice
smart-seq2 protocol

tag: tissue


title: Sample_31_Aire_DC


title: SEM3, 9, ABE

value: ABE


title: RNA-seq_osteoblasts_Runx2_cHet_KO_rep2

tag: strain


title: RNA-seq_osteoblasts_Runx2_cHom_KO_rep1

tag: genotype


title: Sample_30_Aire_DC

value: mesenteric lymph nodes (MLN)


title: Sample_34_Aire_DC

value: three weeks


title: THP-1 ORF CDC37 rep2

description: THP-1 stable cell line overexpressing CDC37 gene / replicate 2

value: Monocytic leukemia


title: SEM3, 1, control

value: control


title: Sample_29_Aire_DC


title: THP-1 ORF BANK1 rep3

description: THP-1 stable cell line overexpressing BANK1 gene / replicate 3


title: Sample_27_Aire_DC


title: THP-1 ORF CDC37 rep1

description: THP-1 stable cell line overexpressing CDC37 gene / replicate 1


title: RNA-seq_osteoblasts_Runx2_cHet_KO_rep1


title: SEM2, 10, Cas9


title: Sample_28_Aire_DC


title: SEM2, 9, ABE


title: SEM3, 5, CBE

value: CBE


title: THP-1 ORF BANK1 rep2

description: THP-1 stable cell line overexpressing BANK1 gene / replicate 2

value: A


title: Sample_25_Aire_DC


title: Sample_26_Aire_DC


title: THP-1 ORF BANK1 rep1

description: THP-1 stable cell line overexpressing BANK1 gene / replicate 1


title: THP-1 ORF ZBTB40 rep2

description: THP-1 stable cell line overexpressing ZBTB40 gene / replicate 2

value: B


title: SEM2, 5, CBE


title: SEM2, 1, control


title: Sample_23_Aire_DC


title: SEM1, 10, Cas9, bis


title: THP-1 ORF ZBTB40 rep1

description: THP-1 stable cell line overexpressing ZBTB40 gene / replicate 1


title: Sample_24_Aire_DC


title: SEM1, 10, Cas9


title: Sample_22_Aire_DC


title: THP-1 ORF CDC37 rep3

description: THP-1 stable cell line overexpressing CDC37 gene / replicate 3


title: THP-1 ORF TYK2 rep3

description: THP-1 stable cell line overexpressing TYK2 gene / replicate 3


title: Sample_21_Aire_DC


title: SEM1, 9, ABE


title: THP-1 ORF TYK2 rep2

description: THP-1 stable cell line overexpressing TYK2 gene / replicate 2


title: Sample_20_Aire_DC


title: THP-1 ORF TYK2 rep1

description: THP-1 stable cell line overexpressing TYK2 gene / replicate 1


title: Sample_19_Aire_DC


title: THP-1 ORF TNFRSF9 rep3

description: THP-1 stable cell line overexpressing TNFRSF9 gene / replicate 3


title: 12Z_siZMYND8_rep3_RNA

city: Grand Rapids

first: Ronald

middle: L.

last: Chandler

state: MI

department: Obstetrics, Gynecology and Reproductive Biology

institute: Michigan State University

description: mRNA (replicate #3) extracted from siZMYND8 treated (ZMYND8 knockdown) 12Z endometriotic epithelial cells.

data_processing:
Raw reads were trimmed with Trim Galore! (http://www.bioinformatics.babraham.ac.uk/projects/trim_galore/) followed by quality control analysis via FastQC (Andrews 2010) and MultiQC (Ewels, Magnusson et al. 2016).
Trimmed reads were aligned to GRCh38.p12 (hg38) assembly and indexed to GENCODE (v28) along with gene feature counting via STAR (Dobin, Davis et al. 2013).
Low count genes with less than 1 count per sample on average were filtered prior to count normalization and differential gene expression (DGE) analysis by DESeq2 with Empirical Bayes shrinkage for fold-change estimation (Love, Huber et al. 2014, Love, Anders et al. 2015).
Wald probabilities were corrected for multiple testing by independent hypothesis weighting (IHW) (Ignatiadis, Klaus et al. 2016) for downstream analyses.
Genome_build: GRCh38.p12
Supplementary_files_format_and_content: Tabular STAR gene-level feature raw read counts and transcriptome-wide differential gene expression analysis results from DESeq2.

source_name: 12Z endometriotic epithelial cells

growth_protocol: Human 12Z endometriotic epithelial cells were cultured in growth media (10% FBS, 1% L-glutamine, 1% Penicillin-Streptomycin).

extract_protocol:
Cells were purified for RNA using the Quick-RNA Miniprep Kit (Zymo Research).
Transcriptome libraries were prepared and sequenced by the Van Andel Genomics Core from 500 ng of total RNA using the KAPA mRNA HyperPrep kit (v4.17) (Kapa Biosystems). RNA was sheared to 300-400 bp. Prior to PCR amplification, cDNA fragments were ligated to IDT for Illumina unique dual adapters (IDT DNA Inc). Quality and quantity of the finished libraries were assessed using a combination of Agilent DNA High Sensitivity chip (Agilent Technologies), QuantiFluor dsDNA System (Promega), and Kapa Illumina Library Quantification qPCR assays (Kapa Biosystems). Individually indexed libraries were pooled and 50 bp, paired-end sequencing was performed on an Illumina NovaSeq 6000 sequencer using a 100-cycle sequencing kit (Illumina). Each library was sequenced to an average raw depth of 20-25 million reads. Base calling was done by Illumina RTA3 and output of NCS was demultiplexed and converted to FastQ format with Illumina Bcl2fastq v1.9.0.

treatment_protocol: Cells were seeded in antibiotic-free media the day before siRNA transfection. 50 nM siRNA (Dharmacon, ON-TARGETplus) were transfected into cells using the Lipofectamine RNAiMAX (ThermoFisher Scientific) reagent, according to the manufacturer protocol, in OptiMEM (Gibco). Growth media was replaced 24 hours following transfection, without antibiotics. 48 hours after transfection, low serum (0.5% FBS) growth media was added with antibiotics. Cells were harvested 72 hours following siRNA transfection.

value: 12Z endometriotic epithelial cells

tag: treatment/condition

value: siZMYND8 (ZMYND8 knockdown by siRNA)


title: Sample_18_Aire_DC


title: THP-1 ORF ZBTB40 rep3

description: THP-1 stable cell line overexpressing ZBTB40 gene / replicate 3


title: 12Z_siZMYND8_rep2_RNA

description: mRNA (replicate #2) extracted from siZMYND8 treated (ZMYND8 knockdown) 12Z endometriotic epithelial cells.


title: THP-1 ORF TNFRSF9 rep1

description: THP-1 stable cell line overexpressing TNFRSF9 gene / replicate 1


title: Sample_17_Aire_DC

description:
Aire(GFP)+ Dendritic Cell from 3-week old mice
smart-seq2 protocol


title: THP-1 ORF SNX32 rep3

description: THP-1 stable cell line overexpressing SNX32 gene / replicate 3


title: Sample_15_Aire_DC


title: 12Z_siH3F3B_rep3_RNA

description: mRNA (replicate #3) extracted from siH3F3B treated (H3.3 knockdown) 12Z endometriotic epithelial cells.

value: siH3F3B (H3F3B knockdown by siRNA)


title: Sample_13_Aire_DC


title: 12Z_siH3F3B_rep2_RNA

description: mRNA (replicate #2) extracted from siH3F3B treated (H3.3 knockdown) 12Z endometriotic epithelial cells.


title: 12Z_siH3F3B_rep1_RNA

description: mRNA (replicate #1) extracted from siH3F3B treated (H3.3 knockdown) 12Z endometriotic epithelial cells.


title: THP-1 ORF SNX32 rep2

description: THP-1 stable cell line overexpressing SNX32 gene / replicate 2


title: 12Z_Control_rep3_RNA

description: mRNA (replicate #3) extracted from non-targeting siRNA (control) treated 12Z endometriotic epithelial cells.

value: Control (non-targeting siRNA)


title: Sample_12_Aire_DC


title: Sample_14_Aire_DC


title: THP-1 ORF SLC22A5 rep6

description: THP-1 stable cell line overexpressing SLC22A5 gene / replicate 3


title: THP-1 ORF SNX32 rep1

description: THP-1 stable cell line overexpressing SNX32 gene / replicate 1


title: SEM1, 5, CBE

tag: treatment


title: SEM1, 1, control


title: THP-1 ORF SLC22A5 rep5

description: THP-1 stable cell line overexpressing SLC22A5 gene / replicate 2


title: THP-1 ORF TNFRSF9 rep2

description: THP-1 stable cell line overexpressing TNFRSF9 gene / replicate 2


title: Sample_11_Aire_DC

tag: age


title: 12Z_siZMYND8_rep1_RNA

description: mRNA (replicate #1) extracted from siZMYND8 treated (ZMYND8 knockdown) 12Z endometriotic epithelial cells.

tag: cell type


title: Sample_16_Aire_DC


title: Sample_9_Aire_DC


title: Sample_10_Aire_DC


title: THP-1 ORF SLC22A5 rep2

tag: overexpressed gene


title: THP-1 ORF SLC22A5 rep3

tag: sampleID


title: Sample_8_Aire_DC


title: 12Z_siCHD4siARID1A_rep2_RNA

description: mRNA (replicate #2) extracted from siCHD4+siARID1A treated (CHD4/ARID1A co-knockdown) 12Z endometriotic epithelial cells.

value: siCHD4+siARID1A (CHD4/ARID1A co-knockdown by siRNA)


title: 12Z_siCHD4siARID1A_rep3_RNA

description: mRNA (replicate #3) extracted from siCHD4+siARID1A treated (CHD4/ARID1A co-knockdown) 12Z endometriotic epithelial cells.


title: THP-1 ORF SLC22A5 rep1

description: THP-1 stable cell line overexpressing SLC22A5 gene / replicate 1

tag: plasmid


title: 12Z_Control_rep1_RNA

description: mRNA (replicate #1) extracted from non-targeting siRNA (control) treated 12Z endometriotic epithelial cells.


title: Sample_7_Aire_DC

value: Aire(GFP)+ Dendritic Cell


title: 12Z_siCHD4siARID1A_rep1_RNA

description: mRNA (replicate #1) extracted from siCHD4+siARID1A treated (CHD4/ARID1A co-knockdown) 12Z endometriotic epithelial cells.


title: THP-1 ORF PTPN2 rep3

description: THP-1 stable cell line overexpressing PTPN2 gene / replicate 3

tag: infection date


title: Sample_6_Aire_DC

description:
Aire(GFP)+ Dendritic Cell from 3-week old mice
smart-seq2 protocol


title: 12Z_Control_rep2_RNA

description: mRNA (replicate #2) extracted from non-targeting siRNA (control) treated 12Z endometriotic epithelial cells.


title: THP-1 ORF PTPN2 rep2

description: THP-1 stable cell line overexpressing PTPN2 gene / replicate 2

tag: days in culture


title: 12Z_siARID1A_rep3_RNA

description: mRNA (replicate #3) extracted from siARID1A treated (ARID1A knockdown) 12Z endometriotic epithelial cells.

value: siARID1A (ARID1A knockdown by siRNA)


title: Sample_5_Aire_DC


title: 12Z_siARID1A_rep2_RNA

description: mRNA (replicate #2) extracted from siARID1A treated (ARID1A knockdown) 12Z endometriotic epithelial cells.


title: THP-1 ORF PTPN2 rep1

description: THP-1 stable cell line overexpressing PTPN2 gene / replicate 1

tag: sequencing batch


title: Sample_4_Aire_DC


title: 12Z_siARID1A_rep1_RNA

description: mRNA (replicate #1) extracted from siARID1A treated (ARID1A knockdown) 12Z endometriotic epithelial cells.


title: THP-1 ORF SLC22A5 rep4


title: THP-1 ORF NFKB1 rep3

description: THP-1 stable cell line overexpressing NFKB1 gene / replicate 3


title: 12Z_siCHD4_rep3_RNA

description: mRNA (replicate #3) extracted from siCHD4 treated (CHD4 knockdown) 12Z endometriotic epithelial cells.

value: siCHD4 (CHD4 knockdown by siRNA)


title: Sample_3_Aire_DC


title: THP-1 ORF NFKB1 rep2

description: THP-1 stable cell line overexpressing NFKB1 gene / replicate 2


title: Sample_2_Aire_DC


title: 12Z_siCHD4_rep2_RNA

description: mRNA (replicate #2) extracted from siCHD4 treated (CHD4 knockdown) 12Z endometriotic epithelial cells.


title: 12Z_siCHD4_rep1_RNA

description: mRNA (replicate #1) extracted from siCHD4 treated (CHD4 knockdown) 12Z endometriotic epithelial cells.


title: 12Z_Control_non-targeting siRNA_rep3_RNA

description: mRNA (replicate #3) extracted from non-targeting siRNA (control) treated 12Z endometriotic epithelial cells.


title: THP-1 ORF LRRC32 rep3

description: THP-1 stable cell line overexpressing LRRC32 gene / replicate 3


title: Riboseq_P0_2

city: Berlin

first: Matthew

middle: Lee

last: Kraushar

state: Berlin

address: Ihnestrasse 63-73

department: Department of Computational Molecular Biology

country: Germany

institute: Max Planck Institute for Molecular Genetics

description: ribosome-protected mRNA fragment

data_processing:
Library strategy: Ribo-Seq
Riboseq reads were stripped of adaptor sequences using cutdapt, and contaminants such as transfer RNAs (tRNAs) and rRNA were removed by alignment to a contaminants index via STAR 2.7.0, consisting of nucleotide sequences from known mouse rRNA and tRNA sequences drawn from the gencode annotation.
Unaligned reads from this analysis were then aligned to mouse GENCODE release M12 (mus musculus)  with the STAR v 2.7.0 splice-aware alignment tool allowing for up to 1 mismatch. The star genome index was built using GENCODE M12. Only uniquely aligning reads were used.
The RiboseQC pipeline v1.1 (https://github.com/ohlerlab/RiboseQC) was used to confirm 3-nucleotide periodicity of the data and deduce P-site positions from the Riboseq reads.
Ribosome P-site count metaplots and fold change were calculated by first normalizing the P-site coverage track for each gene to the gene's total density, excluding genes with < 32 reads or less (low count filtering), and then all genes’ tracks were added together to generate a mean P-site density at each point for each sample. Confidence intervals were calculated by resampling from the 3 replicates for each condition.
Genome_build: GENCODE release M12 (Mus musculus)
Supplementary_files_format_and_content: tsv count table

source_name: neocortex

growth_protocol: Neocortex tissue was sub-dissected from embryos and pups from timed pregnant mice, as described in Kraushar et al. Molecular Cell. 81; 2021.

extract_protocol:
Neocortex tissue was lysed on ice in 20 mM HEPES, 100 mM KCl, 7.5 mM MgCl2, pH 7.4, supplemented with 20 mM Dithiothreitol (DTT), 0.04 mM Spermine, 0.5 mM Spermidine, 1x Protease Inhibitor cOmplete EDTA-free (Roche, 05056489001), 0.3% v/v IGEPAL CA-630 detergent (Sigma, I8896) and clarified by centrifugation at 16100 xg for 5 min at 4 °C with a benchtop centrifuge. Samples were then measured for A260 ODU on a NanoDrop 1000 Spectrophotometer. For digestion of ribosome protected RNA fragments (RPFs), Ribo-seq samples were then mixed with 60U RNAse-T1 plus 96 ng RNAse-A per ODU, and incubated for 30 min at 25 °C, shaking at 400 rpm. To stop RNAse activity, 200 U of SUPERase-In RNAse inhibitor was then added.
Riboseq sequencing library preparation: the RNA was first ligated to a 3’ adapter 4N-RA3, and gel-purified using a 15 % denaturing urea-PAGE gel . Next the 5’ adapter OR5-4N was ligated and gel purified. The RNA was reverse transcribed and PCR-amplified by Phusion High-Fidelity DNA polymerase (Thermo Fischer, F-530XL). The cDNA was visualized on a 2.5 % agarose gel, a ~150 bp sized fragment was excised and purified by Zymoclean Gel DNA Recovery kit (Zymo Research, D4007/D4008).

value: RNAse-T1 & RNAse-A digestion

value: postnatal day 0

value: neocortex


title: Sample_1_Aire_DC

value: mesenteric lymph nodes (MLN)

value: three weeks


title: THP-1 ORF LRRC32 rep2

description: THP-1 stable cell line overexpressing LRRC32 gene / replicate 2

value: Monocytic leukemia


title: Riboseq_P0_1

description: ribosome-protected mRNA fragment


title: THP-1 ORF NFKB1 rep1

description: THP-1 stable cell line overexpressing NFKB1 gene / replicate 1


title: THP-1 ORF LRRC32 rep1

description: THP-1 stable cell line overexpressing LRRC32 gene / replicate 1


title: Riboseq_E17_2

description: ribosome-protected mRNA fragment


title: THP-1 ORF JAK2 rep3

description: THP-1 stable cell line overexpressing JAK2 gene / replicate 3


title: 12Z_Control_Input_4

description: MNase digested chromatin from 12Z human endometriotic epithelial cells treated with non-targeting shRNA particles.

data_processing:
Raw reads for IPs and inputs were trimmed with cutadapt (Martin 2011) and Trim Galore! (http://www.bioinformatics.babraham.ac.uk/projects/trim_galore/) followed by quality control analysis via FastQC (Andrews 2010) and MultiQC (Ewels, Magnusson et al. 2016).
Trimmed reads were aligned to GRCh38.p12 reference genome (Schneider, Graves-Lindsay et al. 2017) via Bowtie2 (Langmead and Salzberg 2012) with flag `--very-sensitive`. Aligned reads were sorted and indexed with samtools (Li, Handsaker et al. 2009).
Only properly-paired read fragments were retained for paired-end libraries via samtools view with flag `-f 3` followed by sorting and indexing.
Picard MarkDuplicates (http://broadinstitute.github.io/picard/) was used to remove PCR duplicates, followed by sorting and indexing.
MACS2 (Zhang, Liu et al. 2008) was used to call broadPeaks on each ChIP replicate against the input chromatin control, with FDR < 0.05 threshold and otherwise default settings.
The resulting peaks were repeat-masked by ENCODE blacklist filtering and filtered for non-standard contigs (Amemiya, Kundaje et al. 2019).
A naive overlapping peak set, as defined by ENCODE (Landt, Marinov et al. 2012), was constructed by calling peaks on pooled replicates followed by bedtools intersect (Quinlan and Hall 2010) to select for peaks of at least 50% overlap with each biological replicate.
Genome_build: GRCh38.p12
Supplementary_files_format_and_content: Tabular blacklist-filtered broadPeak files reported by MACS2 corresponding to significant ZMYND8 binding sites in 12Z cells from either or both ChIP replicates.

extract_protocol:
Cells were treated with 1% formaldehyde in growth media for 10 minutes at ambient temperature. Formaldehyde was quenched by the addition of 0.125 M Glycine and incubation for 5 minutes at room temperature, followed by PBS wash and scraping. 1*10^7 crosslinked cells were used for each ChIP, and ChIP was performed in duplicate. Chromatin from crosslinked cells was fractionated by digestion with micrococcal nuclease using the SimpleChIP Enzymatic Chromatin IP Kit (Cell Signaling) according to the manufacturer protocol, followed by 30 seconds of sonication. ChIP was then performed according to the SimpleChIP Enzymatic Chromatin IP Kit (Cell Signaling). 1:250 anti-ZMYND8 (A302-089, Bethyl) was added to each 1.25 mL IP. Crosslinks were reversed with 0.4 mg/mL Proteinase K (ThermoFisher) and 0.2 M NaCl at 65 °C for 2 hours. DNA was purified using the ChIP DNA Clean & Concentrator Kit (Zymo).
Libraries for input and IP samples were prepared by the Van Andel Research Institute Genomics Core. 10 ng of material was used for input samples, and the entire precipitated sample was used for IPs. Libraries were generated using the KAPA Hyper Prep Kit (v5.16) (Kapa Biosystems). Prior to PCR amplification, end-repaired and A-tailed DNA fragments were ligated to IDT for Illumina UDI Adapters (IDT DNA Inc.). Quality and quantity of the finished libraries were assessed using a combination of Agilent DNA High Sensitivity chip (Agilent Technologies), QuantiFluor® dsDNA System (Promega) and Kapa Illumina Library Quantification qPCR assays (Kapa Biosystems). Individually indexed libraries were pooled, and 50 bp, paired-end sequencing was performed on an Illumina NovaSeq 6000 sequencer using a 100-cycle sequencing kit (Illumina). Each library was sequenced to minimum read depth of 80 million reads per input library and 40 million reads per IP library. Base calling was performed by Illumina NCS v2.0, and NCS output was demultiplexed and converted to FastQ format with Illumina Bcl2fastq v1.9.0.

value: Control (non-targeting shRNA)

tag: antibody

value: None (input chromatin)


title: 12Z_Control_non-targeting siRNA_rep1_RNA

description: mRNA (replicate #1) extracted from non-targeting siRNA (control) treated 12Z endometriotic epithelial cells.


title: Riboseq_E17_1

description: ribosome-protected mRNA fragment


title: THP-1 ORF JAK2 rep1

description: THP-1 stable cell line overexpressing JAK2 gene / replicate 1

value: B


title: THP-1 ORF JAK2 rep2

description: THP-1 stable cell line overexpressing JAK2 gene / replicate 2


title: 12Z_Control_non-targeting siRNA_rep2_RNA

description: mRNA (replicate #2) extracted from non-targeting siRNA (control) treated 12Z endometriotic epithelial cells.

value: 12Z endometriotic epithelial cells


title: 12Z_ZMYND8_rep1_ChIP

description: Eluted ZMYND8 ChIP DNA (replicate #1) from wild-type 12Z human endometriotic epithelial cells.

value: Wild-type

value: anti-ZMYND8 (A302-089, Bethyl)


title: Riboseq_E15.5_2

description: ribosome-protected mRNA fragment


title: 12Z_ZMYND8_rep2_ChIP

description: Eluted ZMYND8 ChIP DNA (replicate #2) from wild-type 12Z human endometriotic epithelial cells.


title: THP-1 ORF ITGAL rep2

description: THP-1 stable cell line overexpressing ITGAL gene / replicate 2

value: ITGAL


title: Riboseq_E14_2

description: ribosome-protected mRNA fragment


title: 12Z_shCHD4_Input_3

data_processing:
Raw reads for IPs and inputs were trimmed with cutadapt (Martin 2011) and Trim Galore! (http://www.bioinformatics.babraham.ac.uk/projects/trim_galore/) followed by quality control analysis via FastQC (Andrews 2010) and MultiQC (Ewels, Magnusson et al. 2016).
Trimmed reads were aligned to GRCh38.p12 reference genome (Schneider, Graves-Lindsay et al. 2017) via Bowtie2 (Langmead and Salzberg 2012) with flag `--very-sensitive`. Aligned reads were sorted and indexed with samtools (Li, Handsaker et al. 2009).
Only properly-paired read fragments were retained for paired-end libraries via samtools view with flag `-f 3` followed by sorting and indexing.
Molecular complexity was estimated from duplicate rates by ATACseqQC (Ou, Liu et al. 2018) and preseqR (Daley and Smith 2013), and libraries were subsampled to equivalent molecular complexity within the experimental design based on these estimates with samtools.
Picard MarkDuplicates (http://broadinstitute.github.io/picard/) was used to remove PCR duplicates, followed by sorting and indexing.
MACS2 (Zhang, Liu et al. 2008) was used to call broad peaks on each ChIP replicate against the input chromatin control with FDR < 0.05 threshold and flags `--nomodel --extsize 146` to bypass model building.
The resulting peaks were repeat-masked by ENCODE blacklist filtering and filtered for non-standard contigs (Amemiya, Kundaje et al. 2019).
A naive overlapping peak set, as defined by ENCODE (Landt, Marinov et al. 2012), was constructed by calling peaks on pooled replicates followed by bedtools intersect (Quinlan and Hall 2010) to select for peaks of at least 50% overlap with each biological replicate.
ChIP-seq differential histone abundance analysis was performed with csaw (Lun and Smyth 2016). First, a consensus peak set was constructed from the union of replicate-intersecting, filtered MACS2 peak regions called in each condition. ChIP reads were counted in these query regions by csaw, then filtered for low abundance peaks with average log2CPM > -3. When comparing ChIP libraries, any global differences in IP efficiency observed between the two conditions were considered a result of technical bias to ensure a highly conservative analysis. As such, we employed a loess-based local normalization to the peak count matrix, as is implemented in csaw (Lun and Smyth 2016), to assume a symmetrical MA distribution. A design matrix was then constructed from one ‘condition’ variable, without an intercept term. The count matrix and loess offsets were then supplied to edgeR (Robinson, McCarthy et al. 2010) for estimating dispersions and fitting quasi-likelihood generalized linear models for differential abundance hypothesis testing. Nearby query regions were then merged up to 500 bp apart for a maximum merged region width of 5 kb, and the most significant probability was used to represent the merged region. Finally, FDR < 0.05 threshold was used to define significant differentially abundant regions.
Genome_build: GRCh38.p12
Supplementary_files_format_and_content: Tabular blacklist-filtered broadPeak files reported by MACS2 corresponding to sites of significant H3.3 abundance in 12Z cells from either or both ChIP replicates per condition. Tabular csaw differential H3.3 abundance results tables for shCHD4 vs. control conditions, across all tested genome-wide regions.

extract_protocol:
Cells were treated with 1% formaldehyde in growth media for 10 minutes at ambient temperature. Formaldehyde was quenched by the addition of 0.125 M Glycine and incubation for 5 minutes at room temperature, followed by PBS wash and scraping. 1*10^7 crosslinked cells were used for each ChIP, and ChIP was performed in duplicate. Chromatin from crosslinked cells was fractionated by digestion with micrococcal nuclease using the SimpleChIP Enzymatic Chromatin IP Kit (Cell Signaling) according to the manufacturer protocol, followed by 30 seconds of sonication. ChIP was then performed according to the SimpleChIP Enzymatic Chromatin IP Kit (Cell Signaling) with the addition of 5 mM sodium butyrate to preserve histone acetylation. 1:125 anti-histone H3.3 (2D7-H1, abnova) was added to each, respective 1.25 mL IP. Crosslinks were reversed with 0.4 mg/mL Proteinase K (ThermoFisher) and 0.2 M NaCl at 65 °C for 2 hours. DNA was purified using the ChIP DNA Clean & Concentrator Kit (Zymo).
Libraries for input and IP samples were prepared by the Van Andel Research Institute Genomics Core. 10 ng of material was used for input samples, and the entire precipitated sample was used for IPs. Libraries were generated using the KAPA Hyper Prep Kit (v5.16) (Kapa Biosystems). Prior to PCR amplification, end-repaired and A-tailed DNA fragments were ligated to IDT for Illumina UDI Adapters (IDT DNA Inc.). Quality and quantity of the finished libraries were assessed using a combination of Agilent DNA High Sensitivity chip (Agilent Technologies), QuantiFluor® dsDNA System (Promega) and Kapa Illumina Library Quantification qPCR assays (Kapa Biosystems). Individually indexed libraries were pooled, and 50 bp, paired-end sequencing was performed on an Illumina NovaSeq 6000 sequencer using a 100-cycle sequencing kit (Illumina). Each library was sequenced to minimum read depth of 80 million reads per input library and 40 million reads per IP library. Base calling was performed by Illumina NCS v2.0, and NCS output was demultiplexed and converted to FastQ format with Illumina Bcl2fastq v1.9.0.

treatment_protocol: Lentiviral shRNA particles were transduced into 12Z cells at a 100-fold multiplicity of infection, and media was replaced 24 hours later. Cells were harvested 72 hours following shRNA transduction.

value: shCHD4 (lentiviral CHD4 shRNA particles)


title: Riboseq_E14_1

description: ribosome-protected mRNA fragment


title: THP-1 ORF ITGAL rep1

description: THP-1 stable cell line overexpressing ITGAL gene / replicate 1


title: Riboseq_E12.5_2

description: ribosome-protected mRNA fragment


title: 12Z_shCHD4_H3.3_rep2_ChIP

description: Eluted H3.3 ChIP DNA (replicate #2) from shCHD4 lentiviral particle treated 12Z human endometriotic epithelial cells.

value: anti-histone H3.3 (2D7-H1, abnova)


title: 12Z_Control_Input_3

value: Control (non-targeting shRNA treated)


title: 12Z_shCHD4_H3.3_rep1_ChIP

description: Eluted H3.3 ChIP DNA (replicate #1) from shCHD4 lentiviral particle treated 12Z human endometriotic epithelial cells.


title: THP-1 ORF IRF3 rep3

description: THP-1 stable cell line overexpressing IRF3 gene / replicate 3


title: THP-1 ORF IRF3 rep1

description: THP-1 stable cell line overexpressing IRF3 gene / replicate 1


title: THP-1 ORF IRF3 rep2

description: THP-1 stable cell line overexpressing IRF3 gene / replicate 2


title: 12Z_Control_non-targeting shRNA_H3.3_rep1_ChIP

description: Eluted H3.3 ChIP DNA (replicate #2) from non-targeting shRNA lentiviral particle treated (control) 12Z human endometriotic epithelial cells.


title: C121 nucleus accumbens (ACB)

city: La Jolla

first: Francesca

last: Telese

state: CA

address: 9500 Gilman Drive

department: Psychiatry

institute: UCSD

data_processing:
FastQC: RNA-seq quality control was performed using FastQC
Trimmomatic: Low-quality reads were trimmed by Trimmomatic
STAR: Reads were then aligned to Rattus norvegicus genome Rnor6.0 using STAR
HTSeq: Gene counts were quantified using HTSeq
DEseq2: DEseq2 was then used to perform differential gene expression analysis between control and ACE-K treatment groups across both sexes to identify DEGs affected by ACE-K Treatment, Sex, and Treatment x Sex interactions, or within males and females separately to identify DEGs affected by treatment within each sex
Assembly: Rnor6.0
Supplementary files format and content: csv files including raw counts for each brain region

organism: Rattus norvegicus

source_name: ACB, CTL

extract_protocol:
Total RNA was extracted according to the manufacturer’s instructions using an AllPrep DNA/RNA Mini Kit (Qiagen, Hilden, Germany) and checked for purity using the NanoDrop One (Thermo Fisher Scientific, Waltham, MA, USA).
All samples were found to have high purity and were sent to the USC Genome Core for library preparation and RNA sequencing. There, the total RNA was checked for degradation using a Bioanalyzer 2100 (Agilent, Santa Clara, CA, USA) to verify the quality for all samples. Libraries were prepared from 1 ug of total RNA using a NuGen Universal Plus mRNA-seq Library Prep Kit (Tecan Genomics Inc. Redwood City, CA, USA). Final library products were quantified using the Qubit 2.0 Fluorometer (Thermo Fisher Scientific Inc., Waltham, MA, USA), and the fragment size distribution was determined with the Bioanalyzer 2100. The libraries were pooled equimolarly, and the final pool was quantified via qPCR using the Kapa Biosystems Library Quantification Kit, according to manufacturer’s instructions. The pool was sequenced using an Illumina NextSeq 550 platform (Illumina, San Diego, CA, USA), in Single-Read 75 cycles format, obtaining about 25 million reads per sample.

value: Brain

tag: brain region

value: nucleus accumbens

value: CTL (water)

value: Sprague Dawley

tag: Sex

value: Female

tag: age at_sacrifice

value: Postnatal day (PN) 80

tag: body weight_at_pn_80


title: 12Z_Control_non-targeting shRNA_H3.3_rep2_ChIP

description: Eluted H3.3 ChIP DNA (replicate #1) from non-targeting shRNA lentiviral particle treated (control) 12Z human endometriotic epithelial cells.


title: C122 nucleus accumbens (ACB)

source_name: ACB, ACE-K

tag: tissue

value: Acesulfame potassium (ACE-K)

tag: strain


title: THP-1 ORF GPR35 rep2

description: THP-1 stable cell line overexpressing GPR35 gene / replicate 2

value: Monocytic leukemia


title: C119 nucleus accumbens (ACB)


title: 12Z_shARID1A_Input_2

description: MNase digested chromatin from 12Z human endometriotic epithelial cells treated with shARID1A lentiviral particles.

data_processing:
Raw reads for IPs and inputs were trimmed with cutadapt (Martin 2011) and Trim Galore! (http://www.bioinformatics.babraham.ac.uk/projects/trim_galore/) followed by quality control analysis via FastQC (Andrews 2010) and MultiQC (Ewels, Magnusson et al. 2016).
Trimmed reads were aligned to GRCh38.p12 reference genome (Schneider, Graves-Lindsay et al. 2017) via Bowtie2 (Langmead and Salzberg 2012) with flag `--very-sensitive`. Aligned reads were sorted and indexed with samtools (Li, Handsaker et al. 2009).
Only properly-paired read fragments were retained for paired-end libraries via samtools view with flag `-f 3` followed by sorting and indexing.
Molecular complexity was estimated from duplicate rates by ATACseqQC (Ou, Liu et al. 2018) and preseqR (Daley and Smith 2013), and libraries were subsampled to equivalent molecular complexity within the experimental design based on these estimates with samtools.
Picard MarkDuplicates (http://broadinstitute.github.io/picard/) was used to remove PCR duplicates, followed by sorting and indexing.
MACS2 (Zhang, Liu et al. 2008) was used to call broad peaks on each ChIP replicate against the input chromatin control with FDR < 0.05 threshold and flags `--nomodel --extsize 146` to bypass model building.
The resulting peaks were repeat-masked by ENCODE blacklist filtering and filtered for non-standard contigs (Amemiya, Kundaje et al. 2019).
A naive overlapping peak set, as defined by ENCODE (Landt, Marinov et al. 2012), was constructed by calling peaks on pooled replicates followed by bedtools intersect (Quinlan and Hall 2010) to select for peaks of at least 50% overlap with each biological replicate.
ChIP-seq differential histone abundance analysis was performed with csaw (Lun and Smyth 2016). First, a consensus peak set was constructed from the union of replicate-intersecting, filtered MACS2 peak regions called in each condition. ChIP reads were counted in these query regions by csaw, then filtered for low abundance peaks with average log2CPM > -3. When comparing ChIP libraries, any global differences in IP efficiency observed between the two conditions were considered a result of technical bias to ensure a highly conservative analysis. As such, we employed a loess-based local normalization to the peak count matrix, as is implemented in csaw (Lun and Smyth 2016), to assume a symmetrical MA distribution. A design matrix was then constructed from one ‘condition’ variable, without an intercept term. The count matrix and loess offsets were then supplied to edgeR (Robinson, McCarthy et al. 2010) for estimating dispersions and fitting quasi-likelihood generalized linear models for differential abundance hypothesis testing. Nearby query regions were then merged up to 500 bp apart for a maximum merged region width of 5 kb, and the most significant probability was used to represent the merged region. Finally, FDR < 0.05 threshold was used to define significant differentially abundant regions.
Genome_build: GRCh38.p12
Supplementary_files_format_and_content: Tabular blacklist-filtered broadPeak files reported by MACS2 corresponding to sites of significant H4K16ac abundance in 12Z cells from either or both ChIP replicates per condition. Tabular csaw differential H4K16ac abundance results tables for shARID1A vs. control conditions, across all tested genome-wide regions.

extract_protocol:
Cells were treated with 1% formaldehyde in growth media for 10 minutes at ambient temperature. Formaldehyde was quenched by the addition of 0.125 M Glycine and incubation for 5 minutes at room temperature, followed by PBS wash and scraping. 1*10^7 crosslinked cells were used for each ChIP, and ChIP was performed in duplicate. Chromatin from crosslinked cells was fractionated by digestion with micrococcal nuclease using the SimpleChIP Enzymatic Chromatin IP Kit (Cell Signaling) according to the manufacturer protocol, followed by 30 seconds of sonication. ChIP was then performed according to the SimpleChIP Enzymatic Chromatin IP Kit (Cell Signaling) with the addition of 5 mM sodium butyrate to preserve histone acetylation. 1:125 anti-histone H4K16ac (Active Motif) was added to each, respective 1.25 mL IP. Crosslinks were reversed with 0.4 mg/mL Proteinase K (ThermoFisher) and 0.2 M NaCl at 65 °C for 2 hours. DNA was purified using the ChIP DNA Clean & Concentrator Kit (Zymo).
Libraries for input and IP samples were prepared by the Van Andel Research Institute Genomics Core. 10 ng of material was used for input samples, and the entire precipitated sample was used for IPs. Libraries were generated using the KAPA Hyper Prep Kit (v5.16) (Kapa Biosystems). Prior to PCR amplification, end-repaired and A-tailed DNA fragments were ligated to IDT for Illumina UDI Adapters (IDT DNA Inc.). Quality and quantity of the finished libraries were assessed using a combination of Agilent DNA High Sensitivity chip (Agilent Technologies), QuantiFluor® dsDNA System (Promega) and Kapa Illumina Library Quantification qPCR assays (Kapa Biosystems). Individually indexed libraries were pooled, and 50 bp, paired-end sequencing was performed on an Illumina NovaSeq 6000 sequencer using a 100-cycle sequencing kit (Illumina). Each library was sequenced to minimum read depth of 80 million reads per input library and 40 million reads per IP library. Base calling was performed by Illumina NCS v2.0, and NCS output was demultiplexed and converted to FastQ format with Illumina Bcl2fastq v1.9.0.

tag: treatment/condition

value: shARID1A (lentiviral ARID1A shRNA particles)

value: None (input chromatin)


title: C120 nucleus accumbens (ACB)

tag: treatment


title: Riboseq_E15.5_1

description: ribosome-protected mRNA fragment

value: RNAse-T1 & RNAse-A digestion

tag: developmental stage

value: neocortex


title: THP-1 ORF ITGAL rep3

description: THP-1 stable cell line overexpressing ITGAL gene / replicate 3

tag: cell type

value: B


title: THP-1 ORF GPR35 rep3

description: THP-1 stable cell line overexpressing GPR35 gene / replicate 3

tag: overexpressed gene


title: THP-1 ORF FUT2 rep3

description: THP-1 stable cell line overexpressing FUT2 gene / replicate 3

tag: sampleID


title: 12Z_Control_Input_2

description: MNase digested chromatin from 12Z human endometriotic epithelial cells treated with non-targeting shRNA (control) lentiviral particles.

value: 12Z endometriotic epithelial cells


title: THP-1 ORF GPR35 rep1

description: THP-1 stable cell line overexpressing GPR35 gene / replicate 1

tag: plasmid


title: 12Z_shARID1A_H4K16ac_rep2_ChIP

description: Eluted H4K16ac ChIP DNA (replicate #2) from shARID1A lentiviral particle treated 12Z human endometriotic epithelial cells.

value: anti-histone H4K16ac (Active Motif)


title: THP-1 ORF FUT2 rep1

description: THP-1 stable cell line overexpressing FUT2 gene / replicate 1

tag: infection date


title: C118 nucleus accumbens (ACB)


title: C115 nucleus accumbens (ACB)


title: C116 nucleus accumbens (ACB)


title: THP-1 ORF FUT2 rep2

description: THP-1 stable cell line overexpressing FUT2 gene / replicate 2

tag: days in culture


title: 12Z_Control_H4K16ac_rep2_ChIP

description: Eluted H4K16ac ChIP DNA (replicate #2) from non-targeting shRNA lentiviral particle treated (control) 12Z human endometriotic epithelial cells.


title: 12Z_Control_H4K16ac_rep1_ChIP

description: Eluted H4K16ac ChIP DNA (replicate #1) from non-targeting shRNA lentiviral particle treated (control) 12Z human endometriotic epithelial cells.


title: THP-1 ORF ETS2 rep3

description: THP-1 stable cell line overexpressing ETS2 gene / replicate 3

tag: sequencing batch


title: C114 nucleus accumbens (ACB)


title: 12Z_shARID1A_Input_1

data_processing:
Raw reads for IPs and inputs were trimmed with cutadapt (Martin 2011) and Trim Galore! (http://www.bioinformatics.babraham.ac.uk/projects/trim_galore/) followed by quality control analysis via FastQC (Andrews 2010) and MultiQC (Ewels, Magnusson et al. 2016).
Trimmed reads were aligned to GRCh38.p12 reference genome (Schneider, Graves-Lindsay et al. 2017) via Bowtie2 (Langmead and Salzberg 2012) with flag `--very-sensitive`. Aligned reads were sorted and indexed with samtools (Li, Handsaker et al. 2009).
Only properly-paired read fragments were retained for paired-end libraries via samtools view with flag `-f 3` followed by sorting and indexing.
Molecular complexity was estimated from duplicate rates by ATACseqQC (Ou, Liu et al. 2018) and preseqR (Daley and Smith 2013), and libraries were subsampled to equivalent molecular complexity within an experimental design based on these estimates with samtools.
Picard MarkDuplicates (http://broadinstitute.github.io/picard/) was used to remove PCR duplicates, followed by sorting and indexing.
MACS2 (Zhang, Liu et al. 2008) was used to call peaks on each ChIP replicate against the input chromatin control with FDR < 0.05 threshold. For H3.3, broad peaks were computed with flags `--nomodel --extsize 146` to bypass model building. For H2A.Zac, narrow peaks were computed with flags `--nomodel --extsize 146` to bypass model building.
The resulting peaks were repeat-masked by ENCODE blacklist filtering and filtered for non-standard contigs (Amemiya, Kundaje et al. 2019).
A naive overlapping peak set, as defined by ENCODE (Landt, Marinov et al. 2012), was constructed by calling peaks on pooled replicates followed by bedtools intersect (Quinlan and Hall 2010) to select for peaks of at least 50% overlap with each biological replicate.
ChIP-seq differential histone abundance analysis was performed with csaw (Lun and Smyth 2016). First, a consensus peak set was constructed for each differential experiment from the union of replicate-intersecting, filtered MACS2 peak regions called in each condition. ChIP reads were counted in these query regions by csaw, then filtered for low abundance peaks with average log2CPM > -3. When comparing ChIP libraries, any global differences in IP efficiency observed between the two conditions were considered a result of technical bias to ensure a highly conservative analysis. As such, we employed a loess-based local normalization to the peak count matrix, as is implemented in csaw (Lun and Smyth 2016), to assume a symmetrical MA distribution. A design matrix was then constructed from one ‘condition’ variable, without an intercept term. The count matrix and loess offsets were then supplied to edgeR (Robinson, McCarthy et al. 2010) for estimating dispersions and fitting quasi-likelihood generalized linear models for differential abundance hypothesis testing. Nearby query regions were then merged up to 500 bp apart for a maximum merged region width of 5 kb, and the most significant probability was used to represent the merged region. Finally, FDR < 0.05 threshold was used to define significant differentially abundant regions.
Genome_build: GRCh38.p12
Supplementary_files_format_and_content: Tabular blacklist-filtered broadPeak files reported by MACS2 corresponding to sites of significant H3.3 and H2A.Zac abundance in 12Z cells from either or both ChIP replicates per condition. Tabular csaw differential H3.3 and H2A.Zac abundance results tables for shARID1A vs. control conditions, across all tested genome-wide regions per assay.

extract_protocol:
Cells were treated with 1% formaldehyde in growth media for 10 minutes at ambient temperature. Formaldehyde was quenched by the addition of 0.125 M Glycine and incubation for 5 minutes at room temperature, followed by PBS wash and scraping. 1*10^7 crosslinked cells were used for each ChIP, and ChIP was performed in duplicate. Chromatin from crosslinked cells was fractionated by digestion with micrococcal nuclease using the SimpleChIP Enzymatic Chromatin IP Kit (Cell Signaling) according to the manufacturer protocol, followed by 30 seconds of sonication. ChIP was then performed according to the SimpleChIP Enzymatic Chromatin IP Kit (Cell Signaling) with the addition of 5 mM sodium butyrate to preserve histone acetylation. 1:125 anti-histone H3.3 (2D7-H1, abnova) or 1:50 anti-histone H2A.Z-acetyl (K4/K7) (D3V1I, Cell Signaling) was added to each, respective 1.25 mL IP. Crosslinks were reversed with 0.4 mg/mL Proteinase K (ThermoFisher) and 0.2 M NaCl at 65 °C for 2 hours. DNA was purified using the ChIP DNA Clean & Concentrator Kit (Zymo).
Libraries for input and IP samples were prepared by the Van Andel Research Institute Genomics Core. 10 ng of material was used for input samples, and the entire precipitated sample was used for IPs. Libraries were generated using the KAPA Hyper Prep Kit (v5.16) (Kapa Biosystems). Prior to PCR amplification, end-repaired and A-tailed DNA fragments were ligated to IDT for Illumina UDI Adapters (IDT DNA Inc.). Quality and quantity of the finished libraries were assessed using a combination of Agilent DNA High Sensitivity chip (Agilent Technologies), QuantiFluor® dsDNA System (Promega) and Kapa Illumina Library Quantification qPCR assays (Kapa Biosystems). Individually indexed libraries were pooled, and 50 bp, paired-end sequencing was performed on an Illumina NovaSeq 6000 sequencer using a 100-cycle sequencing kit (Illumina). Each library was sequenced to minimum read depth of 80 million reads per input library and 40 million reads per IP library. Base calling was performed by Illumina NCS v2.0, and NCS output was demultiplexed and converted to FastQ format with Illumina Bcl2fastq v1.9.0.

tag: antibody


title: Riboseq_E12.5_1

description: ribosome-protected mRNA fragment


title: THP-1 ORF ETS2 rep2

description: THP-1 stable cell line overexpressing ETS2 gene / replicate 2


title: 12Z_Control_Input_1


title: C112 nucleus accumbens (ACB)

tag: brain region


title: 12Z_shARID1A_H2A.Zac_rep2_ChIP

description: Eluted H2A.Zac ChIP DNA (replicate #2) from shARID1A lentiviral particle treated 12Z human endometriotic epithelial cells.

value: anti-histone H2A.Z-acetyl (K4/K7) (D3V1I, Cell Signaling)


title: 12Z_shARID1A_H4K16ac_rep1_ChIP

description: Eluted H4K16ac ChIP DNA (replicate #1) from shARID1A lentiviral particle treated 12Z human endometriotic epithelial cells.


title: 12Z_shARID1A_H2A.Zac_rep1_ChIP

description: Eluted H2A.Zac ChIP DNA (replicate #1) from shARID1A lentiviral particle treated 12Z human endometriotic epithelial cells.


title: THP-1 ORF C10orf55 rep2

description: THP-1 stable cell line overexpressing C10orf55 gene / replicate 2


title: 12Z_shARID1A_H3.3_rep2_ChIP

description: Eluted H3.3 ChIP DNA (replicate #2) from shARID1A lentiviral particle treated 12Z human endometriotic epithelial cells.

value: anti-histone H3.3 (2D7-H1, abnova)


title: THP-1 ORF C10orf55 rep1

description: THP-1 stable cell line overexpressing C10orf55 gene / replicate 1


title: C110 nucleus accumbens (ACB)

value: Brain

tag: Sex


title: 12Z_shARID1A_H3.3_rep1_ChIP

description: Eluted H3.3 ChIP DNA (replicate #1) from shARID1A lentiviral particle treated 12Z human endometriotic epithelial cells.


title: C109 nucleus accumbens (ACB)

value: nucleus accumbens

tag: age at_sacrifice


title: C111 nucleus accumbens (ACB)

value: CTL (water)

tag: body weight_at_pn_80


title: CD8 T Cells from IRF8FLPyMT Tumor, scRNAseq

first: Fengshen

last: Kuo

state: New York

address: 1275 York Ave

description: scRNAseq related to Fig 5

library_source: transcriptomic single cell

data_processing:
Illumina bcl2fastq2 version 2.18.0 and 2.20.0.422 were used for generating FASTQ files.
Raw bulk RNA sequencing FASTQ files were aligned against the mm10 assembly by STAR (v020201). Gene level count values were then computed by the summarizeOverlaps function from the R package "GenomicAlignments" with UCSC mm10 KnownGene as the base gene model. The Union counting mode was used and only mapped paired reads were considered. The sample size factor normalized read counts and FPKM (Fragments Per Kilobase Million) values were then computed from gene level counts by using the counts function and the fpkm function, respectively, from the R package DESeq2.
The scRNAseq gene counting data in HDF5 format were made using the Cell Ranger software v3.0.2 (https://support.10xgenomics.com/single-cell-gene-expression/software/pipelines/latest/what-is-cell-ranger) run with input FASTQ files and aligned against mouse transcriptome, mm10-3.0.0, along the expect-cells option to be 1000.
Assembly: mm10
Supplementary files format and content: Bulk RNAseq gene raw count marix is generated individually per sample and is in Comma-separated values (CSV) format with Entrez ID as the row name.
Supplementary files format and content: scRNAseq gene count marix is generated individually per sample through Cell Ranger count option and is in HDF5 format.

source_name: PyMT Tumor

growth_protocol: Mice were aged and tumor growth was monitored, cells were harvested from mice with large tumor burden, approaching 3000 mm3 among all mammary tumors.

extract_protocol:
Around 10,000 FACS sorted live cells (1000 cells/ml suspension, more than 90% viability evaluated by DAPI nuclear stain) were suspended in 1X PBS (calcium and magnesium free) containing 0.04% BSA (A2153, SIGMA). Cell suspension was used as input in 10x chromium controller system (10x Genomics Inc., product code 120223) and cell barcoding was performed by gel beads in emulsion (GEMs) in assembly chip. GEM RT reaction was performed in thermocycler ( 530C for 45 minutes, 850C for 5 minutes, 40C hold overnight). SPRIselect dynabeads (B23318, Beckman Coulter) were used for GEM recovery post RT
2-50ng of DNA was used for target enrichment. cDNA amplification, fragmentation, end repair and A tailing preparation was performed as per manual instructions. High sensitivity DNA chips and Agilent 2100 bioanalyzer (Agilent Technologies) was used for gene expression library profile quality control and quantification at several steps. Quality control was performed twice before sequencing. Two barcoded scRNA samples were pooled together before sequencing

tag: genotype

tag: tissue

value: PyMT Tumor

value: TCRb+CD8+ T cell

value: female


title: THP-1 ORF ATG16L1 rep2

description: THP-1 stable cell line overexpressing ATG16L1 gene / replicate 2


title: THP-1 ORF ATG16L1 rep3

description: THP-1 stable cell line overexpressing ATG16L1 gene / replicate 3


title: CD8 T Cells from MafBCreIRF8FLPyMT Tumor, scRNAseq

description: scRNAseq related to Fig 5


title: 12Z_Control_H2A.Zac_rep1_ChIP

description: Eluted H2A.Zac ChIP DNA (replicate #1) from non-targeting shRNA lentiviral particle treated (control) 12Z human endometriotic epithelial cells.

tag: treatment/condition

value: Control (non-targeting shRNA treated)


title: THP-1 ORF C10orf55 rep3

description: THP-1 stable cell line overexpressing C10orf55 gene / replicate 3

value: Monocytic leukemia


title: C117 nucleus accumbens (ACB)

tag: treatment

tag: strain

value: Sprague Dawley

value: Female

value: Postnatal day (PN) 80


title: C107 nucleus accumbens (ACB)


title: THP-1 ORF ATG16L1 rep1

description: THP-1 stable cell line overexpressing ATG16L1 gene / replicate 1

tag: cell type


title: C113 nucleus accumbens (ACB)


title: C108 nucleus accumbens (ACB)

value: Acesulfame potassium (ACE-K)


title: 12Z_Control_H3.3_rep2_ChIP

description: Eluted H3.3 ChIP DNA (replicate #2) from non-targeting shRNA lentiviral particle treated (control) 12Z human endometriotic epithelial cells.


title: Cell line EO771, +doxy +miR200c, rep3

city: Aurora

first: Tonya

last: Brunetti

state: CO

address: 12800 East 19th Avenue

department: Immunology and Microbiology

institute: University of Colorado Anschutz Medical Campus

data_processing:
Trimming and read cleanup was performed using Cutadapt v2.9 (parameters:  -a AGATCGGAAGAG -A AGATCGGAAGAG -q 30 --minimum-length=10 –pair-filter=any)
Alignment was performed using three different aligners: STAR v2.7.3a (parameters: --outSAMtype BAM SortedByCoordinate, --twopassMode Basic,  --outSAMattributes NH HI NM MD AS nM jM jI XS, --sjdbOverhang 150 ), TopHat v2.1.1 (parameters: --library-type fr-secondstrand), HISAT2 v2.2.1
Alignment correction and cleanup was performed using Opossum v1.0 (parameters: -SoftClipsExist=False --MapCutoff=40 --ProperlyPaired=True)
Variants were called using Platypus v0.8.1 (parameters: --filterDuplicates=0, --minMapQual=0,--minFlank=0, --maxReadLength=500, --minGoodQualBases=10, --minBaseQual=20)
VCF files were normalized using vt v0.5772
SNPs were annotated using variant effect predictor (VEP) against the GRCm38 genome annotation version 102
RNA gene-level and isoform-level quantifications were performed using Salmon v1.3.0 (parameters: --validateMappings)
Assembly: GRCm38
Supplementary files format and content: Tab-delimited text files for *quants.sf files containing various raw and normalized counts data
Supplementary files format and content: Excel files for *potential_mhc_binders_9mers_intron_retentions.xlsx contain large spreadsheets of intron retentions and annotations
Supplementary files format and content: Excel files for *miR200c_and_plusDoxyEmptyVector.xlsx contain large spreadsheets of neoantigens and annotations

growth_protocol: cell lines were cultured in RPMI 1640 Medium supplemented with 5% fetal bovine serum (FBS) and 10U/mL Penicillin-Streptomycin. All cells were tested for mycoplasma contamination using the Barbara Davis Center BioResources Core at the University of Colorado

extract_protocol:
RNA using TRIzol (ThermoFisher, Cat.# 15596026) per manufacturer's instructions followed by a RNeasy MinElute Cleanup (Qiagen, Cat.# 74204) per manufacturer's instructions
PolyA Selected Total RNA Library Kit, prepared according to manufacturer’s instructions

tag: cell line

value: Epithelial-like mammory gland breast carcinoma

value: plus, doxy, miR200c


title: Cell line EO771, +doxy +miR200c, rep2


title: THP-1 ORF ETS2 rep1

description: THP-1 stable cell line overexpressing ETS2 gene / replicate 1

data_processing:
Illumina HiSeq2000 using HiSeq Control Software
Primary QC analysis was performed using FastQC (v0.11.5; https://www.bioinformatics.babraham.ac.uk/projects/fastqc)
Sequence filtering parameters set by Trimmomatic (v.0.36 ) Primer sequences were filtered out with provided TruSeq sequence data and a quality sliding window of Q over 20 for a 40-nucleotide window; only sequences longer than 70 nucleotides were kept in the filtered FASTQ files. Another FastQC run was performed on all filtered FASTQ files to validate Trimmomatic results.
Filtered FASTQ files were aligned to the human genome with STAR (v. 2.5.3a) using indexes built with STAR from H. sapiens GRCh Build 38 files (reference genome and GTF annotation files) provided by Illumina's iGenome repository.
Default values were selected except for the –quantMode flag that was set to TranscriptomeSAM for downstream quantification. Counts per gene for each sequenced sample were obtained using RSEM (v. 1.2.6)
Gene counts were transformed to FPKM as an approximation of gene relative abundance. Genes were filtered to keep only those with FPKM > 0.3 and count>8 in at least two samples.
Small RNAs and LOC genes without annotations were removed. Quality control of samples was performed, excluding strong outlier samples based on principal component analyses
The log-transformed FPKM were then normalized between samples with cyclicloess (normalizeCyclicloess from sva Bioconductor R package).
RNA-seq and cell culture batches effects were removed using ComBat (sva Bioconductor library v3.18.0).
Genome_build: GRCh 38
Supplementary_files_format_and_content: THP1_overexpression_processed_20210506.txt; tab delimited text file; normalized FPKM table

extract_protocol:
Total RNA was extracted from stably transduced THP-1 cultures using the RNeasy Plus Mini kit (Qiagen) according to manufacturer’s protocol. The RNA samples were quantified, and quality controlled using an Agilent RNA 6000 Nano kit (Agilent) on an Agilent Bioanalyzer 2100 system. Samples with RNA Integrity Number (RIN) below 8 were discarded.
RNA samples were transformed into barcoded DNA libraries using TruSeq Stranded mRNA library preparation kits (Illumina), which were then paired-end sequenced, generating 2x100bp reads, using an Illumina HiSeq2000 sequencer

tag: overexpressed gene

tag: sampleID

value: B


title: Cell line EO771, +doxy +miR200c, rep1


title: C106 nucleus accumbens (ACB)

value: Male


title: THP-1 ORF AIRE rep3

description: THP-1 stable cell line overexpressing AIRE gene / replicate 3

value: AIRE

tag: plasmid

tag: infection date


title: Cell line EO771, +doxy empty vector, rep2

value: plus doxy, empty vector


title: THP-1 ORF AIRE rep2

description: THP-1 stable cell line overexpressing AIRE gene / replicate 2

tag: days in culture

tag: sequencing batch


title: THP-1 ORF AIRE rep1

description: THP-1 stable cell line overexpressing AIRE gene / replicate 1


title: SimpleChIP_LVshARID1A_Input_R1.fastq.gz [re-analyzed]

description: Reanalyzed sample GSM4472110; Input library from lentiviral shARID1A infected 12Z cells

data_processing:
Raw reads for IPs and inputs were trimmed with cutadapt (Martin 2011) and Trim Galore! (http://www.bioinformatics.babraham.ac.uk/projects/trim_galore/) followed by quality control analysis via FastQC (Andrews 2010) and MultiQC (Ewels, Magnusson et al. 2016).
Trimmed reads were aligned to GRCh38.p12 reference genome (Schneider, Graves-Lindsay et al. 2017) via Bowtie2 (Langmead and Salzberg 2012) with flag `--very-sensitive`. Aligned reads were sorted and indexed with samtools (Li, Handsaker et al. 2009).
Molecular complexity was estimated from duplicate rates by ATACseqQC (Ou, Liu et al. 2018) and preseqR (Daley and Smith 2013), and libraries were subsampled to equivalent molecular complexity within an experimental design based on these estimates with samtools.
Picard MarkDuplicates (http://broadinstitute.github.io/picard/) was used to remove PCR duplicates, followed by sorting and indexing.
MACS2 (Zhang, Liu et al. 2008) was used to call peaks on each ChIP replicate against the input chromatin control with FDR < 0.05 threshold. For H2A.Z, narrow peaks were computed with flags `--nomodel --extsize 146` to bypass model building. For pan-H4ac, broad peaks were computed with flags `--nomodel --extsize 146` to bypass model building.
The resulting peaks were repeat-masked by ENCODE blacklist filtering and filtered for non-standard contigs (Amemiya, Kundaje et al. 2019).
A naive overlapping peak set, as defined by ENCODE (Landt, Marinov et al. 2012), was constructed by calling peaks on pooled replicates followed by bedtools intersect (Quinlan and Hall 2010) to select for peaks of at least 50% overlap with each biological replicate.
ChIP-seq differential histone abundance analysis was performed with csaw (Lun and Smyth 2016). First, a consensus peak set was constructed for each differential experiment from the union of replicate-intersecting, filtered MACS2 peak regions called in each condition. ChIP reads were counted in these query regions by csaw, then filtered for low abundance peaks with average log2CPM > -3. When comparing ChIP libraries, any global differences in IP efficiency observed between the two conditions were considered a result of technical bias to ensure a highly conservative analysis. As such, we employed a loess-based local normalization to the peak count matrix, as is implemented in csaw (Lun and Smyth 2016), to assume a symmetrical MA distribution. A design matrix was then constructed from one ‘condition’ variable, without an intercept term. The count matrix and loess offsets were then supplied to edgeR (Robinson, McCarthy et al. 2010) for estimating dispersions and fitting quasi-likelihood generalized linear models for differential abundance hypothesis testing. Nearby query regions were then merged up to 500 bp apart for a maximum merged region width of 5 kb, and the most significant probability was used to represent the merged region. Finally, FDR < 0.05 threshold was used to define significant differentially abundant regions.
Genome_build: GRCh38.p12
Supplementary_files_format_and_content: Tabular blacklist-filtered broadPeak files reported by MACS2 corresponding to sites of significant H2A.Z and pan-H4ac abundance in 12Z cells from either or both ChIP replicates per condition. Tabular csaw differential H2A.Z and pan-H4ac abundance results tables for shARID1A vs. control conditions, across all tested genome-wide regions per assay.

source_name: 12Z cell line

extract_protocol:
Cells were treated with 1% formaldehyde in growth media for 10 minutes at ambient temperature. Formaldehyde was quenched by the addition of 0.125 M Glycine and incubation for 5 minutes at room temperature, followed by PBS wash and scraping. 1*10^7 crosslinked cells were used for each ChIP, and ChIP was performed in duplicate. Chromatin from crosslinked cells was fractionated by digestion with micrococcal nuclease using the SimpleChIP Enzymatic Chromatin IP Kit (Cell Signaling) according to the manufacturer protocol, followed by 30 seconds of sonication. ChIP was then performed according to the SimpleChIP Enzymatic Chromatin IP Kit (Cell Signaling) with the addition of 5 mM sodium butyrate to preserve histone acetylation. 1:250 anti-histone H2A.Z (ab4174, abcam) or 1:50 anti-acetyl-histone H4 (06-866, Millipore) was added to each, respective 1.25 mL IP. Crosslinks were reversed with 0.4 mg/mL Proteinase K (ThermoFisher) and 0.2 M NaCl at 65 °C for 2 hours. DNA was purified using the ChIP DNA Clean & Concentrator Kit (Zymo).
Libraries for input and IP samples were prepared by the Van Andel Research Institute Genomics Core. 10 ng of material was used for input samples, and the entire precipitated sample was used for IPs. Libraries were generated using the KAPA Hyper Prep Kit (v5.16) (Kapa Biosystems). Prior to PCR amplification, end-repaired and A-tailed DNA fragments were ligated to IDT for Illumina UDI Adapters (IDT DNA Inc.). Quality and quantity of the finished libraries were assessed using a combination of Agilent DNA High Sensitivity chip (Agilent Technologies), QuantiFluor® dsDNA System (Promega) and Kapa Illumina Library Quantification qPCR assays (Kapa Biosystems). Individually indexed libraries were pooled, and 100 bp, single-end sequencing was performed on an Illumina NovaSeq 6000 sequencer using a 100-cycle sequencing kit (Illumina). Each library was sequenced to minimum read depth of 80 million reads per input library and 40 million reads per IP library. Base calling was performed by Illumina NCS v2.0, and NCS output was demultiplexed and converted to FastQ format with Illumina Bcl2fastq v1.9.0.

value: 12Z cell line


title: C104 nucleus accumbens (ACB)

value: Brain

tag: brain region

value: nucleus accumbens


title: Cell line EO771,  +doxy empty vector, rep1


title: SimpleChIP_LVshNONtg_Input [re-analyzed]

description: Reanalyzed sample GSM4472105; Input library from lentiviral non-targeting shRNA control (LVshNONtg) infected 12Z cells

value: Control


title: iNCC_Sox2_D5_2_hg38

city: Ithaca

first: Marcos

last: Simoes-Costa

address: 526 Campus Rd

department: Molecular Biology and Genetics

institute: Cornell University

data_processing:
Library strategy: CUT&RUN
Demultiplexing, bcl2fastq
Trimming, CutAapt v2.1.0, -a AGATCGGAAGAGCACACGTCTGAACTCCAGTCA -A AGATCGGAAGAGCGTCGTGTAGGGAAAGAGTGT --minimum-length=25 -j 0
Alignment, bowtie2, Group #1 --local --very-sensitive-local --no-unal --no-mixed --no-discordant --x ~/genome/bowtie2-hg38/hg38 -I 10 -X 1000  Group #2 --x ~/genome/bowtie2-GRCg6a/GRCg6a
Marking Duplicates, Picard MarkDuplicates
Filtering, samtools view, sort, index
Peak calling, MACS2 Group#1 callpeak -f BAMPE -g 2913022398  -q 0.05 --call-summits, Group #2 -g 1218492533
Genome_build: Group #1, UCSC hg38
Genome_build: Group #2, ENSEMBL galGal6 (GRCg6a)
Supplementary_files_format_and_content: .narrowPeak, MACS2 peaks
Supplementary_files_format_and_content: .bed, peak file
Supplementary_files_format_and_content: .bam, alignment files

source_name: iNCC induction Day 5

extract_protocol:
Group #1 - Cultures were dissociated using Accutase and counted to normalize cell input. Group #2, cranial neural fold dissections in HH9 stage chicken embryos.
Libraries were cell input normalized and contructed as in Rothstein & Simoes-Costa, Genome Research 2020

tag: time

tag: condition

value: D5 CHIR induced iNCC

tag: group

value: iNCC induction Day 5

tag: antibody

value: Sox2, R&D Systems (AF 2018) 1:50


title: C105 nucleus accumbens (ACB)

tag: tissue

value: CTL (water)

value: Sprague Dawley

tag: Sex

tag: age at_sacrifice

tag: body weight_at_pn_80


title: Cell line EO771, untreated, rep3

tag: cell type

tag: treatment

value: untreated


title: 12Z_shARID1A_panH4ac_rep2_ChIP

description: Eluted pan-H4ac ChIP DNA (replicate #2) from shARID1A lentiviral particle treated (control) 12Z human endometriotic epithelial cells.

value: 12Z endometriotic epithelial cells

tag: treatment/condition

value: shARID1A (lentiviral ARID1A shRNA particles)

value: anti-acetyl-histone H4 (06-866, Millipore)