title: 1014-1_Cord_BPD

value: Peripheral blood

value: BPD

value: F


title: XIAK49.1E_PANCREAS_KPiC_2

status: Public on Jul 06 2023

type: SRA

city: Oxford

first: Xin

last: Lu

state: Oxfordshire

address: Old Road Campus

department: Ludwig Institute for Cancer Research

country: United Kingdom

institute: University of Oxford

library_source: transcriptomic

data_processing:
Sequence reads were trimmed for adaptor sequence/low-quality sequence using Trim Galore version 0.6.4_dev with Cutadapt version 2.9 using Python version 3.8.1
Trimmed sequence reads were mapped to Mus_musculus.GRCm38.dna.primary_assembly.fa and Mus_musculus.GRCm38.84.gtf using both STAR version 2.7.5c and HISAT2 version 2.2.0
Read counts for features were extracted using featureCounts version 2.0.1 from SubRead following alignment by STAR and HISAT2
Assembly: Mus_musculus.GRCm38.dna.primary_assembly.fa
Supplementary files format and content: tab-delimited text files include count values for each sample

molecule: total RNA

organism: Mus musculus

source_name: pancreas

growth_protocol: 10% DMEM, 10% FBS, supplemented with glutamine, pencillin and streptomycin, 5% CO2

extract_protocol:
RNA was harvested using Rneasy mini plus kit (Qiagen)
3' mRNA libraries for RNA-seq were prepared using Lexogen QuantSeq 3’ mRNA-Seq Library Prep Kit FWD following manufacturer's protocols

tag: tissue

value: pancreas

tag: cell line

value: primary

tag: cell type

value: PDAC

tag: genotype


title: 1013-1_Day28_BPD

city: Research Triangle Park

first: Douglas

middle: A.

last: Bell

state: NC

address: PO Box 12233

department: Immunity Inflammation and Disease Lab

country: USA

institute: National Institute of Environmental Health Sciences

data_processing:
75 bp single-end sequencing was performed on NovaSeq 6000 platform using S4 flow cell (Illumina).
Raw sequencing reads in FASTQ files sequenced to 22-150 million reads
Aligned with STAR version 2.6.0c
Gene read counts obtained using featureCounts from the Subread package version 1.5.1 using release 39 of the GENCODE annotation
Assembly: hg38
Supplementary files format and content: counts: 1st column = Gene symbol, 2nd column = Gene read counts

organism: Homo sapiens

source_name: Peripheral blood

extract_protocol:
Total RNAs were isolated from whole blood cell using PAXgene Blood miRNA Kit (PreAnalytix/Qiagen).
Aliquots of RNA (250 ng) were used to generate poly-adenylated RNA libraries with TruSeq Stranded Total RNA Ribo-Zero Human Gold kit (Illumina). Samples were indexed with NEXTfle-96 RNA-Seq Barcodes (Bioo-scientific, Austin, TX).

tag: sample id

value: whole blood cells

tag: variable-disease

tag: variable-time

tag: variable-o2 treatment

value: yes

tag: variable-o2 level

tag: covariate-ga

tag: covariate-bw

tag: covariate-sex

value: M

tag: covariate-b cell%

tag: covariate


title: Chronic Dop 5day 2week withdrawal H1 R2

city: Raleigh

first: Ryan

middle: W

last: Tam

state: North Carolina

address: 840 Main Campus Drive, Building Partners II, Room 3700

department: Chemical and Biomolecular Engineering

institute: North Carolina State University

data_processing:
CLC Workbench 23.0.2
Sequence reads were trimmed for adaptor sequence/low-quality sequence using CLC genomic benchwork (parameter - Quality limit: 0.05)
Trimmed sequence reads were mapped to ensembl v97 hg38 using CLC genomic benchwork (default settings)
Read count extraction and normalization were performed using CLC genomic benchwork.
Differential expression was performed using CLC genomic benchwork using default settings.
Assembly: hg38
Supplementary files format and content: xlsx files include TPM and RPKM normalized values and raw counts for each sample.

growth_protocol: hMSN-like cells were obtained following previously described protocols [15] and have been described in detail in prior work [51]. Briefly, the feeder-independent H9 and H1 hESCs (WA09 and WA01; WiCell, Madison, WI, USA) were maintained in 6-well tissue culture dishes (Greiner Bio-One, Alphen aan den Rijn, NL) coated with 0.5 µg/mL reduced growth factor Matrigel solution (Corning, Durham, NC, USA) in E8 medium (Stemcell Technologies, Cambridge, MA, USA) and passaged using standard protocols. Cells were grown to ~75–80% confluency prior to differentiation. On days in vitro (DIV) 0 of differentiation, E8 medium was removed, cells were washed with 1X PBS (Gibco, Waltham, MA, USA), and media switched to DMEM-F12/Neurobasal media (2:1) (Gibco) supplemented with N2 (Gibco) and B27 minus Vitamin A (Gibco) (together referred to as N2B27). From DIV 0 to 4, cultures were supplemented with SB431542 (10 µM in 95% EtOH; Selleck Chemicals, Houston, TX, USA), LDN-193189 (100 nM in DMSO; BioVision, Waltham, MA, USA), and dorsomorphin (200 nM; BioVision). Media was changed every day. From DIV 5 to 8, cultures were supplemented with just LDN-193189 and dorsomorphin. Media was changed every day. On DIV 9, cultures were washed with 1X PBS, and media switched to N2B27 supplemented with activin A (25 ng/mL in 4 mM HCl; R&D, Minneapolis, MN, USA). Cells were then lifted using cell scrapers (Greiner Bio-One) since EDTA treatment drastically reduced survivability, pipetted up and down 1 time with a 5 mL serological pipette, and replated with 5.21E5 cells/cm2 onto Matrigel-coated 6-well plates. Half-media was changed the next day and then every other day afterward. On DIV 18, cultures were passaged using 0.5 mM EDTA for 2 min at 37 °C, as we no longer observed issues with survivability with EDTA treatment, and pipetted up and down 10–15 times using P1000 mechanical pipette then replated onto Poly-L-Ornithine (15 µg/mL in water; Sigma, St. Louis, MO, USA) and Laminin (5 µg/mL in PBS; Corning) coated 24-well plates (Greiner Bio-One) at 2.11E5 cells/cm2. Half-media was changed the next day and then every other day afterward. From DIV 20 to 24, media was switched to N2B27 with Vitamin A (Gibco). On day 25 for analysis, BDNF and GDNF (10 ng/mL in 0.1% BSA (w/v); Peprotech) were added to aid neuronal maturation and survival (referred to as Maturation Medium). Cells were lifted, similar to the DIV18 passage, on DIV 30 to 35 and all cells were replated onto Poly-L-Ornithine (2 µg/cm2) and Laminin (1 µg/cm2) coated 24-well plates to maintain neuron attachment. Cells were maintained in a humid incubator at 37 °C with 5% CO2. Striatal interneuron-like cells were obtained following a previously published differentiation protocol [55]. H9 and H1 hESCs were grown to ~80-90% confluency prior to differentiation. On days in vitro (DIV) 0 of differentiation, E8 medium was removed, cells were washed with 1X PBS (Gibco, Waltham, MA, USA), and media switched to DMEM-F12/Neurobasal media (2:1) (Gibco) supplemented with N2 (Gibco) and B27 (Gibco) (together referred to as N2B27). From DIV 0 to 9, cultures were supplemented with SB431542 (10 µM in 95% EtOH; Selleck Chemicals, Houston, TX, USA), LDN-193189 (100 nM in DMSO; BioVision, Waltham, MA, USA), and XAV939 (2 µM in DMSO; R&D, Minneapolis, MN, USA). Media was changed every day. On DIV 9, cells were lifted using cell scrapers (Greiner Bio-One) since EDTA treatment drastically reduced survivability, pipetted up and down 1 time with a 5 mL serological pipette, and replated with 5.21E5 cells/cm2 onto Matrigel-coated 6-well plates. From DIV 10 to 20, cultures were supplemented with SHH (200 ng/mL in 0.1% (w/v) BSA in 1X PBS; R&D, Minneapolis, MN, USA) and Purmorphamine (1 µM in DMSO; Sigma, St. Louis, MO, USA). Half-media was changed every other day afterward. On DIV 20, cultures were passaged using 0.5 mM EDTA for 2 min at 37 °C, as we no longer observed issues with survivability with EDTA treatment, and pipetted up and down 10–15 times using P1000 mechanical pipette then replated onto Poly-L-Ornithine (15 µg/mL in water; Sigma, St. Louis, MO, USA) and Laminin (5 µg/mL in PBS; Corning) coated 24-well plates (Greiner Bio-One) at 2.11E5 cells/cm2. Half-media was changed the next day and then every other day afterward. From DIV 20 to 45, media was supplemented with only BDNF (10 ng/mL in 0.1% BSA (w/v); Peprotech) to aid neuronal maturation and survival. Cells were maintained in a humid incubator at 37 °C with 5% CO2.

extract_protocol:
For RNA extraction, total RNA was extracted as previously described [109]. In brief, hMSN-like cultures were washed 1 time in PBS. Total RNA was isolated using Direct-zol RNA MicroPrep Kit (Zymo Research) according to the manufacturer’s protocol. RNA samples were collected in 2 mL RNAse-free tubes and chilled on ice throughout the procedure. Total RNA was then used for RNA-seq.
RNA libraries for RNA-seq were prepared by Genewiz/Azenta.

treatment_protocol: To perform tonic neurotransmitter preincubations hMSN-like and interneuron-like cells were grown to DIV45, half-media was removed, and new media containing either ascorbic acid (100 µM in water; Sigma, St. Louis, MO, USA), dopamine hydrochloride (10 nM in 100 µM ascorbic acid; Sigma, St. Louis, MO, USA), acetylcholine chloride (750 nM in water; Sigma, St. Louis, MO, USA), or L-glutamic acid (2 µM in water; Sigma, St. Louis, MO, USA) was added and left in the wells to be incubated at 37 °C until the next day. This process was repeated every day for 21 days, then RNA extracted for sequencing. To perform antagonist incubations, hMSN-like cells were grown to DIV45, half-media was removed, replaced with media containing both R(+)-SCH23390 hydrochloride (250 µM; Sigma, St. Louis, MO, USA), S(-)-Sulpiride (250 µM; Sigma, St. Louis, MO, USA) was added and incubated at 37 °C for 60 min. Half media was then removed and media containing either dopamine hydrochloride (1 mM in 100 µM ascorbic acid; Sigma, St. Louis, MO, USA), R(+)-SCH23390 hydrochloride (250 µM; Sigma, St. Louis, MO, USA), S(-)-Sulpiride (250 µM; Sigma, St. Louis, MO, USA), or PBS (1X; Gibco), individually or in combination, was added and incubated at 37 °C for 60 min then extracted RNA for sequencing. Chronic incubation proceeded by removing all culture media, replacing with maturation media, and repeating dopamine, SCH23390, and Sulpiride incubations for 5 days. Chronic time-course doses were time-matched to extract all samples on DIV50. To perform acute incubations, hMSN-like cells were grown to DIV45, half-media was removed, replaced with dopamine hydrochloride (10 nM–1 mM in 100 µM ascorbic acid; Sigma, St. Louis, MO, USA), L-glutamic acid (5 µM in water; Sigma, St. Louis, MO, USA), SKF82958 (1–100 µM; Sigma, St. Louis, MO, USA), quinpirole (1–100 µM; Sigma, St. Louis, MO, USA), or CGS21680 (0.1–10 µM; Sigma, St. Louis, MO, USA), and incubated at 37 C for 60 min prior to harvesting for RNA-seq. Withdrawal incubations were performed by dosing as previously described for chronic dopamine for 5 days, then ceasing dopamine dosage for two weeks. A final dopamine dosage was performed after 2 weeks and RNA extracted. All doses were performed at 37 C for 60 min.

value: differentiated human embryonic stem cell

tag: treatment

value: Dopamine 1 mM 1h for 5 days, remove, 2 weeks RNA Extraction


title: XIAK49.1B_PANCREAS_KPiC_3


title: WT_HDAC2

city: Shanghai

first: Jiwei

last: Chen

address: 130 DongAn Road

country: China

institute: Fudan University

library_source: genomic

data_processing:
Raw reads were processed with Trim Galore v0.6.6 (https://www.bioinformatics.babraham.ac.uk/projects/trim_galore/) to remove adaptors and low-quality reads with the parameter “-q 25” and then aligned to the mouse mm9 and human hg19 genome assemblies using Bowtie v2.4.4 with default parameters . All unmapped reads, low mapping quality reads (MAPQ < 30) and PCR duplicates were removed using SAMtools v1.12. The number of spike-in hg19 reads was counted with SAMtools v1.12 and normalization factor alpha = 1e6/hg19_count was calculated. Normalized bigwig was generated with bamCoverage function from deepTools v3.5.1 using scale factors calculated above and reads mapped to the ENCODE blacklist regions were removed using BEDTools v2.30.0 .
Peaks were called using MACS2 v2.2.7.1 with option ‘nomodel’ and peak annotation was performed with R package ChIPseeker v1.28.3.
Assembly: mm9
Supplementary files format and content: bigwig

molecule: genomic DNA

source_name: Embryonic stem cells

growth_protocol: ESCs were grown in N2B27 based serum free medium containing MEK inhibitor PD0325901 and GSK inhibitor CHIR99021 (2i), and LIF (Sigma).

extract_protocol:
ESCs were fixed with 1% formaldehyde (Thermo Fisher) and sheared with E220 focused ultrasonicator (Covaris), respectively. Sheared chromatins were mixed with 20% of lysate from HEK293T cells processed identically as spike-in for normalization. Mixed chromatin were incubated at 4℃ overnight with antibodies. Protein A/G beads (Santa Cruz Biotechnology) were then incubated with the chromatin and antibody mixture for 2 hours at 4℃. Beads were washed, reverse-crosslinked, and DNA were purified with Qiaquick PCR purification kit (Qiagen).
KAPA HyperPrep Kit (Roche) was used to prepare ChIP-Rx libraries.

value: WT

value: Embryonic stem cells

tag: chip antibody


title: Chronic Dop 5day 2week withdrawal H1 R1


title: WT_HDAC1


title: 1013-1_Cord_BPD

value: CordBlood


title: 1013-1_Day14_BPD


title: XIAK42.6B_PANCREAS_KiC_3


title: CGS 1h R3

value: CGS21680 5 microM 1h, RNA Extraction


title: XIAK28.8C_PANCREAS_KiC_1


title: CGS 1h R2


title: LSD1CI_H3K27ac_2


title: WT_H3K27ac_3


title: XIAK28.8B_PANCREAS_KiC_1


title: 1012-1_Day28_BPD


title: XIAK28.7D_PANCREAS_KiC_1


title: 1012-1_Cord_BPD


title: Acute Dop and Antag SCH Sulp 1h H1 R5

value: SCH23390 250 microM, Sulpiride 250 microM 1h, add Dopamine 1 mM 1h, RNA Extraction


title: LSD1CI_H3K4me1_2


title: WT_H3K4me1_3


title: Acute Dop and Antag SCH Sulp 1h H1 R4


title: 1011-1_Day28_nonBPD

value: nonBPD


title: P5, Ctrl pontine nuclei, replicate 2

city: Houston

first: Huda

middle: Yahya

last: Zoghbi

state: TX

address: 1250 Moursund St

department: Molecular and Human Genetics

institute: Baylor College of Medicine

description: 10x Genomics

library_source: transcriptomic single cell

data_processing:
The demultiplexing, barcoded processing, gene counting and aggregation were made using the Cell Ranger software v5.0.1 with default settings.
Assembly: customized genome composed of mouse mm10 reference genome and WPRE sequence
Supplementary files format and content: Tab-separated values files and matrix files

molecule: polyA RNA

source_name: pontine nuclei

extract_protocol:
The brains of embryos and P5 pups were dissected out in ice-cold HEBG media (0.8X B27, 0.25X GlutaMAX in Hibernate E media). For embryonic studies, hindbrains were collected from Atoh1S193A/- and its littermate control (one embryo per genotype, three independent replicates). For P5 study, pontine nuclei were micro-dissected out and pooled from 11-13 pups (two independent replicates). Tissues were cut into small pieces and incubated with Worthington Papain solution at 37°C for 30 min at 800 rpm. At the end of incubation, the samples were transferred to 15 mL falcon tube, followed by gentle trituration with serologic pipette. The cell pellets were collected by centrifuge at 200 rcf for 3 min at 4°C, washed and resuspended in ice-cold sorting buffer (PBS with 0.05% fetal bovine serum). The single-cell resuspension was loaded to 30 µm cell drainer to remove debris, followed by DAPI staining at RT for 5 min. TdTom+DAPI- cells were sorted by Sony SH800S. The cells were pelleted by centrifuge at 200 rcf for 5 min at 4°C and resuspended in sorting buffer to make the final concentration of 1,000 cells/µL.
The cDNA libraries were constructed by 10x Genomics 3’ v3.1 kit following the user guide. Briefly, ~16,500 cells from one sample were mixed with reversed transcription master mix before loaded into Chromium Chip G. GEM containing cells, reversed transcription reagents, and barcoded gel beads were generated by Chromium Controller. The first strand cDNA was amplified, fragmentated, and ligated with sequencing adaptors and sample indices.

value: neurons

value: pontine nuclei

tag: strain

tag: age


title: P5, Ctrl pontine nuclei, replicate 1

description: 10x Genomics

tag: tissue


title: Acute Dop and Antag SCH Sulp 1h H1 R3

tag: cell line

tag: cell type


title: 1011-1_Day14_nonBPD

value: Peripheral blood

tag: sample id

value: F


title: 1011-1_Cord_nonBPD

tag: variable-disease


title: LSD1KO_H3K27ac_2

tag: genotype


title: WT_input_3


title: E18.5, Atoh1S193A/- hindbrain, replicate 3

description: 10x Genomics

source_name: hindbrain

value: hindbrain


title: Acute Antag Sulp Only 1h H1 R3

value: Sulpiride 250 microM 1h, RNA Extraction


title: 1010-1_Day28_nonBPD

value: whole blood cells

tag: variable-time

tag: variable-o2 treatment

value: no


title: WT_H3K27ac_2


title: 1012-1_Day14_BPD

value: BPD

value: yes

tag: variable-o2 level

tag: covariate-ga

value: M


title: Acute Antag Sulp Only 1h H1 R2

value: differentiated human embryonic stem cell


title: E18.5, Ctrl hindbrain, replicate 3

description: 10x Genomics


title: Acute Antag Sulp Only 1h H1 R1


title: WT_H3K4me1_2

value: WT

value: Embryonic stem cells


title: Acute Antag SCH Only 1h H1 R3

value: SCH23390 250 microM 1h, RNA Extraction


title: CGS 1h R1

value: CGS21680 5 microM 1h, RNA Extraction


title: 1010-1_Day14_nonBPD

value: nonBPD

tag: covariate-bw

tag: covariate-sex


title: LSD1KO_H3K4me1_2


title: E18.5, Atoh1S193A/- hindbrain, replicate 1

description: 10x Genomics

value: neurons


title: E18.5, Ctrl hindbrain, replicate 2

description: 10x Genomics


title: Acute Antag SCH Only 1h H1 R2


title: LSD1KO_input_2


title: E18.5, Ctrl hindbrain, replicate 1

description: 10x Genomics


title: 1009-2_Day28_BPD

tag: covariate-b cell%

tag: covariate


title: LSD1CI_input_2

tag: chip antibody


title: 1010-1_Cord_nonBPD

value: CordBlood


title: WT_input_2


title: E14.5, Atoh1S193A/- hindbrain, replicate 3

description: 10x Genomics

tag: strain


title: Acute Antag SCH Only 1h H1 R1

tag: treatment


title: LSD1CI_H3K4me3


title: 1009-2_Cord_BPD

tag: tissue

value: Peripheral blood


title: E18.5, Atoh1S193A/- hindbrain, replicate 2

description: 10x Genomics

tag: age


title: PBS 1h H1 R3

tag: cell line

tag: cell type

value: PBS 1X 1h, RNA Extraction


title: LSD1KO_H3K4me3


title: 1007-1_Day28_BPD

tag: sample id

tag: variable-disease


title: 1007-1_Day14_BPD

tag: variable-time


title: E14.5, Atoh1S193A/- hindbrain, replicate 2

description: 10x Genomics

value: hindbrain


title: WT_H3K4me3

tag: genotype


title: PBS 1h H1 R2


title: Glutamate 1h H1 R3

value: Glutamate 1 mM 1h, RNA Extraction


title: 1007-1_Cord_BPD

value: whole blood cells

tag: variable-o2 treatment

tag: variable-o2 level


title: LSD1CI_H3K4me2


title: E14.5, Atoh1S193A/- hindbrain, replicate 1

description: 10x Genomics


title: 1006-1_Day28_nonBPD

tag: covariate-ga

tag: covariate-bw

value: F


title: PBS 1h H1 R1


title: Glutamate 1h H1 R2


title: 1009-2_Day14_BPD

value: BPD

tag: covariate-sex

tag: covariate-b cell%


title: E14.5, Ctrl hindbrain, replicate 1

description: 10x Genomics


title: LSD1KO_H3K4me2

tag: chip antibody


title: E14.5, Ctrl hindbrain, replicate 2

description: 10x Genomics

tag: strain


title: WT_H3K4me2


title: Mef2d-RV non-Tfh_bio rep 3

city: Seoul

first: YOUN SOO

last: CHOI

address: 103 Daehak-ro Jongro-gu

department: Department of Biomedical Sciences

country: South Korea

institute: Seoul National University College of Medicine

data_processing:
Reads passing Illumina filters were further filtered for those mapping to tRNA, rRNA, adapters, and spike-in controls using Bowtie v1.0.0 (-S --fr)
Filtered reads were against the GRCh38 reference genome and Gencode v27 annotations using STAR v2.6.1 (default options)
Reads mapping to the reference genome are further filtered to remove low-complexity reads with PRINSEQ v0.20.3 (-lc_method "dust" -lc_threshold 4)
Sequencing read coverage per gene was counted using HTSeq-count v0.7.1 (-m union -s yes -t exon -i gene_id) and features with 0 counts across all samples were removed
Assembly: GRCh38.gencode.v27B
Supplementary files format and content: read count matrix with rows as genes and columns as cells

source_name: Mef2d-RV OTII TCRtg CD4 T cells

extract_protocol:
Enriched CD45.1+ GFP OTII CD4 T cells were then stained with CD4, PD-1, PSGL-1, and CXCR5 to sort PSGL-1+CXCR5- non-Tfh cells or PSGL-1loCXCR5+ Tfh cells into Trizol LS (Invitrogen). As previously described, cells were prepared for RNA-seq analysis (88, 89). Briefly, total RNA was extracted from the sorted cells (5,000-250,000) using the miRNAeasy micro kit (Qiagen). Approximately 1 ng of isolated RNA was amplified using the Smart-seq2 protocol. Whole transcriptome cDNA was purified using Ampure XP magnetic bead solution (0.8:1 [vol: vol] ratio, Beckman Coulter). Quality control steps were included in determining (i) fragment size distribution quality by capillary DNA electrophoresis, (ii) quantity by fluorescent Picogreen assay (Invitrogen), and (iii) the optimal number of PCR preamplification cycles for optimal cDNA amplification, and (iv) fragment size selection efficiency. S
Sequencing was performed according to a paired-end strategy using the NovaSeq sequencer (Illumina). Samples that failed quality control standards were eliminated from further analysis21.

value: Mef2d overexpression

tag: stimulation

value: NP-OVA immunization

tag: cell subset

value: PD1-CXCR5- non-Tfh


title: Glutamate 1h H1 R1

value: differentiated human embryonic stem cell

tag: treatment


title: Chronic Dop 24h H1 R3

tag: cell line

value: Dopamine 1 mM 1h for 5 days, remove, 24h RNA Extraction


title: LSD1CI_H3K27ac_1

tag: cell type

value: Embryonic stem cells


title: E14.5, Ctrl hindbrain, replicate 3

description: 10x Genomics

tag: tissue

tag: age


title: 1006-1_Cord_nonBPD

tag: sample id

tag: variable-disease

tag: variable-time

value: yes

tag: covariate


title: Mef2d-RV Tfh_bio rep 3

tag: genotype

value: PD1+CXCR5+ Tfh


title: Chronic Dop 24h H1 R2


title: 1005-1_Day28_nonBPD

value: nonBPD

tag: variable-o2 treatment

tag: variable-o2 level

tag: covariate-ga

tag: covariate-bw


title: LSD1KO_H3K27ac_1

tag: chip antibody


title: Control non-Tfh_bio rep 3

source_name: Empty-RV OTII TCRtg CD4 T cells

value: Control

tag: stimulation


title: WT_H3K27ac_1

value: WT


title: Control Tfh_bio rep 3

tag: cell subset


title: LSD1CI_H3K4me1_1


title: 1001-1_Day28_nonBPD

value: Peripheral blood

value: no

tag: covariate-sex

value: M

tag: covariate-b cell%


title: 1001-1_Cord_nonBPD

tag: cell type

value: whole blood cells

value: CordBlood


title: Chronic Dop 1h H1 R3

tag: cell line

tag: treatment

value: Dopamine 1 mM 1h for 5 days, RNA Extraction


title: LSD1KO_H3K4me1_1


title: BD10_PB_3, eHAP FNLS control, 2 nM PB, replicate 3

city: San Francisco

first: Irene

last: Beusch

state: California

address: 600 16th Street

department: Biochemistry and Biophysics

institute: UCSF

description:
BD10_PB_3
col32

data_processing:
The RNA-seq data was demultiplexed and trimmed (BBMap BBDuk v.38.94) for TruSeq adapters and polyA.
if using UMI from lexogen kit later do: [umi_tools extract --extract-method=regex --bc-pattern=\'(?P<umi_1>.{10})(?P<discard_1>..).*\']
sequencing reads were then mapped using STAR 2.7.915 [--alignSJoverhangMin 8 --alignSJDBoverhangMin 1 --outFilterMismatchNmax 999 --alignIntronMin 20 --alignIntronMax 1000000 --alignMatesGapMax 1000000 --peOverlapNbasesMin 10 --peOverlapMMp 0.2] and SAMtools (v1.10) for conversion to sorted bam files.
Reads can then be deduplicated using the UMIs from the Lexogen CORALL workflow to prevent removal of natural duplicates if desired.
Assembly: hg38
Supplementary files format and content: tab-delimited file containing coutns per transcript obtained with Rsubreads.

source_name: eHAP FNLS

growth_protocol: All cell lines were maintained at 37 °C with 5% CO2 and were regularly tested negative for mycoplasma infection. Cells were passaged every 2-3 days.  Human eHAP cell lines and derivatives thereof were cultured in IMDM (with L-glutamine, with HEPES; Cytiva #SH30228), supplemented with 10% FBS and 1:100 penicillin/streptomycin. eHAP cell lines were at all times maintained at sub-confluent conditions and ploidy was regularly assessed with flow cytometry.

extract_protocol:
Total RNA was extracted using the RNAqueousTM-96 Total RNA Isolation Kit (ThermoFisher #AM1920) according to the manufacturer’s protocol.
Poly(A)-enriched RNA was obtained with the poly(A) RNA Selection Kit V1.5 (Lexogen #157.96) and RNA-seq libraries were generated using the CORALL Total RNA-Seq Library Prep Kit (Lexogen #117.96). The libraries were then sequenced using paired-end 150 bp reads with 60 million reads per sample on Nova Seq (S4).

treatment_protocol: eHAP FNLS and mutant cells were treated for 3 h with DMSO or 2 nM PB before cells were harvested. This was done for three independent monoclonal cell lines (mutants) or three independent replicates of an eHAP FNLS cell line transduced with a non-targeting sgRNA (control).

value: eHAP FNLS

value: control

value: sgRNA non-targeting, 2 nM PB 3h


title: WT_H3K4me1_1


title: 1006-1_Day14_nonBPD

tag: tissue

tag: sample id

value: F


title: Chronic Dop 1h H1 R1

value: differentiated human embryonic stem cell


title: Control non-Tfh_bio rep 2

value: NP-OVA immunization

value: PD1-CXCR5- non-Tfh


title: Acute Dop 24h H1 R3

value: Dopamine 1 mM 1h, remove, 24h RNA Extraction


title: Mef2d-RV non-Tfh_bio rep 2

value: Mef2d overexpression


title: BD10_PB_2, eHAP FNLS control, 2 nM PB, replicate 2

description:
BD10_PB_2
col31


title: LSD1CI_input_1

value: Embryonic stem cells


title: Control Tfh_bio rep 2

value: Control

value: PD1+CXCR5+ Tfh


title: Acute Dop 24h H1 R2


title: BD10_PB_1, eHAP FNLS control, 2 nM PB, replicate 1

description:
BD10_PB_1
col30


title: Chronic Dop 24h H1 R1

value: Dopamine 1 mM 1h for 5 days, remove, 24h RNA Extraction


title: 1005-1_Cord_nonBPD

value: Peripheral blood

tag: variable-disease

value: nonBPD

tag: variable-time

value: yes


title: Mef2d-RV Tfh_bio rep 2

tag: genotype


title: Mef2d-RV non-Tfh_bio rep 1


title: LSD1KO_input_1


title: WT_input_1

value: WT


title: Acute Dop 24h H1 R1


title: BD10_DMSO_3, eHAP FNLS control, DMSO, replicate 3

description:
BD10_DMSO_3
col29

value: eHAP FNLS

value: sgRNA non-targeting, DMSO 3h


title: Acute Dop 1h H1 R3

value: Dopamine 1 mM 1h, RNA Extraction


title: Mef2d-RV Tfh_bio rep 1


title: Chronic Dop 1h H1 R2

value: Dopamine 1 mM 1h for 5 days, RNA Extraction


title: BD10_DMSO_2, eHAP FNLS control, DMSO, replicate 2

description:
BD10_DMSO_2
col28

value: control


title: Control non-Tfh_bio rep 1

value: NP-OVA immunization


title: BD10_DMSO_1, eHAP FNLS control, DMSO, replicate 1

description:
BD10_DMSO_1
col27


title: postnatal day 5, scRNA-seq

city: Charlottesville

first: Jason

middle: Paul

last: Smith

state: VA

address: 409 Lane Rd

department: Pediatrics

institute: University of Virginia

description: 10x Genomics

data_processing:
The demultiplexing, barcoded processing, gene counting and aggregation were made using the Cell Ranger software v7.0.1
Assembly: refdata-gex-mm10-2020-A
Supplementary files format and content: Tab-separated values files and matrix files

source_name: kidney

extract_protocol:
Pregnant mice were injected with tribromoethanol at enough dose to keep alive but anesthetized. Then, the small fetuses were removed one by one. A small squash from the lung was used to identify the GFP+ pups at E12, using an EvosFLC cell imaging system (LifeTechnologiesTM, California, USA). Once they were identified, the metanephros area was removed under microscopy and harvested in ice cold dPBS. The tissue was then minced carefully with a razor blade and transferred to 1.7mL Eppendorf tube. 300 μL of TryPLETM Express (Gibco, New York, USA) was added and incubated at 37∘C for 5 minutes. Then, 600 μL of DMEM + 5% FBS was added to the Eppendorf tube. The mixture was homogenized by pipetting up and down. The solution was then filtered with a 40 μm nylon cell strainer. Tissue chunks were removed from the mesh surface, placed again in an Eppendorf tube and the process repeated a second time. Meanwhile, the flow-through filtrate was placed in a new Eppendorf tube and centrifuged at 4∘C, 150g for 5 minutes. The supernatant was removed and the pellet resuspended with resuspension buffer. All the tubes were combined in the end and the cells were ready for single-cell capture. For later timpoints, animals were anesthetized with tribromoethanol (300 mg/kg). P5 and P30 mice kidneys were excised and decapsulated. Then, the kidney cortices were dissected, minced with a razor blade, and transferred into a 15 mL tube with 5 mL of enzymatic solution (0.3% collagenase A [Millipore-Sigma], 0.25% trypsin [Millipore-Sigma], and 0.0021% DNase I [Millipore-Sigma]). The tubes were placed inside a shaking incubator (80 RPM) for 15 minutes at 37∘C. The solution was pipetted up/down 10 times with a sterile transfer pipette and allowed to settle for 2 minutes, and the supernatant was collected in a fresh tube on ice. The enzymatic solution was added to the 15 mL tube containing the remaining undigested cortices, and the digestion procedure was repeated a total of 3 times. The supernatants were pooled and centrifuged at 800 g for 4 minutes at 4∘C using a Sorvall RT7 refrigerated centrifuge (Sorvall, Newtown, CT). The cell pellet was resuspended in fresh buffer 1 (130 mM NaCl, 5 mM KCl, 2 mM CaCl2, 10 mM glucose, 20 mM sucrose, 10 mM HEPES, pH 7.4), and the suspension was poured through a sterile 100 μm nylon cell strainer (Corning Inc., Corning, NY) and washed with buffer 1. The flow-through was poured through a sterile 40 μm nylon cell strainer (Corning Inc.) and washed with buffer 1. The flow-through was centrifuged at 1,100 g for 4 min at 4∘C. The cell pellet was resuspended in 1.5 mL of resuspension buffer [PBS, 1% FBS, 1 mM EDTA, DNAase I (Millipore-Sigma)]. DAPI (Millipore-Sigma) was added to the cells to identify the living cells. The GFP positive cells were collected by Fluorescent-Activated Cell Sorting (FACS) and resuspended in DMEM (Dulbecco’s Modified Eagle Medium, Gibco, Netherlands) with 10% FBS (Fetal Bovine Serum) for immediate use. The sorters were either an Influx Cell Sorter (Becton Dickinson, Franklin Lakes, NJ) or a FACS Aria Fusion Cell Sorter (Becton Dickinson), both located at the Flow Cytometry Core Facility at the University of Virginia.
Sorted GFP-positive cells were spun down at 500g for 10 minutes in a Sorvall RT7 refrigerated 4∘C centrifuge (Sorvall, Newtown, CT). Then, the supernatant was carefully removed, and the cell pellet was resuspended in dPBS (Dulbecco’s Phosphate Buffered Saline, Gibco, UK) with 0.04% BSA (UltraPureTM Bovine Serum Albumin, Invitrogen, Lithuania) 10 times with wide-bore tips. The process was repeated once, and the cells were counted with the CellometerMini (Nexcelon Bioscience, Massachussets, USA). Experiments with more than 500 cells/μL, viability higher than 85%, and absence of clumps were chosen to proceed. Single cells were loaded and captured with the Chromium System (10X Genomics, Pleasanton, CA) following the manufacturer’s recommendation using the Chromium Next GEM Chip G with reagents of Chromium Next GEM Single Cell 3' Reagent Kits v3.1 (10X Genomics). The GEMs were generated and incubated to generate the barcoded cDNA. cDNA was cleaned with Dynabeads and amplified by PCR. cDNA was then enzymatically fragmented, end-repaired, poly-A tailed, adaptor-ligated, and amplificated by PCR. The constructed libraries were sequenced on an Illumina HiSeq 2500/4000 platform (150-bp paired-end reads).

value: FoxD1+ cells

value: kidney

tag: strain

tag: age

value: 5 day old pups


title: Control Tfh_bio rep 1


title: embryonic day 18, scRNA-seq

description: 10x Genomics

value: 18 day old embryos


title: Acute Dop 1h H1 R2

value: differentiated human embryonic stem cell


title: Acute Dop 1h H1 R1


title: B05_PB_3, eHAP FNLS SF3B1 T1080I, 2 nM PB, replicate 3

description:
B05_PB_3
col26

value: sgRNA SF3B1_166, 2 nM PB 3h


title: B05_PB_2, eHAP FNLS SF3B1 T1080I, 2 nM PB, replicate 2

description:
B05_PB_2
col25


title: embryonic day 12, scRNA-seq

description: 10x Genomics

value: 12 day old embryos


title: DGA pre Dop H1 R3

value: Acetylcholine 750 nM, Dopamine 10 nM, Glutamate 2 microM 3 weeks, Dopamine 1 mM 1h, RNA Extraction


title: Node/left-right organizer

city: Indianapolis

first: Stephanie

last: Ware

state: IN

address: 1044 W Walnut Street, R4 W227

institute: Indiana University School of Medicine

data_processing:
Bam files were created using the TopHat pipeline.
The differential gene and transcript expression analysis was performed using RNA-seq workflow in Partek Genomics Suite software, version 6.6
Assembly: mm10
Supplementary files format and content: Excel file with RPKM values for left-right organizer and non-organiser cells

source_name: whole embryo sorted left-right organizer cells

extract_protocol:
RNA was harvested using RNeasy Micro kit protocol (Qiagen).
Ovation RNA-Seq System v2 (NuGEN, Tecan Group Ltd., Männedorf, Switzerland) was used to create double stranded cDNA from total RNA. The cDNA library was prepared with the Nextera DNA Sample Preparation kit (Illumina).

value: C57BL/6 x 129 SvEv

value: left-right organizer

tag: developmental stage

value: E.8 (0-4 somites) stage embryos

tag: gfp status

value: GFP positive


title: DGA pre Dop H1 R2


title: embryonic day 12, scATAC-seq

description: 10x Genomics

library_source: genomic single cell

data_processing:
The demultiplexing, barcoded processing, region counting and aggregation were made using the Cell Ranger ATAC software v2.1.0
Assembly: cellranger-arc-mm10-2020-A-2.0.0
Supplementary files format and content: fragments.tsv files

extract_protocol:
Pregnant mice were injected with tribromoethanol at enough dose to keep alive but anesthetized. Then, the small fetuses were removed one by one. A small squash from the lung was used to identify the GFP+ pups at E12, using an EvosFLC cell imaging system (LifeTechnologiesTM, California, USA). Once they were identified, the metanephros area was removed under microscopy and harvested in ice cold dPBS. The tissue was then minced carefully with a razor blade and transferred to 1.7mL Eppendorf tube. 300 μL of TryPLETM Express (Gibco, New York, USA) was added and incubated at 37∘C for 5 minutes. Then, 600 μL of DMEM + 5% FBS was added to the Eppendorf tube. The mixture was homogenized by pipetting up and down. The solution was then filtered with a 40 μm nylon cell strainer. Tissue chunks were removed from the mesh surface, placed again in an Eppendorf tube and the process repeated a second time. Meanwhile, the flow-through filtrate was placed in a new Eppendorf tube and centrifuged at 4∘C, 150g for 5 minutes. The supernatant was removed and the pellet resuspended with resuspension buffer. All the tubes were combined in the end and the cells were ready for single-cell capture. For later timpoints, animals were anesthetized with tribromoethanol (300 mg/kg). P5 and P30 mice kidneys were excised and decapsulated. Then, the kidney cortices were dissected, minced with a razor blade, and transferred into a 15 mL tube with 5 mL of enzymatic solution (0.3% collagenase A [Millipore-Sigma], 0.25% trypsin [Millipore-Sigma], and 0.0021% DNase I [Millipore-Sigma]). The tubes were placed inside a shaking incubator (80 RPM) for 15 minutes at 37∘C. The solution was pipetted up/down 10 times with a sterile transfer pipette and allowed to settle for 2 minutes, and the supernatant was collected in a fresh tube on ice. The enzymatic solution was added to the 15 mL tube containing the remaining undigested cortices, and the digestion procedure was repeated a total of 3 times. The supernatants were pooled and centrifuged at 800 g for 4 minutes at 4∘C using a Sorvall RT7 refrigerated centrifuge (Sorvall, Newtown, CT). The cell pellet was resuspended in fresh buffer 1 (130 mM NaCl, 5 mM KCl, 2 mM CaCl2, 10 mM glucose, 20 mM sucrose, 10 mM HEPES, pH 7.4), and the suspension was poured through a sterile 100 μm nylon cell strainer (Corning Inc., Corning, NY) and washed with buffer 1. The flow-through was poured through a sterile 40 μm nylon cell strainer (Corning Inc.) and washed with buffer 1. The flow-through was centrifuged at 1,100 g for 4 min at 4∘C. The cell pellet was resuspended in 1.5 mL of resuspension buffer [PBS, 1% FBS, 1 mM EDTA, DNAase I (Millipore-Sigma)]. DAPI (Millipore-Sigma) was added to the cells to identify the living cells. The GFP positive cells were collected by Fluorescent-Activated Cell Sorting (FACS) and resuspended in DMEM (Dulbecco’s Modified Eagle Medium, Gibco, Netherlands) with 10% FBS (Fetal Bovine Serum) for immediate use. The sorters were either an Influx Cell Sorter (Becton Dickinson, Franklin Lakes, NJ) or a FACS Aria Fusion Cell Sorter (Becton Dickinson), both located at the Flow Cytometry Core Facility at the University of Virginia.
GFP-positive cells were washed in dPBS with 0.04% BSA twice, and the sorted GFP-positive cells were washed in dPBS with 0.04% BSA twice. The cells were counted with the CellometerMini (Nexcelom Bioscience, Massachussets, USA). Sorted cells with viability higher than 80% and absence of clumps were chosen to proceed. Experiments yielded less than 100,000 cells. Therefore, we used the protocol designed for Low Cell Input Nuclei Isolation. Briefly, we centrifuged the cells at 500g at 4∘C for 10 minutes, removed the supernatant carefully and resuspended the cells in a freshly prepared lysis buffer (1M Tris-HCl, 5M NaCl, 1M MgCl2, 10% BSA, 10% Tween-20). We optimized the lysis incubation time for 7 minutes on ice. Next, we immediately added freshly made washing buffer (1M Tris-HCl, 5M NaCl, 1M MgCl2, 10% Tween-20, 10% Nonidet P40, 5% digitonin and 10% BSA). This washing was performed twice, and then supernatant was removed for a final wash with diluted nuclei buffer (nuclei buffer 20X, 10X Genomics, diluted in nuclease free water to 1X). Once this was completed, we again removed the supernatant, resuspended the nuclei pellet and counted them with a Neubauer Chamber (Spencer, Buffalo, USA). In all our experiments we were able to target between 1000-5000 nuclei. Nuclei were loaded and captured using the Chromium System (10X Genomics, Pleasanton, CA) following the manufacturer’s recommendation100 using the Chromium Next GEM (Cell-Gel Beads in Emulsion) Chip H with reagents of Chromium Next GEM Reagent Kits v1.1 (10X Genomics). Initially, nuclei were incubated with the transposition mix, that includes a transposase, and later the GEMs were barcoded and PCR amplified to generate the cDNA. The cDNA was then cleaned with Dynabeads and SPRIselect, end-repaired, adaptor-ligated and amplificated by PCR. The constructed libraries were sequenced on an Illumina HiSeq 2500/4000 platform (150-bp paired-end reads).


title: postnatal day 5, scATAC-seq

description: 10x Genomics


title: B05_PB_1, eHAP FNLS SF3B1 T1080I, 2 nM PB, replicate 1

description:
B05_PB_1
col24

value: eHAP FNLS


title: postnatal day 30, scATAC-seq

description: 10x Genomics

value: 30 day old pups


title: Non-organiser cells

source_name: whole embryo sorted non-organiser cells

value: non-organiser cells

value: GFP negative


title: 1383D2-day13+14

city: Kyoto

first: Ryuichiro

last: Hirayama

address: 53 Kawahara-cho

country: Japan

institute: Kyoto University

data_processing:
STAR (version 2.7.3a)
featureCounts (version2.0.1)
RSEM (version 1.3.1)
Assembly: GRCh38
Supplementary files format and content: tab-delimited text files include FPKM values for each sample.

extract_protocol:
Total RNA were extracted using RNeasy according to the manufacture's protocol.
For RNA sequencing, total RNA was extracted as described above. The samples preserved at −80°C were shipped and analyzed by DNAFORM. The quality of total RNA was evaluated by a Bioanalyzer (Agilent) to ensure over 8.0 RIN (RNA integrity number) or electrophoresis waveform. Double-stranded cDNA libraries (RNA-seq libraries) were prepared using a SMART Seq Stranded Kit (Clontech) according to the manufacturer’s protocols. RNA-seq libraries were sequenced using paired-end reads (50 nt of read 1 and 25 nt of read 2) on a NextSeq 500 (Illumina). Obtained reads were mapped to the human GRCh38 genome analyzed by STAR (version 2.7.3a). Annotated reads were counted using featureCounts (version2.0.1) and RSEM (version 1.3.1). FPKM values were calculated from mapped reads by normalizing to total counts.

value: kidney organoid

tag: time point


title: 1383D2-day13+28


title: 1383D2-day13+7


title: DGA pre Dop H1 R1


title: B05_DMSO_1, eHAP FNLS SF3B1 T1080I, DMSO, replicate 1

description:
B05_DMSO_1
col21

value: sgRNA SF3B1_166, DMSO 3h


title: DGA pre AA H1 R1

value: Acetylcholine 750 nM, Dopamine 10 nM, Glutamate 2 microM 3 weeks, Ascorbic Acid 100 microM 1h, RNA Extraction


title: DGA pre AA H1 R2


title: B05_DMSO_3, eHAP FNLS SF3B1 T1080I, DMSO, replicate 3

description:
B05_DMSO_3
col23


title: ATAC_seq_TCF1KO_LSK_P45_DN2_Rep2B_Rep3T

city: Philadelphia

first: Naomi

last: Goldman

state: PA

address: 324 BRB II/III, 421 Curie Blvd

institute: University of Pennsylvania

data_processing:
ATAC-seq libraries were pair-end sequenced by Illumina NextSeq 500. BWA was used for alignment. Reads aligned to the mitochondrial genome or chrY as well as reads mapping to multiple genomic loci were discarded from downstream analyses. Additionally, Picard was used to mark and remove duplicates. Furthermore, for each ATAC-seq library the insert size was calculated by Picard.
Assembly: GRCm38
Supplementary files format and content: Bigwig files were generated by bedtools genomecov and wigToBigWig normalizing tracks to tags-per-million

source_name: BM derived CD44+ CD25+ DN2

extract_protocol:
ATAC-seq was performed as previously described with minor modifications (Buenrostro et al., 2013). Fifty thousand cells were pelleted at 550 x g and washed with 1 mL 1x PBS, followed by treatment with 50 μL lysis buffer (10 mM Tris-HCl [pH 7.4], 10 mM NaCl, 3 mM MgCl2, 0.1% IGEPAL CA-630). After pelleting nuclei, the pellets were resuspended in 50 μL transposition reaction with 2.5 μL Tn5 transposase (FC-121-1030; Illumina). The reaction was incubated in a 37°C water bath for 45 minutes. Tagmented DNA was purified using a MinElute Reaction Cleanup Kit (Qiagen) and amplified with varying cycles, depending on the side reaction results. Libraries were purified using a QIAQuick PCR Purification Kit (Qiagen).
Libraries were paired-end sequenced (38bp+38bp) on a NextSeq 550 (Illumina). Libraries were validated for quality and size distribution using a TapeStation 2200 (Agilent).

treatment_protocol: Transduction of Tcf7 KO scid.adh cells was accomplished by addition of retroviral supernatants to culture medium supplemented with polybrene (8mg/ml) and spinfected at 1,750 RPM for 25 minute. 72hrs after transduction, live transduced cells were sorted for downstream experiments. Cells from co-cultures were stained with L/D APCef780, and fluorescent antibodies to B220, CD44, CD45, Thy1.2, Ckit, CD25, and CD11b. Sorting was performed on a BD FACSAria after 7 days to isolate DN1 (CD45+c-KithiCD44hI CD25low) and DN2 cells (CD45+c-KitlowCD44low CD25high).

value: Tcf7eGFP fl/fl Cre+

value: Cultured on OP9-DLL1


title: 1383D2-day13+4


title: A09_PB_3, eHAP FNLS SUGP1 G603N, 2 nM PB, replicate 3

description:
A09_PB_3
col20

value: clone A09_3.04, SUGP1 G603N

value: sgRNA SUGP1_188, 2 nM PB 3h


title: A09_PB_2, eHAP FNLS SUGP1 G603N, 2 nM PB, replicate 2

description:
A09_PB_2
col19

value: clone A09_3.20, SUGP1 G603N


title: Glut pre Dop H1 R3

value: differentiated human embryonic stem cell

value: Glutamate 2 microM 3 weeks, Dopamine 1 mM 1h, RNA Extraction


title: HBEC3 cells, R/Y DKO2, H3K27me3

city: Grand Rapids

first: Gerd

last: Pfeifer

state: MI

address: 333 Bostwick Ave. NE

institute: Van Andel Research Institute

data_processing:
75-bp single-end reads were trimmed with Trim Galore (version 0.4.0) with default parameters, then the trimmed reads were mapped to human genome hg19 with Bowtie2(version 2.3.5)
The aligned reads were deduplicated with the software picard(version 2.19.0)
Then the deduplicated reads were used for peak calling by HOMER (version 4.10) with the following settings: -B -region -size 1000 -minDist 2500 -P 0.01 -F 2.0 -L 2.0 -LP 0.01.
Output bedgraph files from HOMER were processed into bigwig files with UCSC Exe Utilities bedGraphToBigWig(version 1.04.00)
Assembly: hg19
Supplementary files format and content: bigWig, counts coverage

growth_protocol: keratinocyte SFM medium , Cells were cultured at 37C at 5% CO2.

extract_protocol:
For RYBP ChIP, cells were fixed with 2.5 mM EGS (ethylene glycol-bis-succinimidyl-succinate) for 45 minutes, then fixed with 1% formaldehyde for 8 minutes; for H2AK119ub1 and H3K27me3, cells were crosslinked with 1% formaldehyde only for 8 minutes, after quenching and cell lysis, chromatin was sonicated using a Covaris E220 Evo sonicator.
TruSeq ChIP Sample Preparation Kit (Illumina, IP-202-1012, IP-202-1024) was used according to the manufacturer’s instructions to perform ChIP-seq library preparation. 5 ng of DNA was used as starting material for input and IP samples. Libraries were amplified using 14 cycles on the thermocycler. Libraries were validated using the Agilent High Sensitivity DNA Kit.

value: Human Bronchial cells

value: RYBP and YAF2 double knockout

tag: chip antibody

value: Tri-Methyl-Histone H3 (Lys27) (CST, 9733)


title: Glut pre Dop H1 R2


title: ATAC_seq_TCF1KO_LSK_P45_DN2_Rep2B_Rep1T


title: ATAC_seq_TCF1KO_LSK_P45_DN2_Rep2B_Rep2T


title: HBEC3 cells, R/Y DKO1, H3K27me3


title: A09_PB_1, eHAP FNLS SUGP1 G603N, 2 nM PB, replicate 1

description:
A09_PB_1
col18

value: eHAP FNLS

value: clone A09_3.08, SUGP1 G603N


title: 1383D2-day13+21


title: DGA pre AA H1 R3


title: B05_DMSO_2, eHAP FNLS SF3B1 T1080I, DMSO, replicate 2

description:
B05_DMSO_2
col22


title: embryonic day 18, scATAC-seq

description: 10x Genomics

value: FoxD1+ cells

value: kidney

value: 18 day old embryos


title: Glut pre Dop H1 R1


title: ATAC_seq_TCF1KO_LSK_P45_DN2_Rep1B_Rep3T


title: A09_DMSO_3, eHAP FNLS SUGP1 G603N, DMSO, replicate 3

description:
A09_DMSO_3
col17

value: sgRNA SUGP1_188, DMSO 3h


title: Glut pre AA H1 R3

value: Glutamate 2 microM 3 weeks, Ascorbic Acid 100 microM 1h, RNA Extraction


title: ATAC_seq_TCF1KO_LSK_P45_DN2_Rep1B_Rep2T

value: Tcf7eGFP fl/fl Cre+


title: HBEC3 cells, YKO1, H3K27me3

value: YAF2 knockout


title: A09_DMSO_2, eHAP FNLS SUGP1 G603N, DMSO, replicate 2

description:
A09_DMSO_2
col16


title: HBEC3 cells, YKO2, H3K27me3


title: ATAC_seq_TCF1KO_LSK_P45_DN2_Rep1B_Rep1T

value: Cultured on OP9-DLL1


title: A09_DMSO_1, eHAP FNLS SUGP1 G603N, DMSO, replicate 1

description:
A09_DMSO_1
col15


title: HBEC3 cells, RKO2,  H3K27me3

value: RYBP knockout


title: ATAC_seq_TCF1KO_LSK_P45_DN1_Rep2B_Rep3T

source_name: BM derived CD44+ CD25- DN1


title: Glut pre AA H1 R2


title: A08_PB_3, eHAP FNLS SUGP1 E554K, 2 nM PB, replicate 3

description:
A08_PB_3
col14

value: clone A08_A03, SUGP1 E554K

value: sgRNA SUGP1_238, 2 nM PB 3h


title: HBEC3 cells, Scr3,  H3K27me3

value: Scramble gRNA3


title: A08_PB_2, eHAP FNLS SUGP1 E554K, 2 nM PB, replicate 2

description:
A08_PB_2
col13

value: clone A08_2.16, SUGP1 E554K


title: Dop pre Dop H1 R3

value: differentiated human embryonic stem cell

value: Dopamine 10 nM 3 weeks, Dopamine 1 mM 1h, RNA Extraction


title: ATAC_seq_TCF1KO_LSK_P45_DN1_Rep2B_Rep1T


title: HBEC3 cells, EmpV,  H3K27me3

value: Human Bronchial cells

value: Empty Vector

value: Tri-Methyl-Histone H3 (Lys27) (CST, 9733)


title: Dop pre Dop H1 R2


title: ATAC_seq_TCF1KO_LSK_P45_DN1_Rep1B_Rep3T


title: Dop pre Dop H1 R1


title: HBEC3 cells, WT,  H3K27me3

value: Wild type


title: ATAC_seq_TCF1KO_LSK_P45_DN1_Rep1B_Rep2T


title: HBEC3 cells, Input, H3K27me3

value: none (input)


title: A08_DMSO_3, eHAP FNLS SUGP1 E554K, DMSO, replicate 3

description:
A08_DMSO_3
col11

value: eHAP FNLS

value: sgRNA SUGP1_238, DMSO 3h


title: Dop pre AA H1 R3

value: Dopamine 10 nM 3 weeks, Ascorbic Acid 100 microM 1h, RNA Extraction


title: A08_DMSO_2, eHAP FNLS SUGP1 E554K, DMSO, replicate 2

description:
A08_DMSO_2
col10


title: ATAC_seq_TCF1KO_LSK_P45_DN1_Rep1B_Rep1T

value: Tcf7eGFP fl/fl Cre+


title: Glut pre AA H1 R1

value: Glutamate 2 microM 3 weeks, Ascorbic Acid 100 microM 1h, RNA Extraction


title: Dop pre AA H1 R2


title: A08_DMSO_1, eHAP FNLS SUGP1 E554K, DMSO, replicate 1

description:
A08_DMSO_1
col9

value: clone A08_2.11, SUGP1 E554K


title: A03_PB_3, eHAP FNLS PHF5A 2xTL, 2 nM PB, replicate 3

description:
A03_PB_3
col8

value: clone A03_2.12, PHF5A 2xTL (6nt insertion at T40-L41)

value: sgRNA PHF5A_6, 2 nM PB 3h


title: HBEC3 cells, R/Y DKO1, H2AK119ub1

value: RYBP and YAF2 double knockout

value: Ubiquityl-Histone H2A (Lys119) (CST, 8240)


title: HBEC3 cells, RKO1,  H3K27me3

value: RYBP knockout


title: HBEC3 cells, R/Y DKO2, H2AK119ub1


title: ATAC_seq_TCF1KO_LSK_L7d_DN1_Rep2B_Rep2T

value: Cultured on OP9-DLL1


title: A08_PB_1, eHAP FNLS SUGP1 E554K, 2 nM PB, replicate 1

description:
A08_PB_1
col12

value: sgRNA SUGP1_238, 2 nM PB 3h


title: A03_PB_2, eHAP FNLS PHF5A 2xTL, 2 nM PB, replicate 2

description:
A03_PB_2
col7

value: clone A03_1.12, PHF5A 2xTL (6nt insertion at T40-L41)


title: HBEC3 cells, YKO2, H2AK119ub1

value: Human Bronchial cells

value: YAF2 knockout


title: A03_PB_1, eHAP FNLS PHF5A 2xTL, 2 nM PB, replicate 1

description:
A03_PB_1
col6

value: clone A03_1.11, PHF5A 2xTL (6nt insertion at T40-L41)


title: Acetyl pre Dop H1 R2

value: differentiated human embryonic stem cell

value: Acetylcholine 750 nM 3 weeks, Dopamine 1mM 1h, RNA Extraction


title: HBEC3 cells, YKO1, H2AK119ub1


title: Acetyl pre Dop H1 R3


title: Acetyl pre Dop H1 R1


title: ATAC_seq_TCF1KO_LSK_L7d_DN1_Rep1B_Rep3T


title: A03_DMSO_3, eHAP FNLS PHF5A 2xTL, DMSO, replicate 3

description:
A03_DMSO_3
col5

value: eHAP FNLS

value: sgRNA PHF5A_6, DMSO 3h


title: HBEC3 cells, RKO2,  H2AK119ub1


title: ATAC_seq_TCF1KO_LSK_L7d_DN1_Rep1B_Rep2T

value: Tcf7eGFP fl/fl Cre+


title: A03_DMSO_2, eHAP FNLS PHF5A 2xTL, DMSO, replicate 2

description:
A03_DMSO_2
col4


title: Dop pre AA H1 R1

value: Dopamine 10 nM 3 weeks, Ascorbic Acid 100 microM 1h, RNA Extraction


title: Acetyl pre AA H1 R3

value: Acetylcholine 750 nM 3 weeks, Ascorbic Acid 100 microM 1h, RNA Extraction


title: ATAC_seq_TCF1KO_LSK_L7d_DN1_Rep1B_Rep1T


title: HBEC3 cells, RKO1,  H2AK119ub1


title: ATAC_seq_TCF1KO_LSK_L7d_DN1_Rep2B_Rep3T


title: ATAC_seq_TCF1KO_LSK_L1d_DN2_Rep2B_Rep1T


title: HBEC3 cells, Scr3,  H2AK119ub1

value: Scramble gRNA3


title: ATAC_seq_TCF1KO_LSK_L7d_DN1_Rep2B_Rep1T


title: HBEC3 cells, EmpV,  H2AK119ub1

value: Empty Vector


title: ATAC_seq_TCF1KO_LSK_L1d_DN2_Rep1B_Rep1T


title: Acetyl pre AA H1 R1


title: AA pre Dop H1 R3

value: Ascorbic Acid 100 microM 3 weeks, Dopamine 1 mM 1h, RNA Extraction


title: ATAC_seq_TCF1KO_LSK_L1d_DN1_Rep2B_Rep3T


title: HBEC3 cells, WT,  H2AK119ub1

value: Wild type

value: Ubiquityl-Histone H2A (Lys119) (CST, 8240)


title: ATAC_seq_TCF1KO_LSK_L1d_DN1_Rep2B_Rep2T

value: Cultured on OP9-DLL1


title: HBEC3 cells, Input, H2AK119ub1

value: none (input)


title: HiS-NET-seq, K562-dTAG-BRD4, 40min-DMSO, R3

city: Innsbruck

first: Univ.-Prof. Dr. Andreas

last: Mayer

address: Innrain 80/82

department: Institute for Genomics and RNomics

country: Austria

institute: Biocenter, Medical University Innsbruck

data_processing:
Data processing of HiS-NET-seq data sets was performed similarly as described in Gajos et al., 2021.
Pre-processing: The ten 5’ end nucleotides correspond to the molecular barcode. Those nucleotides are trimmed from the sequencing reads but remain associated with the sequencing read using a custom python script.
Mapping: Reads are aligned to the human reference genome (GRCh38.p12) using the STAR aligner (v2.5.3a) (Dobin et al., 2013) with the following parameters : -clip3pAdapterSeq ATCTCGTATGCCGTCTTCTGCTTG -clip3pAdapterMMp 0.21 -clip3pAfterAdapterNbases 1 -outFilterMultimapNmax 1 -outSJfilterOverhangMin 3 1 1 1 -outSJfilterDistToOtherSJmin 0 0 0 0 -alignIntronMin 11 -alignEndsType EndToEnd. The alignment was performed with the GENCODE annotation v28.
Filtering: We removed putative contaminations with custom python scripts such as reverse transcription mispriming events (barcode sequences correspond exactly to the genomic sequence adjacent to the aligned read), PCR duplicates (reads that align to the same genomic position and contain identical barcode), and RNA-processing intermediates (exact single bp 3’ ends of introns, exons, and polyA sites).
Data Track: For each strand the single-nucleotide 5'-positions from each sequencing read, corresponding to the 3’-end of the purified nascent RNA fragment, were extracted and summarized in a Pol II occupancy data track.
Masking: RNA species from Pol I, Pol III, microRNA, miscellaneous RNA, rRNA, snoRNA, snRNA, tRNA, vault RNA, Y RNA, and blacklisted regions from ENCODE were masked (Kozomara et al., 2019; Frankish et al., 2019; Jurka et al., 2005; ENCODE Project Consortium).
Spike-ins: Adjustments were made for samples with spiked-in mouse control cells (NIH/3T3). For mapping, a joint reference from the human (GRCh38.p12) and mouse (GRCm38.p6) (Frankish et al., 2019) genomes were used. Furthermore, RNA-processing intermediates from both species were removed using the GENCODE v28 and M18 annotations (Frankish et al., 2019). Next, RNA species from other sources than Pol II (see masking) were masked in the human and mouse genomes. Finally, the sequencing reads mapping to the human genome were separated from those mapping to the mouse genome. The Pol II occupancy in untreated mouse cells was used for data normalization.
Assembly: GRCh38_p12
Supplementary files format and content: *.bw: BigWig file format, Pol II occupancy track (raw counts), stranded, mouse or human

source_name: K562 dTAG-BRD4

growth_protocol: K562 dTAG-BRD4 cells were cultured in RPMI containing 10% FBS, 5% penicillin-streptomycin. NIH/3T3 cells, which were used as spike-ins, were grown in DMEM containing 10% FBS, 5% penicillin-streptomycin. K562 dTAG-BRD4 cells were plated at 5 x 10^5 cells/ml, NIH/3T3 were plated at 2 x 10^6 cells/T75 flask every 48 hours.

extract_protocol:
The cells were lysed in 0.15% NP-40 in PBS to isolate the nuclei. All lysis buffers were supplemented with a protease inhibitor cocktail, RNase inhibitor and α-amanitin. Next, the nuclei were resuspended in glycerol buffer, followed by the addition of nuclei lysis buffer and gentle vortexing, as described by Mayer and Churchman (2016). The chromatin wasdissolved in QIAzol (Qiagen). Next, the dchroomatin of K562 and NIH/3T3 cells was combined at an 8:1 ratio. Chromatin-associated RNA was extracted using the standard Phenol-chloroform procedure, treated with TURBO Dnase (Ambion) and purified by a phenol-chloroform-isoamyl alcohol mixture and isopropanol precipitation. Per sample, 100 ug of the purified chromatin-associated RNA were biotinylated using MTSEA biotin-XX linker (Biotinum) and purified with magnetic μMACS streptavidin MicroBeads (Miltenyi Biotec).
Libraries were generated as described by Gajos et al., 2021. 2-3 µg of the 4sU-enriched RNA were used per sample.
OTHER: HIS-NET-seq

treatment_protocol: 1 x 10^8 K562 dTAG-BRD4 cells at a concentration of 1 x 10^6/ml were treated with 500 nM dTAG7 or DMSO for 40 or 120 min. For the last 10 min of the treatment period, 4sU was added to a final concentration of 500 µM. Untreated NIH/3T3 cells were detached by trypsinization and incubated with 4sU for 10 min while in suspension.

value: K562 dTAG-BRD4

value: bone marrow, CML

value: DMSO

tag: treatment time

value: 40 min

tag: spike-in controls

tag: 4su labelling

value: 10 min


title: HiS-NET-seq, K562-dTAG-BRD4, 40min-DMSO, R2

tag: cell type


title: HiS-NET-seq, K562-dTAG-BRD4, 40min-DMSO, R1

tag: cell line


title: ATAC_seq_TCF1KO_LSK_L1d_DN1_Rep2B_Rep1T

tag: treatment


title: AA pre Dop H1 R1

value: differentiated human embryonic stem cell


title: AA pre AA H1 R3

value: Ascorbic Acid 100 microM 3 weeks, Ascorbic Acid 100 microM 1h, RNA Extraction


title: AA pre AA H1 R2


title: HiS-NET-seq, K562-dTAG-BRD4, 40min-dTAG7, R4


title: ATAC_seq_TCF1KO_LSK_L1d_DN1_Rep1B_Rep3T

value: Tcf7eGFP fl/fl Cre+


title: HBEC3 cells, RKO1, RYBP

value: Human Bronchial cells

value: RYBP knockout

value: RYBP(millipore, AB3637)


title: HiS-NET-seq, K562-dTAG-BRD4, 40min-DMSO, R4


title: HBEC3 cells, RKO2, RYBP


title: ATAC_seq_TCF1KO_LSK_L1d_DN1_Rep1B_Rep2T


title: HiS-NET-seq, K562-dTAG-BRD4, 40min-dTAG7, R3


title: HBEC3 cells, Scr3, RYBP

value: Scramble gRNA3


title: HiS-NET-seq, K562-dTAG-BRD4, 40min-dTAG7, R2


title: AA pre Dop H1 R2

value: Ascorbic Acid 100 microM 3 weeks, Dopamine 1 mM 1h, RNA Extraction


title: ATAC_seq_TCF1KO_LSK_L1d_DN1_Rep1B_Rep1T


title: HiS-NET-seq, K562-dTAG-BRD4, 40min-dTAG7, R1


title: HBEC3 cells, RKO3, RYBP


title: HBEC3 cells, EmpV, RYBP

value: Empty Vector


title: A03_DMSO_1, eHAP FNLS PHF5A 2xTL, DMSO, replicate 1

description:
A03_DMSO_1
col3

value: eHAP FNLS

value: clone A03_1.11, PHF5A 2xTL (6nt insertion at T40-L41)

value: sgRNA PHF5A_6, DMSO 3h


title: Acetyl pre AA H1 R2

value: Acetylcholine 750 nM 3 weeks, Ascorbic Acid 100 microM 1h, RNA Extraction


title: eHAP FNLS, t8, DMSO control, part 1

description:
t8_ctrl_run1
col4

data_processing:
IlluminaHiSeq4000 was PE100 run (requires trimming of R1 and R2), Illumina NextSeq550 was a R1 55 nt and R2 20 nt (only requires trimming of R1).
Sequencing reads werer trimmed (BBMap BBDuk v.38.94). Hard lefttrim for R1 with 5'-cccttggagaaccaccttgttgg-3', retain only 20 nt. For R2 where necessary only retain 20 nt.
Sequencing reads corresponding to the same sample were pooled.
Reads were counted by alignment to a reference file of all sgRNAs present in the pool.
For each individual sgRNA, each barcode was randomly assigned to either of two replicates.
Assembly: hg38
Supplementary files format and content: tab-delimited textfile containing counts per sgRNA for all treatments and time points
Library strategy: sgRNA sequencing

source_name: eHAP

extract_protocol:
At screen end point cells were harvested and gDNA was extracted with QIAamp DNA Blood Maxi Kit (Qiagen #51194). Genome weight was estimated based on measured ploidy of cells.
The sequencing library was prepared using NEBnext® Ultra II Q5 Master Mix (NEB #M0544) and custom primers (forward i5: 5'-AATGATACGGCGACCACCGAGATCTACACTCTTTCCCTACACGACGCTCTTCCGATCT[N]cccttggagaaccaccttgTTG-3' with N=(0-8) nt stagger and reverse primer for multiplexing: 5'-CAAGCAGAAGACGGCATACGAGAT[8N]GTGACTGGAGTTCAGACGTG-3')  to have a balanced read sample. A barcode in the reverse primer was used for identification of the sequencing libraries. Libraries were gel purified and cleaned up. Libraries were balanced and quality was assessed with Bioanalyzer High Sensitivity DNA Kit, Agilent #5067-4626). R1 was used for sgRNA identification, R2 was used to read out barcode associated with each single sgRNA in the pool

treatment_protocol: The screen was performed at 500x sgRNA representation for entire duration. 66 million eHAP FNLS cells were infected with the lentiviral spliceosome library (marked with BFP) at an MOI of ~0.3, such that every sgRNA was represented in approximately 500 cells. Puromycin selection was started at 48 h post transduction. At six days post-transduction, cells were assessed by flow cytometry to only contain sgRNA carrying cells (BFP-positive cells >95%). Cells were then split into treatment arms (DMSO vs. 2 nM PB; with identical DMSO concentration in both treatment arms). Cells were propagated and treatment was renewed every second day.

value: eHAP

value: expressing FNLS

value: DMSO, 8 days


title: HiS-NET-seq, K562-dTAG-BRD4, 120min-DMSO, R2

value: K562 dTAG-BRD4

value: bone marrow, CML

value: DMSO

value: 120 min

value: 10 min


title: HBEC3 cells, WT, RYBP

value: Human Bronchial cells

value: Wild type


title: ATAC_seq_TCF1KO_LSK_HMGd_DN1_Rep2B_Rep1T

value: Tcf7eGFP fl/fl Cre+

value: Cultured on OP9-DLL1


title: eHAP FNLS t8, 2 nM PB, part 1

description:
t8_2PB_run1
col5

value: 2 nM pladienolide B, 8 days


title: HBEC3 cells, Input, RYBP

value: none (input)


title: ATAC_seq_TCF1KO_LSK_HMGd_DN1_Rep1B_Rep3T


title: HiS-NET-seq, K562-dTAG-BRD4, 120min-DMSO, R1


title: HiS-NET-seq, K562-dTAG-BRD4, 120min-dTAG7, R2


title: AA pre AA H1 R1

value: differentiated human embryonic stem cell

value: Ascorbic Acid 100 microM 3 weeks, Ascorbic Acid 100 microM 1h, RNA Extraction


title: ATAC_seq_TCF1KO_LSK_HMGd_DN1_Rep1B_Rep2T


title: HiS-NET-seq, K562-dTAG-BRD4, 120min-dTAG7, R1


title: standard NET-seq

value: WT

value: control

value: 0 min


title: HBEC3 cells, QKO3, batch2

data_processing:
Sequencing was performed with an Illumina NextSeq 500 machine and 75 bp paired-end runs were performed.
paired-end RNAseq reads were trimmed using TrimGalore with defaut parameters.
Trimmed reads were aligned to the mouse genome mm9 with STAR (v2.5.1) , and gene count were performed using a STAR (v2.5.1)
The limma (v3.38.2) statistical package19 was used to call the differential expressed gene. Statistical significance for differentially expressed genes was q<0.05 with FC > 2.
Assembly: hg19
Supplementary files format and content: bigWig, counts coverage

extract_protocol:
Total RNA was isolated with PureLink RNA Mini Kit (Invitrogen).
Total RNA integrity was verified with an Agilent 2100 Bioanalyzer (Agilent Technologies) and quantified with a NanoDrop 8000 instrument (Thermo Fisher). RNA-seq libraries were prepared from total RNA with the KAPA RNA HyperPrep kit with RiboErase (KAPA Biosystems)

value: RYBP,TET1,TET2,TET3 quadruple knockout


title: ATAC_seq_TCF1KO_LSK_HMGd_DN1_Rep1B_Rep1T


title: HiS-NET-seq, K562, input, R2


title: HBEC3 cells, QKO1, batch2


title: ATAC_seq_TCF1KO_LSK_HMGd_DN1_Rep2B_Rep2T


title: HBEC3 cells, QKO2, batch2


title: ATAC_seq_TCF1KO_LSK_EV_DN1_Rep2B_Rep3T


title: HiChIP, K562-dTAG-BRD4, 120min-DMSO, R3

data_processing:
For HiChIP data processing, we used the HiC-Pro v3.0.0 (Servant et al., 2015) pipeline at a 10 kb resolution for the applied restriction enzyme (MboI: ^GATC) and the human reference genome (GRCh38.p12).
Next, we used HiCcompare (Stansfield et al., 2018) to perform data normalization and comparison between the pooled HiChIP samples.
We defined significant 3D interaction changes between loci with an average expression >9, a minimum distance of 10 kb, and a padj value of 0.05 or lower.
Assembly: GRCh38.p12
Supplementary files format and content: *.matrix: tab seperated table, raw interaction counts (3rd column) between two genomic loci (1st and 2nd column, bin ID listed in 1000.bed)
Supplementary files format and content: *.bed: BED file format, genomic loci with ID (4th column)
Supplementary files format and content: *.res: tab separated table, HiCcompare results with normalized interaction frequencies (11th and 12th column), logarithmic fold-change between conditions (13th column), and adjusted p-values (18th column).

growth_protocol: K562 dTAG-BRD4 cells were cultured in RPMI containing 10% FBS, 5% penicillin-streptomycin and were plated at 5 x 10^5 cells/ml every 48 hours.

extract_protocol:
HiChiP libraries were prepared as described by Mumbach et al. (2016). In brief, the cells were crosslinked with 1% formaldehyde for 10 min and gently lysed in 10 mM Tris-HCl pH 8.0, 10 mM NaCl, 0.2% NP40 supplemented with a protease inhibitor cocktail. The nuclei were partially permebilized with 0.5% SDS, which was then quenched by Triton X-100. Next, the nuclei were incubated with 375 U MboI overnight. Biotin-dATP incorporation was catalyzed by Klenow fragment, followed by the re-ligation of the DNA fragments and sonication for 6 min in a Covaris S220 instrument. 5 µg of an H3K27ac-specific antibody (Abcam) were coupled to magnetic Protein A/G beads (Thermo Fisher) and added to the chromatin for overnight incubation. Then, the chromatin was washed, eluted and purified.
Streptavidin-coupled magnetic beads were added to 50 ng DNA of each sample to enrich for biotin-marked DNA fragments. While still bead-bound, the DNA was treated with 2.5 µl Tn5 transposase for 10 min and PCR-amplified. The libraries were size-selected and purified using AMPure XP beads.
OTHER: HiChIP

treatment_protocol: 5 x 10^6 K562 dTAG-BRD4 cells at a concentration of 1 x 10^6/ml were treated with 500 nM dTAG7 or DMSO for 120 min.

value: K562 dTAG-BRD4


title: HBEC3 cells, TET123 TKO3, batch2

value: TET1,TET2,TET3 triple knockout


title: HiS-NET-seq, K562, input, R1

value: bone marrow, CML


title: HiS-NET-seq, K562, 4sU, R2


title: ATAC_seq_TCF1KO_LSK_EV_DN1_Rep2B_Rep2T


title: HiChIP, K562-dTAG-BRD4, 120min-DMSO, R1

source_name: K562 dTAG-BRD2

value: K562 dTAG-BRD2

value: DMSO

value: 120 min


title: control, eHAP FNLS, t0, part 1

description:
t0_ctrl_run1
col3

value: eHAP

value: expressing FNLS

value: no treatment


title: HiS-NET-seq, K562, 4sU, R1

value: 10 min


title: plasmid control part 1

description:
pla_ctrl_run1
col2

source_name: plasmid control

value: plasmid control


title: HBEC3 cells, TET123 TKO1, batch2

value: Human Bronchial cells


title: ATAC_seq_TCF1KO_LSK_EV_DN1_Rep1B_Rep3T

value: Tcf7eGFP fl/fl Cre+

value: Cultured on OP9-DLL1


title: HiChIP, K562-dTAG-BRD4, 120min-dTAG7, R2

source_name: K562 dTAG-BRD0

value: K562 dTAG-BRD0


title: ATAC_seq_TCF1KO_LSK_EV_DN1_Rep1B_Rep2T


title: HiChIP, K562-dTAG-BRD4, 120min-dTAG7, R1

source_name: K562 dTAG-BRD-1

value: K562 dTAG-BRD-1


title: ChIP-Rx, K562-dTAG-BRD4, Input, R2

data_processing:
We used Bowtie2 v2.3.5.1 (Langmead et al., 2012) to align the sequencing reads to a joint reference which consisted of the human (GRCh38.p12) and mouse (GRCm38.p6) reference genomes using the paired-end mode with the parameter -k 1.
Potential PCR duplicates were marked using PICARD’s v2.24.2 (https://gatk.broadinstitute.org/hc/en-us/articles/360037052812-MarkDuplicates-Picard-) markDuplicates function.
DNA-binding profiles of BRD4 were extracted using MACS2's v2.2.7.1 (Zhang et al., 2008) bdgcmp function with the -m FE parameter.
Finally, the DNA-binding profiles corresponding to the human genome were separated from the BRD4 binding sites in the mouse genome.
Assembly: GRCh38_p12 and GRCm38_p6
Supplementary files format and content: *.bw: BigWig file format, BRD4 occupancy track (fold enrichment over matched input control), unstranded, mouse or human
Library strategy: ChIP-Rx

extract_protocol:
K562 dTAG-BRD4 cells were crosslinked with 1% formaldehyde for 5 min at room temperature, NIH/3T3 cells were crosslinked for 8 min. After combining NIH/3T3 and K562 dTAG-BRD4 cells in a ratio of 1:5, cells were lysed as described by Baluapuri et al. (2019), followed by sonication using a Covaris E220evolution at intensity 4, duty cycle 5%, 200 cycles per burst for 20 min. Lysates were pre-cleared by centrifugation. Lysates and antibodies coupled to Dynabeads Protein G (Thermo Fisher Scientific) were incubated at 4°C overnight with rotation. Afterwards, the beads were washed with buffers prepared according to Baluapuri et al. (2019). DNA fragments were purified using the ChIP DNA Clean & Concentrator kit (Zymo).
Libraries were constructed from 6 to 30 ng DNA using the NEBNext Ultra II DNA kit (NEB) according to the manufacturer's instructions. After purifying the amplified libraries with AMPure XP beads (Beckman Coulter), fragments of 200 to 500 bp were extracted from an 8% TBE gel. Library quality and quantity were assessed using BioAnalyzer HS DNA (Agilent) and Qubit dsDNA HS (Thermo Fisher) assays, respectively.
OTHER: ChIP-Rx

treatment_protocol: 4 x 10^7 K562 dTAG-BRD4 cells at a concentration of 1 x 10^6/ml were treated with 500 nM dTAG7 or DMSO for 40 or 120 min.

tag: matched input

value: ChIP-Rx, K562-dTAG-BRD4, Input, R2


title: ChIP-Rx, K562-dTAG-BRD4, Input, R1

value: K562 dTAG-BRD4

value: ChIP-Rx, K562-dTAG-BRD4, Input, R1


title: HBEC3 cells, RKO2, batch2

value: RYBP knockout


title: HiChIP, K562-dTAG-BRD4, 120min-DMSO, R2

source_name: K562 dTAG-BRD3

value: K562 dTAG-BRD3

value: bone marrow, CML


title: ATAC_seq_TCF1KO_LSK_EV_DN1_Rep1B_Rep1T


title: ATAC-seq_OV organoid, Day 5, GFP positive

city: Edinburgh

first: Liusaidh

middle: Jane

last: Owen

address: Crewe Road South

department: MRC HGU

institute: University of Edinburgh

data_processing:
FastQC v0.11.4 was used to perform quality control and cutadapt v1.16 to trim adapter sequences from the sequencing reads.
Bowtie2 v2.3.4.3 was used to align trimmed reads to the mm10 genome. Duplicate reads, unmapped reads, and reads mapping to the mitochondrial genome were removed using samtools v1.6.
To enrich for fragments indicative of binding of TFs, reads corresponding to fragment sizes > 100bp were filtered out (i.e. retained only sub-nucleosomal reads for downstream analysis).
Peaks were called using the callpeak function of MACS2 v2.1.1 with parameters -f BAMPE -g mm -keep-dup all -B -SPMR -nomodel, on the paired-end bam files of each sample separately. Bedgraph files from macs2 were converted to BigWig files using bedGraphtoBigWig.
Using the MACS2 peak coordinates, a consensus peak set was created using the bedtools merge command with default settings to merge overlapping peaks. From this consensus set of peaks the peaks overlapping blacklisted regions of the mm10 genome, as listed on http://mitra.stanford.edu/kundaje/akundaje/release/blacklists/ were removed. FeatureCounts from the R package Rsubread was used to generate read count values for the consensus peaks. A conditional quantile normalization was used to normalize signal in consensus ATAC-seq peaks while correcting for the effects of peak width and GC content. Finally ChIPseeker was used to annotate peaks by their genomic context, collapsing peak annotations into four categories: promoter, intergenic, intronic and exonic.
Assembly: mm10
Supplementary files format and content: Big wig files of subnucleosomal read depth genome wide. Tab separated value file showing the normalised read count per ATACseq peak for each sample along with chromosomal coordinates of the peak, the peaks genomic context and the nearest gene.

source_name: Rx-GFP K/I EB5

growth_protocol: Rx-GFP mESCs from the Riken BRC Cell Bank (cell number AES0145) were maintained under feeder free conditions. Cells were cultured on 0.1% gelatin coated tissue culture flasks  in GMEM media supplemented with 1X non- essential amino acids, 1 mM Sodium pyruvate, 10 μM 2-Mercaptoethanol, 10% knock out serum replacement (KOSR), 1% FCS and LIF. Care was taken to avoid over-confluency or undesirable differentiation of cells.

extract_protocol:
Following FACS, cells were pelleted at 1200 RPM for 4 minutes at 4◦C and then resuspended in ice cold PBS at a concentration of 100,000 cells per ml. 50,000 cells (500 μl of cell suspension) were pelleted, resuspended in 100 μl and centrifuged again before the supernatant was removed. Cells were then resuspended in 100 μl of cold ATAC lysis buffer (10 mM TrisCl, pH 7.4, 10 mM NaCl, 3 mM MgCl2, 0.1% IGEPAL CA-630) and kept on ice for 15 minutes, with occasional gentle pipetting. Samples were pelleted at 1200 RPM for 5 minutes at 4◦C, supernatant discarded and resuspended in transposition mix consisting of 50 μl ChIPmentation buffer (10 mM Tris pH8, 5 mM MgCl2, 10% Dimethylformamide) and 2.5 μl Tn5 transposase enzyme (Illumina, 15027866) per sample. The transposition reaction was carried out at 37◦C for 30 minutes.
Samples were purified using the Qiagen MinElute Reaction Cleanup Kit as per manufacturer’s instructions, eluting in 10 μl elution buffer. Libraries were then PCR amplified and purified using AMPure XP beads. Libraries were sequenced (75bp paired-end reads) on the Illumina Hi-seq 4000 platform.

treatment_protocol: SFEBq differentiation of OV organoids was performed as described previously (Wataya et al., 2008; Eiraku et al., 2011). Briefly, mESCs were dissociated and 4500 cells were plated in differentiation media (supplemented GMEM with 1.5% KOSR) in a Nunclon low cell adhesion 96 well plate. This is defined as day 0. On day 1, growth factor reduced Matrigel was added to a final concentration of 2%. Differentiation status was assessed using fluorescent imaging with the presence of GFP used as a measure of Rax expression. On days 0-3 and 5 of optic vesicle differentiation, 48 organoids were pooled, dissociated and sorted using FACS to select for live single cells. The sample collected at day 5 was also sorted on the basis of GFP expression giving a GFP positive and a GFP negative sample for this time point.

value: Rx-GFP K/I EB5

value: ESC

value: optic vesicle organoid differentiation

tag: time

value: Day 5


title: HBEC3 cells, RKO1, batch2


title: ChIP-Rx, K562-dTAG-BRD4, 120min-DMSO, R2

value: DMSO

value: 120 min


title: ATAC-seq_OV organoid, Day 5, GFP negative


title: HiChIP, K562-dTAG-BRD4, 120min-dTAG7, R3

source_name: K562 dTAG-BRD1

value: K562 dTAG-BRD1


title: HBEC3 cells, Scr3, batch2

value: Human Bronchial cells

value: Scramble gRNA3


title: ChIP-Rx, K562-dTAG-BRD4, 120min-DMSO, R1


title: ATAC_DN3-KO_HMG-E10_P45_Rep2

source_name: Scid.adh cell line

value: WT TCF-1 RV


title: ATAC_DN3-KO_HMG-E10_P45_Rep3


title: ATAC_DN3-KO_HMG-E10_P45_Rep1


title: ATAC-seq_OV organoid, Day 3

value: Day 3


title: ATAC-seq_OV organoid, Day 2

value: Day 2


title: ChIP-Rx, K562-dTAG-BRD4, 40min-dTAG7, R2

value: K562 dTAG-BRD4

value: 40 min

value: ChIP-Rx, K562-dTAG-BRD4, Input, R2


title: HBEC3 cells, WT, batch2

value: Wild type


title: ChIP-Rx, K562-dTAG-BRD4, 40min-dTAG7, R1

value: bone marrow, CML

value: ChIP-Rx, K562-dTAG-BRD4, Input, R1


title: HBEC3 cells, TET123 TKO2, batch2

value: TET1,TET2,TET3 triple knockout


title: ATAC_DN3-KO_HMG-E10_L7_Rep4

value: DL7 TCF-1 RV


title: ATAC_seq_TCF1KO_LSK_EV_DN1_Rep2B_Rep1T

value: Tcf7eGFP fl/fl Cre+

value: Cultured on OP9-DLL1


title: ATAC-seq_OV organoid, Day 1

value: Rx-GFP K/I EB5

value: ESC

value: optic vesicle organoid differentiation

value: Day 1


title: ChIP-Rx, K562-dTAG-BRD4, 120min-dTAG7, R2

value: 120 min


title: HBEC3 cells, R/Y DKO3, batch1

value: Human Bronchial cells

value: RYBP and YAF2 double knockout


title: ATAC_DN3-KO_HMG-E10_L7_Rep3


title: ATAC-seq_OV organoid, Day 0

value: Day 0


title: HBEC3 cells, R/Y DKO2, batch1


title: ChIP-Rx, K562-dTAG-BRD4, 120min-dTAG7, R1

value: K562 dTAG-BRD4


title: expt3_d12_mLN_GCB_Cxcr3lo_C

city: Birmingham

first: Alexander

last: Rosenberg

state: AL

address: 1720 2nd Avenue South, Suite 276/11

institute: University of Alabama at Birmingham

description: Ultra-Low Input RNA-Seq

data_processing:
Trimming: TrimGalore v.0.4.4 with settings: phred33 scores, paired, nextera adapters.
Alignment: STAR aligner v.2.5.2a with default settings.
Counting: Htseq-count v0.6.1p1 with settings: unstranded and using GRCm39 annotations.
edgeR for downstream analysis to find differentially expressed genes among pairs of sample groups.
Assembly: GRCm39
Supplementary files format and content: ArroyoDiaz_2022_RNAseq_count_table.txt: Raw count table (tab-delimited text).

source_name: C57BL/6 mouse

extract_protocol:
RNA was isolated using the Single Cell RNA Purification kit (Norgen).
RNA libraries were prepared for sequencing using standard Illumina protocols. All libraries were prepared at Genewiz.

tag: mouse pool

tag: tissue

value: medial LN

value: Cxcr3 Low Germinal Center B Cell

value: WT

value: Influenza Infection

tag: day post infection


title: ATAC_DN3-KO_HMG-E10_L6_Rep4

value: DL6 TCF-1 RV


title: expt3_d12_mLN_GCB_Cxcr3lo_B

description: Ultra-Low Input RNA-Seq

tag: genotype


title: HBEC3 cells, R/Y DKO1, batch1


title: expt3_d12_mLN_GCB_Cxcr3lo_A

description: Ultra-Low Input RNA-Seq


title: HBEC3 cells, EmpV, batch2

value: Empty Vector


title: ATAC_DN3-KO_HMG-E10_L6_Rep1


title: ATAC_DN3-KO_HMG-E10_L6_Rep3


title: HBEC3 cells, RYBP/T123 QKO2

data_processing:
Paired-end reads were trimmed using TrimGalore(version 0.5.0) with the following parameters : --length 50, --clip_R1 10, --clip_R2 18, --three_prime_clip_R1 10, and --three_prime_clip_R2 10.
Trimmed reads were aligned to the hg19 using Bismark version 0.19.0
Duplication were removed using the deduplicate_bismark script provided with Bismark
CpG methylation values were called by the bismark_methylation_extractor script provided with Bismark and the following parameters: --no_overlap, --comprehensive, --merge_non_CpG, and --cytosine_report.
Assembly: hg19
Supplementary files format and content: bigWig, methylation coverage with cutoff 3

extract_protocol:
Genomic DNA was extracted using a Quick-DNA Miniprep Plus kit (Zymo Research, D4070).
The Accel-NGS Methyl-Seq DNA Library Kit (Swift Biosciences, 30024) was used according to the manufacturer’s instructions to perform bisulfite conversion and library preparation.
WGBS

value: RYBP,TET1,TET2,TET3 quadruple knockout


title: HBEC3 cells, YKO2, batch1

value: YAF2 knockout


title: HBEC3 cells, RKO3, batch2

value: RYBP knockout


title: HBEC3 cells, YKO3, batch1


title: expt3_d12_mLN_GCB_Cxcr3hi_C

description: Ultra-Low Input RNA-Seq

value: Cxcr3 High Germinal Center B Cell


title: HBEC3 cells, RYBP/T123 QKO1


title: HBEC3 cells, YKO1, batch1


title: ATAC_DN3-KO_HMG-E10_L2_Rep4

value: DL2 TCF-1 RV


title: HBEC3 cells, RKO3, batch1


title: HBEC3 cells, TET123 TKO2

value: TET1,TET2,TET3 triple knockout


title: expt3_d12_mLN_GCB_Cxcr3hi_A

description: Ultra-Low Input RNA-Seq


title: ATAC_DN3-KO_HMG-E10_L7_Rep1

value: DL7 TCF-1 RV


title: ATAC_DN3-KO_HMG-E10_L2_Rep2


title: HBEC3 cells, TET123 TKO1

value: Human Bronchial cells


title: HBEC3 cells, RKO2, batch1


title: expt2_d30_mLN_GCB_C

description: Ultra-Low Input RNA-Seq

value: Germinal Center B Cell


title: expt2_d30_mLN_GCB_B

description: Ultra-Low Input RNA-Seq


title: HBEC3 cells, TET2/3 DKO3

value: TET2,TET3 double knockout


title: HBEC3 cells, TET2/3 DKO2


title: ATAC_DN3-KO_HMG-E10_L1del_Rep3

value: DL1 TCF-1 RV


title: ATAC_DN3-KO_HMG-E10_L2_Rep1


title: expt3_d12_mLN_GCB_Cxcr3hi_B

description: Ultra-Low Input RNA-Seq

value: medial LN


title: HBEC3 cells, RYBP/T123 QKO3


title: HBEC3 cells, TET2/3 DKO1


title: HBEC3 cells, TET123 TKO3


title: ATAC_DN3-KO_HMG-E10_L1del_Rep2


title: expt2_d30_mLN_GCB_A

description: Ultra-Low Input RNA-Seq


title: expt2_d30_lung_Bmem_C

description: Ultra-Low Input RNA-Seq

value: lung

value: Memory B Cell


title: ATAC_DN3-KO_HMG-E10_L1del_Rep1


title: HBEC3 cells, Scr2, batch1

value: Scramble gRNA2


title: expt2_d30_lung_Bmem_B

description: Ultra-Low Input RNA-Seq

value: WT

value: Influenza Infection


title: ATAC_DN3-KO_HMG-E10_HMG_del_Rep3

value: DHMG TCF-1 RV


title: HBEC3 cells, TET2 KO3

value: TET2 knockout


title: HBEC3 cells, TET2 KO2


title: ATAC_DN3-KO_HMG-E10_HMG_del_Rep2


title: expt2_d30_lung_Bmem_A

description: Ultra-Low Input RNA-Seq


title: HBEC3 cells, WT, batch1

value: Wild type


title: HBEC3 cells, EmpV, batch1

value: Empty Vector


title: HBEC3 cells, Scr1, batch1

value: Scramble gRNA1


title: ATAC_DN3-KO_HMG-E10_HMG_del_Rep1


title: HBEC3 cells, RKO1, batch1

value: RYBP knockout


title: expt1_d12_mLN_GCB_IFNgRko_C

description: Ultra-Low Input RNA-Seq

source_name: 50/50 mixed WT and IFNgR-/- BM chimera C57B mouse

value: IFNgR-/- Germinal Center B Cell

value: IFNgR-/-


title: HBEC3 cells, RYBP KO3

value: Human Bronchial cells


title: shp16shp15-3_S21

city: Tel Aviv

first: Metsada

last: Pasmanik-Chor

address: Ramat Aviv

department: Biology

country: Israel

institute: Tel Aviv University

data_processing:
Partek Flow (build 8.0.19.0610).
Trim bases based on quality score, min Phred score 20.
STAR - 2.6.1d aligned to hg38.
Quantify to annotation model (Partek E/M). Gene level qualntifiaction, RPKM normalization.
Genome_build: hg38
Supplementary_files_format_and_content: gene_rpkm_07012020_1-12.xls

source_name: COLO-357 (metastatic pancreatic adenocarcinoma)

growth_protocol: Cells were grown in DMEM containing 10% fetal bovine serum (FBS).

extract_protocol:
Total RNA extraction, library preparation and sequencing were conducted as previously described (Boldes T, Merenbakh-Lamin K, Journo S, et al. R269C variant of ESR1: high prevalence and differential function in a subset of pancreatic cancers. BMC Cancer 2020;20:531).
as previously described (Boldes T, Merenbakh-Lamin K, Journo S, et al. R269C variant of ESR1: high prevalence and differential function in a subset of pancreatic cancers. BMC Cancer 2020;20:531).

treatment_protocol: Cells were transfected using Jet Pei (Polyplus Transfection, France). Lentiviral infection was conducted as described (Zinger L, Merenbakh-Lamin K, Klein A, et al. Ligand-binding Domain-activating Mutations of ESR1 Rewire Cellular Metabolism of Breast Cancer Cells. Clin Cancer Res 2019;25:2900-2914).

value: PDAC cell line COLO-357

value: CDKN2A and CDKN2B (p15+p16) silencing

value: Co-silencing


title: expt1_d12_mLN_GCB_IFNgRko_B

description: Ultra-Low Input RNA-Seq

value: medial LN


title: ATAC_DN3-KO_HMG-E10_EV_Rep2b

value: empty vector RV


title: HBEC3 cells, Scr3, batch1

value: Scramble gRNA3


title: shp16shp15-1_S23


title: HBEC3 cells, RYBP KO1


title: shp16shp15-2_S22


title: ATAC_DN3-KO_HMG-E10_EV_Rep2a


title: expt1_d12_mLN_GCB_WT_C

description: Ultra-Low Input RNA-Seq

value: WT Germinal Center B Cell

value: WT


title: HBEC3 cells, RYBP KO2


title: expt1_d12_mLN_GCB_WT_B

description: Ultra-Low Input RNA-Seq

value: Influenza Infection


title: HBEC3 cells, Scramble gRNA3


title: HBEC3 cells, TET2 KO1

value: TET2 knockout


title: HBEC3 cells, Empty vector


title: shp16-2_S2

value: CDKN2A (p16) silencing

value: shRNAs against p16


title: expt1_d12_mLN_GCB_WT_A

description: Ultra-Low Input RNA-Seq


title: ATAC_DN3-KO_HMG-E10_EV_Rep1a


title: HBEC3 cells, Wild type

value: Wild type


title: shp16-3_S1


title: lib46030 CD23Cre Germinal center B cells at 14 days after NP-KLHImmunization

city: Chicago

first: Mark

last: Maienschein-Cline

state: IL

address: 900 S Ashland Ave, Rm 1058, M/C 937

department: Research Resources Center

institute: University of Illinois at Chicago

description: Live B220+ GL7hi CD95hi

data_processing:
Base calls were processed to FASTQs on BaseSpace (Illumina).
Reads were processed using workflows managed on the Galaxy platform; reads were trimmed by 1 base at the 3’ end, and then trimmed from both ends until base calls had a minimum quality score of at least 30 (Galaxy FASTQ Trimmer tool v1.0.0)
FastqMcf (v1.1.2, https://benthamopen.com/ ABSTRACT/TOBIOIJ-7-1) was used to remove any remaining adapter sequence.
Reads were aligned by STAR1 2.4.2a to GRCm38
Gene counts were generated using HTSeq-count (v0.4.1). Quality metrics were compiled from PICARD (v1.134, http://broadinstitute.github.io/picard/), FASTQC (v0.11.3, http://www.bioinformatics.babraham.ac.uk/projects/fastqc/), Samtools (v1.2), and HTSeq-count (v0.4.1).
Assembly: GRCm38
Supplementary files format and content: CSV of raw gene counts per sample

source_name: Spleen

extract_protocol:
RNA was harvested using Rneasy mini plus kit (Qiagen).
SMART-SeqV4_Bulk_Half_Vol
Nextera XT Half Volume index set D

value: Spleen

value: Germinal Center B Cells (Bulk Sort)

value: intraperitonial NPKLH immunization, Day 14 after Immunization


title: ATAC_DN3-KO_HMG-E10_EV_Rep3


title: expt1_d12_mLN_GCB_IFNgRko_A

description: Ultra-Low Input RNA-Seq

value: IFNgR-/- Germinal Center B Cell

value: IFNgR-/-


title: shp16-1_S3

value: PDAC cell line COLO-357


title: A375 cells, shUSP22-3

city: Changsha

first: Huiyan

last: Sun

address: National Clinical Research Center for Geriatric Disorders, Xiangya Hospital, Central South University, Changsha, Hunan, China.

institute: Xiangya hospital Central South University

data_processing:
Quality control：Raw data (raw reads) of fastq format were firstly processed through in-house perl scripts. In this step, clean data (clean reads) were obtained by removing reads containing adapter, reads containing ploy-N and low quality reads from raw data. At the same time, Q20, Q30 and GC content the clean data were calculated.
Reads mapping to the reference genome：Reference genome and gene model annotation files were downloaded from genome website directly. Index of the reference genome was built using Hisat2 v2.0.5 and paired-end clean reads were aligned to the reference genome using Hisat2 v2.0.5.  We selected Hisat2 as the mapping tool for that Hisat2 can generate a database of splice junctions based on the gene model annotation file and thus a better mapping result than other non-splice mapping tools.
Novel transcripts prediction：The mapped reads of each sample were assembled by StringTie (v1.3.3b) (Mihaela Pertea.et al. 2015) in a reference-based approach. StringTie uses a novel network flow algorithm as well as an optional de novo assembly step to assemble and quantitate fulllength transcripts representing multiple splice variants for each gene locus.
Quantification of gene expression level：featureCounts v1.5.0-p3 was used to count the reads numbers mapped to each gene. And then FPKM of each gene was calculated based on the length of the gene and reads count mapped to this gene. FPKM, expected number of Fragments Per Kilobase of transcript sequence per Millions base pairs sequenced, considers the effect of sequencing depth and gene length for the reads count at the same time, and currently the most commonly used method for estimating gene expression levels.
Differential expression analysis： Differential expression analysis of two  conditions/groups (two biological replicates per condition) was performed using the DESeq2 R package (1.16.1).
GO and KEGG enrichment analysis of differentially expressed genes
Assembly: mm10
Supplementary files format and content: tab-delimited text files include RPKM values for each Sample

source_name: skin

growth_protocol: A375 melanoma cells were maintained in DMEM supplemented with 10% fetal bovine serum (FBS) and antibiotics in humidified atmosphere with 5% CO2 at 37C

extract_protocol:
RNA was harvested using MagZol Reagent (Magen). 1.3 ug of total RNA.
RNA was harvested using NEBNext® Ultra™ RNA Library Prep Kit for Illumina® was used for the construction of sequencing libraries.

value: skin

value: human melanoma cell

value: USP22 knockdown


title: WT_Noninfected, rep 6

city: Aurora

first: Andrew

last: Goodspeed

state: CO

address: 12801 E 17th Ave, Mail Stop 8303

institute: University of Colorado Anschutz Medical Campus

data_processing:
Illumina adapters were removed using BBDuk and reads <50bp after trimming were discarded
Reads were aligned and quantified using STAR (2.6.0a) to the Ensembl mouse transcriptome (GRCm38.p6, release 96)
Lowly abundant genes were removed as those <1 mean raw count or <1 mean count per million (CPM) across the dataset
Differential gene expression was calculated using the voom function in limma
Assembly: GRCm38
Supplementary files format and content: Text file containing STAR gene count values

source_name: Non-infected wild-type murine tracheal epithelial cells

growth_protocol: Epithelial cells were grown and differentiated using culture media previously described in literature (PMID: 12388377; PMID: 12959927; PMID: 21372793; PMID: 23522475)

extract_protocol:
Differentiated epithelial cells were lysed in situ for RNA extraction and subsequent bulk RNA analysis
RNA libraries were prepared for sequencing using standard Nugen Universal Plus mRNA-Seq kit

treatment_protocol: The murine tracheal epithelial cells were divided into two groups: (1) unchallenged (non-infected) and (2) challenged with influenza strain A/Puerto Rico/8/1934 H1N1 (IAV PR8). Once mTEC cultures were fully confluent and well-differentiated (ALI day 21 or later), the epithelial cells were infected at multiplicity of infection of 0.1 with IAV PR8 diluted in culture medium. Viral inoculum was added to the apical chamber of each transwell (into the insert and over the luminal surface ciliated mTECs) at 200 µl inoculum per insert. The cells were then incubated for two hours at 35oC to allow adsorption of virus to the cells. After 2 hours, the inoculum was removed and TWIs were washed once with 200 µl of culture medium and the infected cells were returned to culture at 37oC, 5% CO2 in a humidified chamber. 48 hrs post-inoculation, apical washed were performed to removed released viral particles. The cells were then lysed for RNA extraction for bulk RAN-seq analysis.

value: Noninfected


title: lib46029 CD23Cre Germinal center B cells at 14 days after NP-KLHImmunization

description: Live B220+ GL7hi CD95hi


title: WT_Noninfected, rep 5


title: shp15-2_S5

value: CDKN2B (p15) silencing

value: shRNAs against p15


title: ATAC_DN3-KO_HMG-E10_EV_Rep1b

value: empty vector RV


title: shp15-1_S6


title: lib46028 CD23Cre Germinal center B cells at 14 days after NP-KLHImmunization

description: Live B220+ GL7hi CD95hi


title: A375 cells, shUSP22-1


title: A375 cells, shControl-3

value: WT


title: A375 cells, shUSP22-2


title: shNS-3_S7

value: control

value: NS-shRNA


title: WT_Noninfected, rep 3


title: lib46026 CD23Cre Germinal center B cells at 14 days after NP-KLHImmunization

description: Live B220+ GL7hi CD95hi


title: shNS-2_S13


title: shp15-3_S4


title: lib46027 CD23Cre Germinal center B cells at 14 days after NP-KLHImmunization

description: Live B220+ GL7hi CD95hi


title: lib46025 STAT3fl/fl.CD23Cre Germinal center B cells at 14 days after NP-KLHImmunization

description: Live B220+ GL7hi CD95hi


title: WT_Noninfected, rep 2


title: WT_Noninfected, rep 1


title: A375 cells, shControl-1


title: lib46024 STAT3fl/fl.CD23Cre Germinal center B cells at 14 days after NP-KLHImmunization

description: Live B220+ GL7hi CD95hi


title: lib46023 STAT3fl/fl.CD23Cre Germinal center B cells at 14 days after NP-KLHImmunization

description: Live B220+ GL7hi CD95hi


title: Human hepatocytes, replicate 1, snRNAseq (dataset II)

city: Westmead

first: Suzanne

last: Scott

state: NSW

address: 214 Hawkesbury Rd

country: Australia

institute: Children's Medical Research Institute

description:
Nuclear RNA
hep1

data_processing:
The demultiplexing, barcoded processing, gene counting and aggregation 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).
Assembly: Combined GRCh38, mm10, AAV reference
Supplementary files format and content: Tab-separated values files and matrix files

source_name: Liver

extract_protocol:
To identify and characterize cell states associated with AAV transduction in human heptocytes, we employed a droplet-based (10x chromium) single nucleus transcriptomics approach. Hepatic nuclei were isolated and profiled from (a) human hepatocytes before engraftment, (b) liver samples from FRG mice engrafted with human hepatocytes, and (c) liver samples from FRG mice engrafted with human hepatocytes, treated with two AAV vectors (LLK03, REDH).
Libraries were prepared from single nucei.  Breifly, nuclei were isoalted from flash-frozen chunks of tissue, which were homogenized using a Kimble Dounce tissue grinder, followed by straining with a 40 μm cell strainers, two rounds of centrifugation at 500g.  Nuclei were stained with DAPI and proridium iodide, followed by FACS sorting to isolate PI-positive nuclei. Libraries were then prepared using a 10x Genomics Chromium Single Cell 3’v3 Reagent Kits were used according to manufacturer’s instructions.

value: Liver

value: Hepatocytes

value: Unengrafted

tag: sample type


title: WT_Infected, rep 4

source_name: IAV-infected wild-type murine tracheal epithelial cells

value: Infected


title: Human hepatocytes, replicate 2, snRNAseq (dataset II)

description:
Nuclear RNA
hep2


title: A375 cells, shControl-2


title: lib46022 STAT3fl/fl.CD23Cre Germinal center B cells at 14 days after NP-KLHImmunization

description: Live B220+ GL7hi CD95hi

value: Spleen

value: Germinal Center B Cells (Bulk Sort)

value: intraperitonial NPKLH immunization, Day 14 after Immunization


title: WT_Infected, rep 3


title: lib46021 STAT3fl/fl.CD23Cre Germinal center B cells at 14 days after NP-KLHImmunization

description: Live B220+ GL7hi CD95hi


title: 293T cells,RSL3 & Fer-1,4h

city: shanghai

first: changzhou

last: feng

address: No. 197, Ruijin 2nd Road

institute: Shanghai Institute of Hematology, Ruijin Hospital Affiliated to Shanghai Jiao Tong University School of Medicine

data_processing:
Basecalls performed using CASAVA version 1.8
Adapters and low-quality read ends were removed using Trimmomatic, based on quality control statistics generated by FastQC.
Quality filtered reads were then mapped to GRCh38 using Hisat2
The expression level of each transcript was calculated according to the fragments per kilobase of exon per million mapped reads (FRKM) . StringTie was used to quantify gene and isoform abundances.
R statistical package software EdgeR (Empirical analysis of Digital Gene Expression in R) was utilized for differential expression analysis.
Assembly: GRCh38
Supplementary files format and content: tab-delimited text files include RPKM values for each Sample

growth_protocol: 293T cells were maintained in DMEM supplemented with 10% fetal bovine serum (FBS) and antibiotics in humidified atmosphere with 5% CO2 at 37C

extract_protocol:
RNA was extracted using TRIzol reagent (Invitrogen, 15596018).
Isolation of mRNA was performed using the NEBNext PolyA mRNA Magnetic Isolation Module. TruSeq RNA preparation kit (KAPA) was used with 1 ug of total RNA for the construction of sequencing libraries.

treatment_protocol: 293T cells treated with DMSO, EC359(0.5μM), and RSL3(0.5μM) alone, or co-treated with Fer-1(2μM).

value: Human embryonic kidney


title: WT_Infected, rep 5


title: WT_Infected, rep 2


title: 293T cells,EC59 & Fer-1,4h


title: GCB STAT3 replicate 1

data_processing:
Raw reads were mapped to the reference genome using BWA MEM. PCR duplicates were removed using Picard MarkDuplicates. Peaks were called using Macs2 against an input sample previously published on GEO (GSM3926504). High-confidence peak calls were determined based on a threshold of 50.
Assembly: mm9
Supplementary files format and content: bed

source_name: Mouse splenic germinal center B cells

extract_protocol:
ChIP-seq was performed using the ChIP Assay Kit following the manufacturer’s protocol (Millipore). DNA was sheared to ~200 bp with a Bioruptor sonication water bath for four 15 min cycles consisting of 45s on and 30s off. ChIP was performed with anti-STAT3 (D3Z2G, Cell Signaling Technologies). Resulting DNA was purified with the MinElute PCR Purification Kit (Qiagen). DNA was incubated with a tagmentation enzyme (Illumina) at 37 °C for 30 min and DNA was purified with the MinElute PCR Purification Kit (Qiagen).
DNA libraries were made with the Nextera Indexing kit (Illumina), and DNA was purified with the QiaQuick PCR Purification Kit (Qiagen).

treatment_protocol: Mice were immunized intraperitoneally (i.p.) with 200µg of 4-Hydroxy-3-nitrophenylacetyl-Keyhole Limpet Hemocyanin (NP-KLH, Biosearch Technologies) mixed with Complete Freund’s Adjuvant (CFA, Sigma-Aldrich) on d0 followed by 100µg NP-KLH in Incomplete Freund’s Adjuvant (IFA, Sigma-Aldrich) on d7. Spleens from 10 mice were harvested, processed into single cell suspensions, and pooled on 14d post immunization. Samples were generated from two experiments using a total of 20 mice. Following RBC lysis using Tris-Ammonium Chloride, B cells were first enriched by negative selection (Stem Cell Technologies) and then stained for GC B cell markers viability- , anti-B220 BV605, anti-GL7 FITC and anti-CD95 PE-Cy7. GC B Cells were sorted using a FACS Aria SORP high-performance cell sorter (BD Biosciences) and immediately cross-linked in 37% formaldehyde (Sigma) diluted in PBS (Corning) to a final concentration of 1.5% formaldehyde. Cells were fixed for 12 minutes at 37 C and then reaction was stopped by adding Glycine (Cell Signaling Technologies) and a 10 minute incubation at room temperature. Cells were then washed with PBS and the pellet was snap frozen and stored at -80 C until processing.

tag: tissue/cell type

value: Germinal center B cell

value: Mouse splenic germinal center B cells


title: DP16_Noninfected, rep 6

source_name: Non-infected Dp16 murine tracheal epithelial cells

value: Noninfected


title: shNS-1_S24

value: PDAC cell line COLO-357


title: 293T cells,EC539,4h


title: DP16_Noninfected, rep 5


title: WT_Noninfected, rep 4


title: 293T cells,DMSO,4h

value: DMSO


title: scD1, wt replicate 2, scRNAseq

description: 10x Genomics

data_processing:
The demultiplexing, barcoded processing, gene counting and aggregation were made using the Cell Ranger software v3.1.0
Assembly: mm10
Supplementary files format and content: Count matrix files

source_name: whole embryos

extract_protocol:
Embryos were harvested at E8.0 in ice-cold phosphate buffered saline (PBS) containing 10% (v/v) of fetal bovine serum (FBS/PBS). Embryos aged between 0-1 somite were selected for further analysis and the extraembryonic tissue was removed. Embryos were washed with Ca+2/Mg+2- free Dulbecco’s phosphate buffered saline DPBS. Embryos were incubated in TrypLETM Express (Gibco) at 37oC for 6-7 min with intermittent pipetting using wide bore tips. Once embryos were completely dissociated, the enzyme was quenched using 10% FBS/DPBS (Ca+2/Mg2+ free) before the cells were resuspended in Ca2+/Mg2+ free DPBS containing 5% (v/v) endotoxin free FBS. Cell viability (i.e., lack of dead cells), number and size were confirmed with a hemocytometer.
Library was performed according to the manufacter’s instructions (single cell 3’ library v3 protocol, 10x Genomics).

tag: strain

value: C57BL/6 x 129 SvEv

value: whole embryos

tag: developmental stage

value: E.8 (0-2 somites) stage embryos