title: Untreated replicate 3

description:
tablecounts_raw.csv
D203T04

value: untreated


title: Untreated replicate 1

status: Public on May 06 2020

type: SRA

city: Paris Cedex 05

first: Nicolas

last: Servant

address: 26 rue d'ulm

country: France

institute: Institut Curie

description:
tablecounts_raw.csv
D203T02

library_source: transcriptomic

data_processing:
Raw sequencing reads were aligned on the human hg38 reference genome using the STAR mapper [7] (2.5.3a), up to the generation of a raw count table per gene (Gencode annotation v26). Expressed genes (TPM>=1 in at least one sample) have then been selected for downstream analysis. The raw count table was then normalized using the TMM method from the edgeR R package [5] (v3,22,3), and the limma voom (v3.36.3) functions were applied to detect genes with differential expression between untreated and EGF samples. Genes with an adjusted pvalue < 0,05 and a log2 fold-change > 1 were called significant.
Genome_build: hg38
Supplementary_files_format_and_content: table of raw counts per gene

molecule: total RNA

organism: Homo sapiens

source_name: Basal breast cancer cells

extract_protocol:
MDA-MB-468 cells were grown to ~ 70% confluence on 140 cm2 dishes and treated with EGF (100 ng mL-1) for 72 h in biological duplicate. Untreated cells were used as an appropriate control. Whole cell extracts (~ 9 x 105 cells) were collected by centrifugation at 500 x g for 5 min at 4ºC and washed twice in ice-cold PBS1x. Total RNA were extracted using RNeasy Mini Kit (Qiagen) according to the manufacturer’s protocol and subjected to DNase I treatment using the RNase-free DNase set (Qiagen).  The total RNA was then analyzed using Agilent RNA 6000 Nano Kit on the Agilent 2100 Bioanalyzer System to determine RNA concentration, qualify RNA Integrity Number (RIN) which was high with RIN = 10 for all samples and confirm removal of DNA contamination.  After quality control of RNA, the ERCC RNA Spike-In (Ambion) was added to RNA samples (RNA samples: RNA Spike-In control diluted at 1/100, 5:1).
RNA sequencing libraries were prepared from 1µg of total RNA using the Illumina TruSeq Stranded mRNA Library preparation kit which allows to perform a strand specific sequencing. A first step of polyA selection using magnetic beads is done to focus sequencing on polyadenylated transcripts. After fragmentation, cDNA synthesis was performed and resulting fragments were used for dA-tailing followed by ligation of TruSeq indexed adapters. PCR amplification was finally achieved to generate the final barcoded cDNA libraries. The 4 libraries were equimolarly pooled and subjected to qPCR quantification using the KAPA library quantification kit (Roche). Sequencing was carried out on the NovaSeq 6000 instrument from Illumina based on a 2*100 cycle mode (paired-end reads, 100 bases) using a S1 flow cell in order to obtain around 35 million clusters (70 million raw paired-end reads) per sample

treatment_protocol: Cells were harvested by trypsinization. After adding media to stop trypsin, cells were centrifuged at 500 x g for 5 min at room temperature.

tag: cell line

value: MDA-MB-468 cells

tag: passages

tag: treatment


title: JS1902_05

city: West Haven

first: Sarah

last: Slavoff

state: CT

address: 300 Heffernan Dr

department: Chemistry

country: USA

institute: Yale University

description: Reads filtered for number of T-to-C mutations (0-5+) and strandedness (min/pos)

data_processing:
Reads were filtered for unique sequences using FastUniq
Reads were trimmed using cutadapt (--minimum-length=20)
Reads were aligned to the GRCh38 genome using STAR aligned with default parameters and --mp 4,2 .
Reads were processed using Picard tools, including FixMateInformation, SortSam and BuildBamIndex
Reads were filtered for those that aligned uniquely (flag: 83/163, 99/147), with MAPQ ≥ 2, and without insertions.
HTSeq-count was used to identify mapped reads in UCSC transcripts (union mode)
T-to-C mutations were identified using Rsamtools and a custom R script (available upon request). Only mutations at positions with a base quality score of greater than 45 that were at least three nt from the end of the read were counted. Reads were excluded where there were greater than five T-to-C mutations and these mutations did not account for at least one third of the observed mutations (NM tag).
SNP sites were filtered from our data in two ways: 1, bcftools was used to identify T-to-C SNP sites in control samples, and these sites were excluded from analysis; 2, sites exhibiting high T-to-C mutation rates in non-s4U treated controls were excluded from analysis.
To examine the distribution of reads with each minimum number of T-to-C mutations, bam files were filtered using Picard tools.
STAR aligner was used to make genome-coverage tracks (inputAlignmentsFromBam mode, outWigType bedGraph), and tracks were normalized using DESeq2 (estimateSizeFactors).
Tracks were converted to binary format using IGVtools (toTDF).
Genome_build: GRCh38
Supplementary_files_format_and_content: bedGraph track. Bedgraph files represent different numbers of T to C mutations identified: TC.0 = 0 mutations, TC.1 = 1 mutation, TC.2 = 2 mutations, etc.

source_name: immortalized 293Ts

growth_protocol: 293T cells were grown in DMEM containing 1% penicillin/streptomycin, 10% FBS, at 37°C.

extract_protocol:
RNA was harvested with Trizol reagent, followed by chloroform extraction and ethanol precipitation. Genomic DNA was depleted using Turbo DNAse, followed by cleanup with RNAclean beads. RNA was treated with oxidative-nucleophilic-aromatic-substitution chemistry as described in Schofield et al. Nature Methods 2018.
Libraries were generated following standard protocols using the SMARTer Stranded Total RNA-Seq Kit-Pico Input Mammalian, V2 (Takara Bio USA, cat. 634413).

treatment_protocol: s4U plus and minus cells had their media replaced with growth media supplemented with 500 uM s4U or left unsupplemented, respectively. Cells were incubated with s4U for 2h.

tag: genotype

value: MSI2 KO

tag: cell type

value: immortalized 293Ts

tag: s4u feed

value: Yes


title: Untreated replicate 2

description:
tablecounts_raw.csv
D203T03

growth_protocol: MDA-MB-468 cells were grown to ~ 70% confluence on 140 cm2 dishes and treated with EGF (100 ng mL-1) for 72 h in biological duplicate. Untreated cells were used as an appropriate control. Whole cell extracts (~ 9 x 105 cells) were collected by centrifugation at 500 x g for 5 min at 4ºC and washed twice in ice-cold PBS1x.


title: JS1902_03

value: Wild Type

value: No


title: DDX5 Knockdown rep3

city: West Lafayette

first: Elizabeth

middle: J

last: Tran

state: IN

address: 175 S. University Street

department: Biochemistry

institute: Purdue University

data_processing:
Data quality control (minimum Phred score 30, minimum read length 50 bp) was performed was performed using TrimGalore toolkit (version 0.4.4).
Quality trimmed reads were mapped to human reference genome (GRCh38) downloaded from Ensembl (https://www.ensembl.org) database, using the STAR aligner (version 2.5.4b).
Reads aligned to each gene feature were counted via HTSeq package.
Genome_build: GRCh38
Supplementary_files_format_and_content: Counts file describing the counts of the reads aligned to each gene feature in each replicate.

source_name: H69AR (ATCC® CRL-11351™)

growth_protocol: H69AR cells were obtained from the American Type Culture Collection (ATCC) and cultured using RPMI-1640 medium (ATCC® 30-2001™) supplemented with 20% FBS (Atlanta Biologicals S11150) and 10U Penicillin-Streptomycin (ThermoFisher 15140122).

extract_protocol:
Total RNA was extracted using Trizol reagent (ThermoFisher, 15596026) and treated with DNAse (ThermoFisher, AM2238).
Library preparation was done using the TruSeq® Stranded Total RNA Library Prep Kit (Illumina, 20020596), which depletes ribosomal RNAs.

treatment_protocol: The SMARTvector inducible lentiviral shRNAs (Dharmacon) were used for depleting DDX5. Specifically, H69AR cells expressing either a control shRNA (Dharmacon, VSC11651) or shDDX5 (ATCCAATCCACTTAGAGCA) was treated with 1 µg/ml doxycycline for 3 days before RNA extraction.

tag: lentivirus

tag: genotype/variation

value: DDX5 knockdown


title: HSCEpigenomics_SP3KO_scRNA

city: PHILADELPHIA

first: Kai

last: Tan

state: PA

address: 3501 CIVIC CENTER BLVD.

institute: Children's Hospital of Philadelphia

description:
scRNA-Seq
processed data file: gene_by_cell_count_matrix.txt
processed data file: cell_annotation.csv
processed data file: gene_annotation.csv

data_processing:
Data was processed using the 10X Genomics CellRanger pipeline. Sequencing reads were mapped to mm10 reference genome.
Filtering was performed on the raw gene-barcode matrix output by the Cell Ranger cellranger count function, removing barcodes that have fewer than 1000 transcripts and 1000 expressed genes genome build mm10.
Genome_build: mm10 (GRCm38)
Supplementary_files_format_and_content: gene_by_cell_count_matrix.txt contains a sparse matrix where rows are genes (with gene information in the gene_annotation.csv), columns are cells (with cell information in the cell_annotation.csv), and values are UMI counts for the given gene in the given cell.
Supplementary_files_format_and_content: cell_annotation.csv contains a data frame with annotations (i.e. 10X barcode, cell type, etc.) for each cell.
Supplementary_files_format_and_content: gene_annotation.csv contains a data frame with gene id and gene short name.

organism: Mus musculus

source_name: Embryonic day 10.5 major arteries endothelium

extract_protocol:
E10.5 embryos were collected from Sp3+/- females mated to Sp3+/- males. Dorsal aorta regions were isolated and dissociated using 0.125% collagenase (Sigma) for 40 min. Cells were then washed and stained with the following antibodies (1:200): Ter119, ESAM, CD41, CD144, CD31, and DAPI (Invitrogen, D3571) for viability. The cells were pooled by genotype and sorted for endothelial cells on a BD Influx into 1xPBS + 10% FBS in 1.5 LowBind Tubes.
Libraries were prepared using 10x Genomics protocol (Chromium Single Cell 3' Reagent Kits v3, CG000183 RevA).

tag: strain

tag: tissue

value: Dorsal Aorta

tag: age

value: Embryonic day 10.5

value: major arteries endothelium


title: JS1902_02


title: DDX5 Knockdown rep2


title: EGF 72 replicate 1

description:
tablecounts_raw.csv
D203T06

value: EGF (100 ng mL-1) for 72 h


title: HSCEpigenomics_WT_scRNA

value: WT


title: DDX5 Knockdown rep1


title: RNA-Seq sample

city: Duluth

first: Marshall

last: Hampton

state: MN

address: 1117 University Drive

institute: University of Minnesota Duluth

data_processing:
Contigs assembled with Trinity v.2.4.0
Reads aligned to contigs using NCBI blastn
Araport 11 Arabidopsis thaliana proteins used for annotation
Genome_build: None available
Supplementary_files_format_and_content: NesoS60_contig_counts.xls

organism: Nesocodon mauritianus

source_name: flower

extract_protocol: Illumina’s Truseq RNA Preparation v2 kit (RS-122-2001)

value: Nectary tissue


title: JS1902_06


title: WT rep3

value: control shRNA

value: wild type


title: JS1902_04


title: HE_H3K27ac_Rep2

library_source: genomic

data_processing:
Sequencing reads were mapped to the mouse genome (mm9) using Bowtie2 (v2.2.2) with default parameter setting
Uniquely mapped reads from both ChIPed and input DNA were used to compute a normalized signal for each 200 bp bin across the genome
ChIP-Seq peaks were called by MACS using default parameter setting
Genome_build: mm9
Supplementary_files_format_and_content: bigWig

molecule: genomic DNA

source_name: Embryonic day 10.5 major arteries hemogenic endothelium

extract_protocol:
5 stages of hematopoietic stem and progenitor cells were sorted from mouse embryos or adult bone marrow. ChIP reactions were performed using 20,000 FACS-sorted cells. Iped and Input DNA were purified with MinElute PCR Purification Kit.
Libraries were constructed using Rubicom ThruPLEX DNA-Seq Kit. Libraries were size-selected at 100-700 bp by AMPure XP beads. Libraries were quantified with KAPA qPCR and bioanalyzer prior to single-end sequencing.

value: AGM

tag: chip antibody

value: H3K27ac (ab4729, Abcam)

value: major arteries hemogenic endothelium


title: WT rep2


title: WT rep1


title: IMR8A2

city: Austin

first: Michael

middle: S

last: Sacks

state: TX

address: 201 East 24th St, Stop C0200

department: Oden Institute for Computational Engineering and Sciences and the Department of Biomedical Engineering

institute: The University of Texas at Austin

data_processing:
Raw reads (in FASTQ format) were processed by Novogene with Perl scripts to remove low quality reads and those containing adapters or poly-N sequences.
The index of the reference genome was built using Bowtie (v2.2.3).
Paired-end reads were aligned to the reference genome using TopHat (v2.0.12).
HTSeq (v0.6.1) was used to count the reads numbers mapped to each gene.
Genome_build: 3.1
Supplementary_files_format_and_content: tab delimited text file (geneCounts.csv) include raw counts for all samples

molecule: polyA RNA

organism: Ovis aries

source_name: mitral valve anterior leaflet

extract_protocol:
Isolation of total mRNA was performed using the RNeasy mini kit (Qiagen, Valencia, CA). 1% β-mercaptoethanol was added to the lysis buffer and 30mg of previously flash-frozen frozen tissue were homogenized with a TissueRuptor (Qiagen). Isolated RNA purity was preliminarily assessed on a NanoDrop spectrophotometer (NanoDrop technologies, Wilmington, DE). RNA concentration was measured on a Qubit 2.0 fluorometer (Invitrogen, Whaltham, MA). RNA integrity assessment was performed on an Agilent Bioanalyzer 2100 using a Nano 6000 assay kit (Agilent Technologies, Santa Clara, CA). A RNA integrity number (RIN) > 7.1 was considered the minimum requirement for library preparation.
RNA library preparation and sequencing was performed at Novogene (Sacramento, CA). RNA quality control by 1% agarose gels and Agilent Bioanalizer 2100 was repeated before library construction. A total amount of 1 µg RNA per sample was used as input material for the RNA sample preparations. Sequencing libraries were generated using NEBNext Ultra RNA Library Prep Kit for Illumina (NEB, USA) following manufacturer's recommendations and index codes were added to attribute sequences to each sample. Briefly, mRNA was purified from total RNA using poly-T oligo-attached magnetic beads. Fragmentation was carried out using divalent cations under elevated temperature in NEBNext First Strand Synthesis Reaction Buffer (5X). First strand cDNA was synthesized using random hexamer primer and M-MuLV Reverse Transcriptase (RNase H-). Second strand cDNA synthesis was subsequently performed using DNA Polymerase I and RNase H. Remaining overhangs were converted into blunt ends via exonuclease/polymerase activities. After adenylation of 3' ends of DNA fragments, NEBNext Adaptor with hairpin loop structure were ligated to prepare for hybridization. In order to select cDNA fragments of preferentially 150-200 bp in length, the library fragments were purified with AMPure XP system (Beckman Coulter, Beverly, USA). 3 µL USER Enzyme (NEB, USA) was used with size-selected, adaptor-ligated cDNA at 37C for 15 min followed by 5 min at 95C before PCR. PCR was performed with Phusion High-Fidelity DNA polymerase, Universal PCR primers, and Index (X) Primer. At last, PCR products were purified (AMPure XP system) and library quality was assessed on the Agilent Bioanalyzer 2100 system. The clustering of the index-coded samples was performed on a cBot Cluster Generation System using PE Cluster Kit cBot-HS (Illumina) according to the manufacturer's instructions. After cluster generation, the library preparations were sequenced on an Illumina platform and 125 bp/150 bp paired-end reads were generated.

treatment_protocol: IMR was induced in adult (30-40 kg) Dorset sheep raised for laboratory work and supplied by commercial vendors following established procedures. Briefly, sheep were anaesthetised with sodium thiopental (10-15 mg/kg) intraveneously, intubated, and ventilated with isoflurane (1.5-2%) and oxygen. Surface electrocardiogram, arterial blood pressure, and other vital signs were continuously monitored throughout the duration of the procedure. IMR was induced by a thoracotomy to facilitate ligation of the second and third obtuse marginal branches of the circumflex coronary artery. Permanent occlusion of these arteries reliably results in a transmural posterior MI that includes the entire posterior PM and involves approximately 20% of the LV mass, causing a gradual onset of severe IMR within eight weeks. The degree of MR was assessed with echocardiography. MV leaflets were collected and flash-frozen at pre-MI as well as 4 and 8 weeks post-MI.

tag: time

value: 8 weeks post-MI

tag: mitral regurgitation

value: YES


title: Puro_3

city: San Francisco

first: Andrei

last: Goga

state: CA

address: 513 Parnassus Ave

department: Cell & Tissue Biology

institute: UCSF

data_processing:
Raw image data file from high-throughput sequencing(like Illumina ) was transformed to Sequenced Reads(called Raw Data or Raw Reads) by CASAVA base recognition (Base Calling).
Raw reads filtering conditions are as follows: (1) Remove reads containing adaptors; (2) Remove reads containing N > 10% (N represents base that could not be determined); (3) The Qscore (Quality value) of over 50% bases of the read is <= 5. RNA-seq Adapter information: NEBNext® UltraTM RNA Library Prep Kit  RNA 5' Adapter (5'Adaptor) 5'AATGATACGGCGACCACCGAGATCTACACTCTTTCCCTACACGACGCTCTTCCGATCT RNA 3' Adapter (3'Adaptor,The underlined 6bp bases is Index) 5'GATCGGAAGAGCACACGTCTGAACTCCAGTCACATCACGATCTCGTATGCCGTCTTCTGCTTG
Indexes of the reference genome was built using STAR and paired-end clean reads were aligned to the reference genome using STAR (v2.5). STAR used the method of Maximal Mappable Prefix(MMP) which can generate a precise mapping result for junction reads.
HTSeq v0.6.1 was used to count the read numbers mapped of each gene. And then FPKM of each gene was calculated based on the length of the gene and reads count mapped to this gene.
(For DESeq2 with biological replicates) Differential expression analysis between two conditions/groups (two biological replicates per condition) was performed using the DESeq2 R package (2_1.6.3).
Genome_build: Homo Sapiens(GRCh37/hg19)
Supplementary_files_format_and_content: excel file containing readcounts, FPKM, log2FC and p values for differential expression analysis

source_name: mammary epithelial cell line

extract_protocol:
RNA isolated using Qiagen RNeasy Plus Mini kit (gDNA elimination column used) from MCF10A human mammary epithelial cells
mRNA was purified from total RNA using poly-T oligo-attached magnetic beads. The mRNA was first fragmented randomly by addition of fragmentation buffer, then NEB library was prepared. Q-PCR is used to accurately quantify the library effective concentration (> 2nM), in order to ensure the library quality. Libraries fed into Illumina Platform, and paired-end reads were generated

value: mammary epithelial cell line

value: Puro/Blast-expressing


title: IMR8A3

value: NO


title: IMR8A4


title: IMR8A1


title: Puro_2


title: ECs, TD_14

type: RNA

city: Marburg

first: Wilhelm

last: Bertrams

address: Hans-Meerwein-Str. 2

department: Institute for Lung Research

country: Germany

institute: Philipps University Marburg

description: NOD1 stimulation

data_processing:
Raw Ct data are shown in Matrix non-normalized. U6 snRNA (average of 4 values) was used for normalization in Matrix normalized, yielding deltaCt values.
Fold changes were calculated with the the ddCt method (2^-ddCt)

label: FAM/NFQ-MGB

source_name: EC (primary lung endothelial cell)

label_protocol: RNA was reverse transcribed with the Life Technologies miR RT kit with megaplex primers. Reverse transcribed RNA was then run on a TLDA plate with FAM-labelled primers

extract_protocol: Total RNA was isolated by cell lysis in Trizol and subsequent phenol/chloroform precipitation

treatment_protocol: Primary lung endothelial cells were isolated from wildtype mice and stimulated with the NOD1 ligand TD or NOD2 ligand MDP. Unstimulated samples were used as controls. RNA was extracted from these samples.

value: EC (primary lung endothelial cell)

value: NOD1 ligand TD


title: ECs, TD_11

description: NOD1 stimulation


title: IMR4A3

value: 4 weeks post-MI


title: RNA-seq_thrb_KO_3

city: Saint Louis

first: Joseph

last: Corbo

state: MO

address: 660 S. Euclid

department: Pathology and Immunology

institute: Washington University School of Medicine

data_processing:
Sequence reads were trimmed using Cutadapt 1.9.1 with parameters -a CAAGCAGAAGACGGCATACGAGAT -m 36 -o
Sequence reads were aligned to danRer10 using STAR 2.7.2b with an index prepared for 50 bp reads
Normalized read counts were generated using HTseq 0.9.1
Differential gene counts between "WT" and "thrb KO" (thrb stl628/st628) samples were calculated and used to create a matrix of differential gene explression (titled "res.thrb_wtvko.csv") and MA plot using DESeq2 1.18.1 in R 3.4.3
Genome_build: danRer10
Supplementary_files_format_and_content: csv, differential read counts as calculated in DEseq2.1

organism: Danio rerio

source_name: Retina

extract_protocol:
To perform RNA-seq on WT and thrb stl628/stl628 retina, we first isolated the retinas of 6-month-old WT and clutch-matched thrb stl628/stl628 zebrafish. Two retinas from a male and two retinas from a female zebrafish were combined to make one sample. Three WT and thrb stl628/stl628 samples were collected and RNA extracted using trizol-chloroform extraction. RNA concentrations ranged between 27-55 ng/μl, and all RNA integrity number scores were > 8.7. 2-4 μg mRNA were used for library preperation.
First, Ribosomal RNA was removed by poly-A+ selection using Oligo-dT beads (mRNA Direct kit, Life Technologies). Then, mRNA was fragmented in buffer containing 40 mM tris acetate pH 8.2, 100 mM potassium acetate, and 30 mM magnesium acetate heated to 94o C for 150 seconds. Subsequently, mRNA was reverse transcribed to yield cDNA using SuperScript III RT enzyme (Life Technologies), and a second-strand reaction was performed to produce cDNA. cDNA ends were made blunt, an “A” base was added to the 3’ ends, and Illumina sequencing adapters were ligated to the ends. Ligated fragments were then amplified for 12 cycles using primers incorporating unique index tags.

tag: genetic background

value: AB*

value: 6 months-old

value: thrb-/-


title: RNA-seq_thrb_KO_2


title: ECs, TD_8

description: NOD1 stimulation


title: Puro_1


title: Tol-7000(3)

city: Suita

first: Kenichi

last: Ogawa

state: Osaka

address: 1-5, Yamadaoka

country: Japan

institute: Osaka University

description: PCC6803 HL Tolerant Strain cultured under Light 7000 in replicate 3

hyb_protocol: Gene Expression Hybridization kit (Agilent Technologies)

scan_protocol: Agilent G2565CA microarray scanner (Agilent Technologies), Scan control software (Agilent Technologies), and Feature extraction software (Agilent Technologies)

data_processing:
The software program Matlab 2013b (The Mathworks Inc., Natick, MA) was used for data analysis. After removing signal intensities that the Feature extraction software judged unreliable, the median of the probe signal intensities representing each ORF was defined as the gene expression level.
The median of the intensities of the 10 probes representing each ORF was defined as the expression level of the corresponding gene. Expression data was normalized by quantile normalization to compare the expression data each other.
Quantile normalized median of the probe signal intensities (see Supplemental_Data_gene_expression_ratios.xlsx).

organism: Synechocystis sp. PCC 6803

source_name: PCC6803 HL tolerant, Light7000

label_protocol: Fairplay III microarray labeling kit (Agilent Technologies)

growth_protocol: The glucose-tolerant wild type strain of Synechocystis sp. PCC 6803 and PCC 6803 HL Tolerant Strain were cultivated in modified BG-11 medium. The main culture was performed using test tube (φ3 cm×20 cm, AGC Techno Glass, Shizuoka, Japan) with 30 mL modified BG11 medium at 34°C under continuous light illumination at 4000 ~ 9000 µmol m-2 s-1 by point source LED (LA-HDF158AS, HAYASHI watch-works, Tokyo, Japan). The cultures were aerated and mixed by sterile air. The light intensity (4000 ~ 9000 µmol m-2 s-1) of the present study was too high compared to other studies. Because the point source LED used in the present study was very small and all the culture broth in the test tube was not exposed by the described light intensity, the cells could grow under such HL intensity condition. The light intensity described in this paper was the highest light intensity measured by the surface of the test tube using for culture experiment.

extract_protocol: Ambion Ribopure yeast kit (Life Technologies Co., Carlsbad, CA)

treatment_protocol: The culture broth was mixed with an equal volume of 10% (w/v) phenol in ethanol and then centrifuged to harvest the cells at mid-log growth phase (OD730 = around 0.6). Harvested cells were immediately frozen in liquid nitrogen and stored at –80°C.

value: HL Tolerant Strain

tag: light intensity

tag: replicate


title: IMR4A4


title: Wild-7000(3)

description: PCC6803 Wild Type cultured under Light 7000 in replicate 3

source_name: PCC6803 Wild, Light7000

value: Wild Type


title: RNA-seq_thrb_KO_1


title: Wild-7000(2)

description: PCC6803 Wild Type cultured under Light 7000 in replicate 2


title: Tol-4000(2)

description: PCC6803 HL Tolerant Strain cultured under Light 4000 in replicate 2

source_name: PCC6803 HL tolerant, Light4000


title: ECs, TD_5

description: NOD1 stimulation


title: Myc_3

tag: cell line

value: Myc-expressing


title: IMR4A2


title: RNA-seq_WT_3

value: wild-type


title: Tol-4000(3)

description: PCC6803 HL Tolerant Strain cultured under Light 4000 in replicate 3


title: Tol-7000(1)

description: PCC6803 HL Tolerant Strain cultured under Light 7000 in replicate 1


title: Wild-4000(3)

description: PCC6803 Wild Type cultured under Light 4000 in replicate 3

source_name: PCC6803 Wild, Light4000


title: Tol-7000(2)

description: PCC6803 HL Tolerant Strain cultured under Light 7000 in replicate 2


title: ECs, MDP_15

description: NOD2 stimulation

value: NOD2 ligand MDP


title: Tol-4000(1)

description: PCC6803 HL Tolerant Strain cultured under Light 4000 in replicate 1


title: Myc_2


title: RNA-seq_WT_2


title: IMR4A5


title: ECs, MDP_12

description: NOD2 stimulation


title: IMR4A1


title: Myc_1


title: Wild-7000(1)

description: PCC6803 Wild Type cultured under Light 7000 in replicate 1


title: CA4

value: pre-MI


title: Wild-4000(2)

description: PCC6803 Wild Type cultured under Light 4000 in replicate 2


title: ECs, MDP_6

description: NOD2 stimulation


title: Wild-4000(1)

description: PCC6803 Wild Type cultured under Light 4000 in replicate 1


title: RNA-seq_WT_1


title: HRas_3

value: HRas-expressing


title: HRas_2


title: M3: Sp-Male-BK 3

city: Lausanne

first: Sandra

last: Calderon

address: Génopode Building

department: Center for Integrative Genomics

country: Switzerland

institute: University of Lausanne

data_processing:
Purity-filtered reads were adapters and quality trimmed with Cutadapt v.1.8. Reads matching to rRNA sequences were removed with fastq_screen v. 0.9.3. Remaining reads were further filtered for low complexity with reaper v. 15-065.
Reads were aligned against the Spiroplasma poulsonii MSRO (v2) genome using STAR v. 2.5.2b
The number of read counts per gene locus was summarized with htseq-count v. 0.6.1 using Spiroplasma poulsonii MSRO (v2) gene annotation.
Genome_build: Spiroplasma poulsonii MSRO (accession: JTLV02000000)
Supplementary_files_format_and_content: Tab-delimited text files include htseq gene counts for each sample.

organism: Spiroplasma poulsonii

source_name: Hemolymph of male BK ORr flies

extract_protocol:
RNA was extracted from (i) the hemolymph of 30 1-week-old infected flies pelleted by 30 min of centrifugation at 16,000 x g by the TRIzol method following the manufacturer’s instructions. Three independent replicates were prepared for each condition.
RNA-seq libraries were prepared using 25 ng of total RNA and the Illumina TruSeq Stranded mRNA reagents (Illumina; San Diego, California, USA) without the polyA selection step.

value: MSRO

tag: host age

value: 1 week

tag: host tissue

value: hemolymph

tag: host sex

value: male


title: CA3

value: NO


title: ECs, Ctrl_13

description: unstimulated

tag: cell type

value: EC (primary lung endothelial cell)

tag: treatment

value: control


title: M2: Sp-Male-BK 2


title: CA2


title: HRas_1

value: mammary epithelial cell line


title: M1: Sp-Male-BK 1


title: AurKB_3

value: AurKB-expressing


title: ECs, Ctrl_10

description: unstimulated


title: ECs, Ctrl_7

description: unstimulated


title: F3: Sp-Fem-BK 3

source_name: Hemolymph of female BK ORr flies

value: female


title: ECs, MDP_9

description: NOD2 stimulation


title: AurKB_2


title: 5EU, 5IU

city: Göttingen

first: Björn

last: Schwalb

address: Fassberg 11

institute: MPI

description: Labeled RNA

data_processing:
Direct RNA nanopore sequencing data preprocessing of synthetic RNAs. Direct RNA nanopore sequencing reads were obtained for each of the samples (Supplementary Table 1).  FAST5 files were base-called using Guppy 3.1.5 (Oxford Nanopore Technologies) with the following parameters: guppy_basecaller -c rna_r9.4.1_70bps_hac.cfg. Direct RNA nanopore sequencing reads were mapped with GraphMap 0.5.2 46 to the synthetic RNA reference sequence with the following parameters: graphmap align --evalue 1e-10. Further data processing was carried out using the R/Bioconductor environment.
Direct RNA nanopore sequencing data preprocessing of human K562 cells. Direct RNA nanopore sequencing reads were obtained for each of the samples (Supplementary Table 1).  FAST5 files were base-called using Guppy 3.1.5 (Oxford Nanopore Technologies) with the following parameters: guppy_basecaller -c rna_r9.4.1_70bps_hac.cfg. Direct RNA nanopore sequencing reads were mapped with Minimap2 2.10 47 to the GRCh38/hg38 genome assembly (Human Genome Reference Consortium) with the following parameters: minimap2 -ax splice -k14 --secondary=no. Samtools 1.3.1 48 was used to quality filter SAM files, whereby alignments with MAPQ smaller than 20 (-q 20) were skipped. Further data processing was carried out using the R/Bioconductor environment and custom python scripts.
RNA-seq data preprocessing and antisense bias correction. Paired-end 75 base reads with additional 6 base reads of barcodes were obtained for each of the samples (Supplementary Table 1).  Reads were demultiplexed and mapped with STAR 2.3.0 (Dobin and Gingeras 2015) to the hg20/hg38 (GRCh38) genome assembly (Human Genome Reference Consortium). Samtools (Li, Handsaker et al. 2009) was used to quality filter SAM files, whereby alignments with MAPQ smaller than 7 (-q 7) were skipped and only proper pairs (-f2) were selected. Further data processing was carried out using the R/Bioconductor environment. We used a spike-in (RNAs) normalization strategy essentially as described (Schwalb, Michel et al. 2016) to allow observation of antisense bias ratio  (ratio of spurious reads originating from the opposite strand introduced by the reverse transcription reaction). Read counts (kij) for spike-ins were calculated using HTSeq (Anders, Pyl et al. 2015). The number of transcribed bases (tbj) for all samples was calculated as the sum of the coverage of evident (sequenced) fragment parts (read pairs only) for all fragments with an inner mate interval not entirely overlapping a Refseq annotated intron (UCSC RefSeq GRCh38) in addition to the sum of the coverage of non-evident fragment parts (entire fragment). The number of transcribed bases (tbj) or read counts (kj) for all features (RefSeq-TUs) were corrected for antisense bias cj as follows using the parameter calculated as described above. RPKs were calculated upon antisense bias corrected read counts (kj) falling into the region of a RefSeq-TU divided by its length in kilobases. Coverages were calculated upon antisense bias corrected number of transcribed bases (tbj) falling into the region of a RefSeq-TU divided by its length in bases.
Genome_build: hg20/hg38 (GRCh38) genome assembly (Human Genome Reference Consortium http://hgdownload.soe.ucsc.edu/downloads.html#human)
Supplementary_files_format_and_content: bigwig files containing the coverage

organism: synthetic construct

source_name: ERCC Spike-in Mix

growth_protocol: Human K562 erythroleukemia cells were obtained from DSMZ (Cat. # ACC-10). K562 cells were cultured antibiotic-free in accordance with the DSMZ Cell Culture standards in RPMI 1640 medium (Thermo Fisher Scientific) containing 10 % heat inactivated fetal bovine serum (FBS) (Thermo Fisher Scientific), and 1x GlutaMAX supplement (Thermo Fisher Scientific) at 37 °C in a humidified 5 % CO2 incubator. Cells used in this study display the phenotypic properties, including morphology and proliferation rate, that have been described in literature. Cells were verified to be free of mycoplasma contamination using Plasmo Test Mycoplasma Detection Kit (InvivoGen). Biological replicates were cultured independently.

extract_protocol:
Direct RNA nanopore sequencing library kit (SQK-RNA001, Oxford Nanopore Technologies)
Nanopore: Total RNA was extracted from K562 cells using QIAzol (Quiagen) according to manufacturer’s instructions. Poly(A) RNA was purified from 1 mg of total RNA using the µMACS mRNA Isolation Kit (Milteny Biotec) following the manufacturer’s protocol. The quality of poly(A) RNA selection was assessed using the TapeStation System (Agilent). Poly(A) selected RNAs were subsequently subjected to direct RNA nanopore sequencing library preparation (SQK-RNA001, SQK-RNA002, Oxford Nanopore Technologies) following manufacturer’s protocol with 1000 ng input. All libraries were sequenced on a MinION Mk1B (MIN-101B) for 48 h, unless reads sequenced per second stagnated dramatically.
RNA-seq: Total RNA was extracted from K562 cells using QIAzol according to manufacturer’s instructions except for the addition of 150 ng RNA spike-in mix 25 together with QIAzol. To isolate polyA RNA from 75 µg of total RNA, two subsequent rounds of purification by Dynabeads Oligo (dT)25 (invitrogen) were performed. Purification based on manufacturer’s instructions was performed twice, using 1 mg of Dynabeads Oligo (dT)25 beads for the first round and 0.5 mg for the second round of purification. The quality of polyadenylated RNA selection was assessed using RNA ScreenTape on a TapeStation (Agilent). Sequencing libraries were prepared using the NuGEN Ovation Universal RNA-seq kit according to manufacturer’s instructions. Fragments were amplified by 10 cycles of PCR, and sequenced on an Illumina NextSeq 550 in paired-end mode with 75 bp read length.

treatment_protocol:
Nanopore: K562 cells were kept at low passage numbers (<6) and at optimal densities (3x10^5 - 8x10^5) during all experimental setups. Per biological replicate, K562 cells were diluted 24 h before the experiment was performed (Supplementary Table 1). Per 5EU 60 min sample (6 replicates), cells were incubated at 37 °C, 5 % CO2 for 1 h after a final concentration of 500 µM 5-Ethynyluridine (5EU, Jena Bioscience) was added. Per 5EU 24 h sample (3 replicates), cells were incubated at 37 °C, 5% CO2 for 24 h. 5EU was added 3 times during the 24h incubation, i.e. every 8 hours (0h, 8h, 16h) at a final concentration of 500 µM. Control samples were not labeled (3 replicates). Per 5EU 60 min HS (heat shock) sample (5 replicates), cells were incubated at 42 °C for 5 min (until cell suspension reached 42 °C), and then 5EU was added at a final concentration of 500 µM. Further, heat shock treatments were performed in a water bath (LAUDA, Aqualine AL12) at 42 °C. for 1 h. Temperature was monitored by thermometer. To avoid transcriptional changes by freshly added growth medium, fresh growth medium was added 24 h prior to heat shock treatments 45. Exactly after the labeling duration, cells were centrifuged at 37 °C and 1,500 x g for 2 min.
RNA-seq: Two biological replicates of K562 cells were diluted 24 h before the experiment was performed. Per replicate, 3.6 x 107 cells in growth medium were labeled at a final concentration of 500 µM 4-thio-uracil (4sU, Sigma-Aldrich), and incubated at 37 °C, 5 % CO2 for 5 min. Exactly after 5 min of labeling, cells were harvested at 37 °C and 1,500 x g for 2 min.

tag: method protocol

value: Direct RNA nanopore sequencing (synthetic RNAs)

tag: treatment condition

value: 5EU, 5IU, IVT


title: 5BrU, 4SU, 6SG

description: Labeled RNA

value: 5BrU, 4SU, 6SG, IVT


title: ECs, Ctrl_4

description: unstimulated


title: F2: Sp-Fem-BK 2

tag: strain


title: AurKB_1

tag: tissue

tag: genotype/variation


title: F1: Sp-Fem-BK 1


title: Synthetic control

description: RNA

value: IVT


title: LPM_H3K27ac_ChIP_rep2

status: Public on May 05 2020

city: La Jolla

first: Christopher

middle: K

last: Glass

address: 9500 Gilman Drive

department: School of Medicine

institute: University of California, San Diego

description:
H3K27ac_ChIP
processed data file: KC-H_KC_N_H3k27ac_at_peaks.txt

data_processing:
Preprocessing: FASTQ files from sequencing experiments were mapped to the mouse mm10 genome. Bowtie2 with default parameters was used to map ChIP-seq experiments (Langmead and Salzberg, 2012). HOMER was used to convert aligned reads into “tag directories” for further analysis (Heinz et al., 2010).
LXR ChIP-seq experiments were performed in replicate with corresponding input experiments. Peaks were called with HOMER for each tag directory with relaxed peak finding parameters -L 0 -C 0 -fdr 0.9 against the corresponding input directory. Only peaks with > 16 normalized tags (normalized to total tags of 1e7) were kept. The pooled tag directory from two replicates was used for track visualization.
H3K27ac analysis: To annotate H3K27ac signal around ATAC-seq or LXR ChIP-seq peaks the parameter -size 2000 was used. Subsequently, DESeq2 (Love et al., 2014) was used to identify differential H3K27ac signal or chromatin accessibility with > 2 fold-change and p-adj < 0.05, unless stated otherwise in the text.
To identify motifs enriched in peak regions over the background, HOMER’s motif analysis (findMotifsGenome.pl) including known default motifs and de novo motifs was used. The background sequences were either from random genome sequences or from peaks from the comparing condition indicated in the main text and in the figure legends.
Genome_build: mm10 (GRCm38)
Supplementary_files_format_and_content: Tab-delimited text files of LXR ChIP seq peaks and IDR peaks.
Supplementary_files_format_and_content: KC-H_ATF3_IDR.txt: Peak
Supplementary_files_format_and_content: KC-H_KC_N_H3k27ac_at_peaks.txt: ChIP tag counts at distal ATAC peaks
Supplementary_files_format_and_content: KC-H_LXR_IDR.txt: Peak
Supplementary_files_format_and_content: KC-H_P300_IDR.txt: Peak
Supplementary_files_format_and_content: KC-N_ATF3_IDR.txt: Peak
Supplementary_files_format_and_content: KC-N_LXR_IDR.txt: Peak

source_name: Sorted large peritoneal macrophages

extract_protocol:
Hepatic macrophages (H3K27ac): Mice were humanely euthanized by exposure to CO2 and liver non-parenchymal cells processed for fluorescence activated cell sorting of Kupffer cells, with modifications from published methodology (Mederacke et al., 2015; Muse et al., 2018; Seki et al., 2007). In brief, livers were retrograde perfused for 3 min at a rate of 5-7 ml/min through the inferior vena cava with HBSS without Ca++ or Mg++ supplemented with 0.5 mM EGTA, 0.5 mM EDTA, and 20 mM HEPES. Perfusions were then switched to 40 ml of a digestion buffer, held at 37C, comprised of HBSS with Ca++ and Mg++ supplemented with 0.033 mg/ml of Liberase TM (Roche), 20 μg/ml DNaseI (Worthington), and 20 mM HEPES. Livers were then excised, minced, and digested for an additional 20 minutes in vitro at 37C with gentle rotation in 20 ml of fresh digestion buffer. The perfusion and digestion steps were performed in the presence of 1 μM flavopiridol to offset transcriptional changes associated with digestion. After tissue digestion, cells were passed through a 70-micron cell strainer and hepatocytes removed by 2 low-speed centrifugation steps at 50 X G for 2 min. Nonparenchymal cells in the supernatant were further separated from debris by pelleting for 15 min at 600 X G in 50 ml of 20% isotonic Percoll (Sigma Aldrich) at room temperature. Cells were then washed from Percoll containing buffer and suspended in 10 ml 28% OptiPrep (Sigma Aldrich) and carefully underlaid beneath 3 ml of wash buffer. The resulting gradient was centrifuged at 1,400 X G for 25 minutes at 4C with no break and cells enriched at the interface were saved and subjected to isotonic erythrocyte lysis. Enriched non-parenchymal cells were then washed, suspended in PBS, then stained for 10 minutes with Zombie NIR (BioLegend) and purified anti-CD16/32 (93, BioLegend) to label dead cells and block Fc receptors. Cells were then immunolabeled with specific antibodies of interest, washed, and sorted using a Beckman Coulter MoFlo Astrios EQ configured with spatially separated 355 nm, 405 nm, 488 nm, 561 nm, and 642 nm lasers.
Kupffer cell nuclei (LXR ChIP): Mice were humanely euthanized by exposure to CO2, then briefly perfused with HBSS without Ca++ or Mg++ supplemented with 0.5mM EGTA, 0.5mM EDTA, and 20mM HEPES. Perfusions were then switched sequentially to 1 mg/ml disuccinimidyl glutarate in PBS for 30 minutes, then 1% formaldehyde in PBS for 10 minutes. Finally, the fixation was quenched by perfusion with 20 ml 0.125M glycine. Livers were excised, finely minced with a razor, and washed twice with 20 ml ice cold NF1 buffer (10mM Tris pH 8.0, 5mM MgCl2, 0.1M sucrose, 0.5% Triton X-100) followed by centrifugation 1,200 XG for 7 minutes at 4C. Liver pellets were next suspended in NF1 buffer and homogenized using the “loose” pestle of a Dounce homogenizer for 10 strokes, followed by incubation on ice for 30 minutes. Samples were finally homogenized using the “tight” pestle of a Dounce homogenizer for an additional 50-70 strokes, with periodic assessment for released nuclei by microscopy. The homogenized liver was then strained through a 70-micron mesh strainer, centrifuged at 1,200 XG for 7 minutes at 4C, then washed once more in PBS. Finally, the homogenized liver was suspended in PBS with 2mM EDTA and passed through a 40-micron mesh strainer. Nuclei were purified by FACS using a Sony SH800 based on TdTomato expression and forward scatter. Acquired nuclei were snap frozen in a dry ice ethanol bath and stored at -80C prior to use in ChIP-seq experiments.
Peritoneal macrophages: As described in Gosselin et al. 2014, Cell PMID 25480297.
ChIP protocol - H3K27ac: ChIP for H3K27ac was performed essentially as describe previously (Eichenfield et al., 2016). In brief, FACS purified cells were fixed with 1% paraformaldehyde for 10 minutes at room temperature. Next, 2.625 M glycine was added to 125 mM to quench fixation and cells were collected by centrifugation with the addition of 0.01% Tween-20 at 1,200 X G for 10 minutes at 4C. Cells were washed once with 0.01 % Tween-20 in PBS and collected by centrifugation at 1,200 X G for 10 minutes at 4C. Cell pellets were then snap frozen and stored at -80C. For ChIP reactions, cell pellets were thawed on ice and lysed in 80 μl LB3 (10 mM Tris/HCl pH 7.5, 100 mM NaCl, 1 mM EDTA, 0.5mM EGTA, 0.1% deoxcycholate, 0.5% sarkosyl, 1 × protease inhibitor cocktail, and 1 mM sodium butyrate). Lysate was sonicated using a Covaris for 12 cycles with the following setting: time, 60 seconds; duty, 5.0; PIP, 140; cycles, 200; amplitude, 0.0; velocity, 0.0; dwell, 0.0. Samples were collected and 10% Triton X-100 was added to 1% final concentration. One percent of the sonicated lysate was saved as a ChIP input. For each chromatin immunoprecipitation, aliquots of ~500,000 cells were added to 20 μl Dynabeads Protein A with 2 μg anti-H3K27ac (Active Motif) and incubated with slow rotation at 4C overnight. The following day, beads were collected using a magnet and washed three times each with wash buffer I (20 mM Tris/HCl pH 7.5, 150 mM NaCl, 1% Triton X-100, 0.1% SDS, 2 mM EDTA, and 1 × protease inhibitor cocktail) and wash buffer III (10 mM Tris/HCl pH 7.5, 250 mM LiCl, 1% Triton X-100, 0.7% Deoxycholate, 1 mM EDTA, and 1 × protease inhibitor cocktail). Beads were then washed twice with ice cold 10 mM Tris/HCl pH 7.5, 1 mM EDTA, 0.2% Tween-20. Sequencing libraries were prepared for ChIP products while bound to the Dynabeads Protein A initially suspended in 25 μl 10 mM Tris/HCl pH 8.0 and 0.05% Tween-20.
ChIP protocol - LXR: For LXR ChIP-seq, FACS purified Kupffer cell nuclei were resuspended in wash buffer (10mM HEPES/KOH pH7.9, 85mM KCl, 1mM EDTA, 0.2% IGEPAL CA-630, 1x protease inhibitor cocktail (Sigma), 1 mM PMSF) for 5 minutes on ice. Nuclei were spun down and resuspended in 130 μl RIPA-NR1 lysis buffer (20 mM Tris/HCl pH7.5, 1 mM EDTA, 0.5 mM EGTA, 0.1% SDS, 0.4% Na-Deoxycholate, 1% NP-40 alternative, 0.5 mM DTT, 1x protease inhibitor cocktail (Sigma), 1 mM PMSF) and chromatin was sheared by sonication. Samples were sonicated in a 96 Place microTUBE Rack (Covaris cat#500282) using a Covaris E220 for 18 cycles with the following setting: time, 60 seconds; duty, 5.0; PIP, 140; cycles, 200; amplitude, 0.0; velocity, 0.0; dwell, 0.0. Samples were recovered and spun down at max speed, 4°C for 10 minutes. 1% supernatant was taken as input DNA and remaining supernatant was transferred to PCR strips and brought up to a volume of 200 μl using RIPA-NR1 lysis buffer (20 mM Tris/HCl pH7.5, 1 mM EDTA, 0.5 mM EGTA, 0.1% SDS, 0.4% Na-Deoxycholate, 1% NP-40 alternative, 0.5 mM DTT, 1x protease 37 inhibitor cocktail (Sigma), 1 mM PMSF). 30 μl Dynabeads Protein A/G coated with 2 μg each of the indicated LXR specific antibodies (Santa Cruz Biotechnology: sc-1000X, sc- 133221X, sc-271064X) was added to the sample, and immunoprecipitation was carried out with slow rotation at 4°C overnight. Beads were then collected using a magnet and washed with 175 μl ice cold buffer as indicated by incubating samples on ice for 3 minutes: 3 times RIPA-NR1 lysis buffer (20 mM Tris/HCl pH7.5, 1 mM EDTA, 0.5 mM EGTA, 0.1% SDS, 0.4% Na-Deoxycholate, 1% NP-40 alternative, 0.5 mM DTT, 1x protease inhibitor cocktail (Sigma), 1 mM PMSF), 6 times LiCl-NR1 buffer (10 mM Tris/HCl pH7.5, 250mM LiCl, 1 mM EDTA, 0.7% Na-Deoxycholate, 1% NP-40 alternative, 1x protease inhibitor cocktail (Sigma), 1 mM PMSF), 3 times TET (10 mM Tris/HCl pH 8.0, 1 mM EDTA, 0.2% Tween-20, 1x protease inhibitor cocktail (Sigma), 1 mM PMSF), and 1 time IDTE (10 mM Tris/HCl pH 8.0, 0.1 mM EDTA, 1x protease inhibitor cocktail (Sigma), 1 mM PMSF). Bead complexes were resuspended in 25 μl TT (10 mM Tris/HCl pH 8.0, 0.05% Tween-20, 1x protease inhibitor cocktail) and sequencing libraries were prepared on-bead as described below.
ChIP libraries were prepared while bound to Dynabeads using NEBNext Ultra II Library preparation kit with reaction

value: Sorted large peritoneal macrophages

tag: cell subtype

value: Large peritoneal macrophages

value: Control diet

tag: chip antibody

value: H3K27ac (Active Motif 39133, RRID:AB_2561016)


title: Lung ST2+ [RRa1-10]

city: BOSTON

first: ROD

middle: A

last: (MD

state: MA- MASSACHUSETTS

address: 55 FRUIT ST., BULFINCH-148

department: Medicine

institute: MASSACHUSETTS GENERAL HOSPITAL

data_processing:
Transcriptome mapping was performed with STAR (PMID:23104886) using the Ensembl annotation of mm9 reference genome. Read counts for individual genes were produced using HTSeq (PMID:25260700).
Read counts for individual genes were produced using HTSeq (PMID:25260700). Differential expression analysis was performed using the EdgeR package (PMID:19910308) after normalizing read counts and including only those genes with CPM > 1 for at least one out of all samples.
mm9

source_name: Lung memory CD4+ST2+ T cells

extract_protocol:
Lungs and secondary lymphoid organs (LN and spleen) were harvested, minced, and digsted with collagenase/liberase followed by generation of a single cell suspension. CD4+ST2+ and CD4+ST2- T cells were isolated from the lung parenchyma and secondary lymphoid organs via fluorescence-acivated cell sorting. RNA was isolated using a Qiagen RNeasy Micro Kit (Cat. No. 74004).
Library construction was performed using the Clontech SMARTer kit.

treatment_protocol: We sensitized and challenged C57BL/6 mice with intranasal house dust mite extract to generate memory CD4+ T helper type 2 (Th2) cells. Approximately 6 weeks after treatment, mice were administered intravenous, fluorophore-labelled anti-CD45 antibody to label intravascular leukocytes prior to harvest.

value: intranasal house dust mite extract

tag: time point

value: ~6 weeks after treatment

value: Lung memory CD4+ST2+ T cells


title: SLO ST2- [RRa1-11]

source_name: SLO memory CD4+ST2- T cells

value: SLO memory CD4+ST2- T cells


title: SLO ST2+ [RRa1-4]

source_name: SLO memory CD4+ST2+ T cells

value: SLO memory CD4+ST2+ T cells


title: SLO ST2+ [RRa1-12]


title: SLO ST2+ [RRa1-8]


title: Lung ST2- [RRa1-9]

source_name: Lung memory CD4+ST2- T cells

value: Lung memory CD4+ST2- T cells


title: Lung ST2+ [RRa1-6]


title: CD45PosCD146Neg_control_rep1_scRNAseq_10XGenomics

data_processing:
Data was mapped using Cell Ranger (10X Genomics) to mm10 and analyzed using the Seurat R toolkit for single cell genomics (Satija et al., 2015).
Supplementary_files_format_and_content: cell bar codes tsv, gene name tsv, UMI matrix

source_name: CD45Pos CD146Neg hepatic immune cells from control mice

extract_protocol:
Mice were humanely euthanized by exposure to CO2 and liver non-parenchymal cells processed for fluorescence activated cell sorting of Kupffer cells, with modifications from published methodology (Mederacke et al., 2015; Muse et al., 2018; Seki et al., 2007). In brief, livers were retrograde perfused for 3 min at a rate of 5-7 ml/min through the inferior vena cava with HBSS without Ca++ or Mg++ supplemented with 0.5 mM EGTA, 0.5 mM EDTA, and 20 mM HEPES. Perfusions were then switched to 40 ml of a digestion buffer, held at 37C, comprised of HBSS with Ca++ and Mg++ supplemented with 0.033 mg/ml of Liberase TM (Roche), 20 μg/ml DNaseI (Worthington), and 20 mM HEPES. Livers were then excised, minced, and digested for an additional 20 minutes in vitro at 37C with gentle rotation in 20 ml of fresh digestion buffer. The perfusion and digestion steps were performed in the presence of 1 μM flavopiridol to offset transcriptional changes associated with digestion. After tissue digestion, cells were passed through a 70-micron cell strainer and hepatocytes removed by 2 low-speed centrifugation steps at 50 X G for 2 min. Nonparenchymal cells in the supernatant were further separated from debris by pelleting for 15 min at 600 X G in 50 ml of 20% isotonic Percoll (Sigma Aldrich) at room temperature. Cells were then washed from Percoll containing buffer and suspended in 10 ml 28% OptiPrep (Sigma Aldrich) and carefully underlaid beneath 3 ml of wash buffer. The resulting gradient was centrifuged at 1,400 X G for 25 minutes at 4C with no break and cells enriched at the interface were saved and subjected to isotonic erythrocyte lysis. Enriched non-parenchymal cells were then washed, suspended in PBS, then stained for 10 minutes with Zombie NIR (BioLegend) and purified anti-CD16/32 (93, BioLegend) to label dead cells and block Fc receptors. Cells were then immunolabeled with specific antibodies of interest, washed, and sorted using a Beckman Coulter MoFlo Astrios EQ configured with spatially separated 355 nm, 405 nm, 488 nm, 561 nm, and 642 nm lasers on the basis of CD45Pos, CD146Neg (to exclude endothelial cells) and viability (negative for the live/dead dye Zombie NIR).
Purified cell preparations were washed twice with PBS and suspended in PBS supplemented with 400 μg/ml non-acetylated bovine serum albumin (Sigma Aldrich, B6917). Individual cells were then coupled to beads using the 10X Genomics Chromium controller. Single-cell sequencing libraries were prepared following the 10X Genomics Protocol using v2 chemistry and sequenced to a median depth of approximately 50,000 reads per cell using an Illumina HiSeq 2500 on rapid run.

tag: strain background

value: liver

tag: diet

value: control

value: Bulk CD45+ CD146- hepatic immune cells


title: CD45PosCD146Neg_nash_rep1_scRNAseq_10XGenomics

source_name: CD45Pos CD146Neg hepatic immune cells from nash mice

value: nash-amln


title: CD45PosCD146Neg_nash_rep2_scRNAseq_10XGenomics


title: CD45PosCD146Neg_control_rep2_scRNAseq_10XGenomics


title: Lung ST2+ [RRa1-2]


title: SLO ST2- [RRa1-3]


title: Lung ST2- [RRa1-5]


title: EJ37

city: Montreal

first: Cindy

last: Audiger

state: Québec

address: 2900, boul. Édouard-Montpetit

country: Canada

institute: Univeristy of Montreal

data_processing:
illumine HiSeq system
Sequence reads were then aligned to the genome by using a standard illumine sequence analysis pipeline by the Laboratory of Statistical Genomics and Systems Biology at the University of Cincinnati
To study differential gene expression, individually indexed and compatible libraries were proportionally pooled (20-50 million reads per sample) for clustering in cBot system (Illumina, San Diego, CA, USA)
Libraries at the final concentration of 15pM was clustered onto a single read flow cell using TruSeq SR Cluster Kit v3, and sequenced for 50bp using TruSeq SBS kit on an Illumina HiSeq system.
Sequence reads were then aligned to the genome by using a standard illumine sequence analysis pipeline by the Laboratory of Statistical Genomics and Systems Biology at the University of Cincinnati
Genome_build: mm8
Supplementary_files_format_and_content: tab-delimited text file include RPKM values for the 9 samples. Both gene ID and sample name are depicted

source_name: spleen

extract_protocol:
RNA was extracted according to the mir-VANA RNA extraction kit protocol
The NEBNext Ultra Directional RNA library Prep Kit was used for library preparation.
Bioanalyzer (Agilent, Santa Clara, CA) analyses using a DNA high sensitivity chip were performed to determine quality and yield f the purified library

tag: age

value: 8-12 weeks old

value: spleen

value: mcDC


title: Lung ST2- [RRa1-1]


title: SLO ST2- [RRa1-7]


title: CC060_F_Ctr-2

city: Jefferson

first: Volodymyr

last: Tryndyak

address: 3900 NCTR Rd

department: Division of Biochemical Toxicology

institute: US-FDA National Center for Toxicological Research

data_processing:
Sequencing was performed in an Illumina NextSeq 500 sequencing machine (Illumina) at a 2 × 75 bp paired-end resolution.
Sequence reads were mapped to the mouse reference genome (NCBI38/mm10) using TopHat v2.1.1 with Bowtie2 v2.2.5.0 (Trapnell et. al., Bioinformatics. 2009) with default parameters.
Transcript abundance estimates, normalizations and differential gene expressions were generated using Cufflinks v2.2.1 (Trapnell et. al., Nat. Biotechnol. 2010) with default parameters.
Genome_build: mm10
Supplementary_files_format_and_content: tab-delimited text files include normalized FPKM values for each Sample.

source_name: Liver

extract_protocol:
Livers were removed and flash frozen on dry ice. Total RNA was extracted from liver tissue samples using miRNeasy Mini kits (Qiagen, Valencia, CA, USA).
RNA libraries for RNA-sequencing were prepared from 500 ng RNA using Illumina TruSeq Stranded Total RNA library preparation kits with Ribo-Zero Gold for rRNA depletion and TruSeq Stranded mRNA library preparation kits as recommended in the manufacturer’s protocol.

treatment_protocol: Mice in the experimental group were maintained on the obesogenic high-fat and high-sucrose (HF/HS) diets (Teklad Custom Diet - TD.88137, Envigo, Madison, WI, USA) for 12 weeks.  This diet contains 42% Kcal from milkfat high in palmitic acid, 0.2% of total cholesterol, and is high in sucrose (>30%).  Mice in the control group received an ingredient-matched low-fat and reduced sucrose diet (Teklad Custom Diet – TD.08485, Envigo).

value: Liver

tag: Sex

value: Female

tag: time of treatment

value: 12 weeks


title: CC060_M_Ctr-2

value: Male


title: CC060_M_HFD-2

value: HF/HS diet-2


title: CC060_M_HFD-3

value: HF/HS diet-3


title: CC060_M_Ctr-3


title: CC060_F_Ctr-3


title: CC060_M_Ctr-1


title: CC060_M_HFD-1

value: HF/HS diet-1


title: CC043_M_HFD-3


title: CC060_F_Ctr-1


title: CC043_M_HFD-2


title: CC043_M_Ctr-2


title: CC043_F_HFD-3


title: CC043_F_HFD-2


title: CC043_M_Ctr-3


title: CC043_F_Ctr-3


title: CC042_M_HFD-3


title: CC042_M_HFD-2


title: CC043_M_HFD-1


title: CC042_M_HFD-1


title: CC060_F_HFD-3


title: CC060_F_HFD-1


title: CC060_F_HFD-2


title: CC043_F_HFD-1


title: CC042_M_Ctr-1


title: CC042_M_Ctr-2


title: CC042_F_HFD-3


title: CC042_F_HFD-1


title: CC043_F_Ctr-2


title: CC042_F_HFD-2


title: CC042_F_Ctr-3


title: CC043_M_Ctr-1


title: CC041_M_HFD-3


title: CC041_M_HFD-2


title: CC041_M_HFD-1


title: CC042_F_Ctr-1


title: CC041_M_Ctr-2


title: CC042_F_Ctr-2


title: CC041_M_Ctr-1


title: CC043_F_Ctr-1


title: CC041_M_Ctr-3


title: CC041_F_HFD-2


title: CC041_F_HFD-3


title: CC042_M_Ctr-3


title: CC040_M_HFD-1


title: CC041_F_HFD-1


title: CC041_F_Ctr-2


title: CC041_F_Ctr-3


title: CC040_M_HFD-2


title: CC040_M_HFD-3


title: CC040_M_Ctr-3


title: CC041_F_Ctr-1


title: CC040_M_Ctr-2


title: CC040_M_Ctr-1


title: CC040_F_HFD-2


title: CC040_F_Ctr-3


title: CC040_F_HFD-1


title: CC040_F_HFD-3


title: CC040_F_Ctr-2


title: CC032_M_HFD-2


title: CC040_F_Ctr-1


title: CC032_M_HFD-3


title: CC032_M_HFD-1


title: CC032_F_HFD-2


title: CC032_F_HFD-3


title: CC032_M_Ctr-3


title: CC032_F_Ctr-3


title: CC032_M_Ctr-2


title: CC032_M_Ctr-1


title: CC032_F_HFD-1


title: CC032_F_Ctr-1


title: CC019_M_HFD-3


title: CC019_M_HFD-2


title: CC032_F_Ctr-2


title: CC019_M_Ctr-3


title: CC019_F_HFD-2


title: CC019_F_HFD-3


title: CC019_F_Ctr-3


title: CC019_F_Ctr-1


title: CC019_M_Ctr-2


title: CC019_F_Ctr-2


title: CC019_M_Ctr-1


title: CC013_M_Ctr-3


title: CC013_M_HFD-1


title: CC013_M_HFD-2


title: CC013_F_HFD-3


title: CC013_M_Ctr-1


title: CC013_M_HFD-3


title: CC019_F_HFD-1


title: CC013_F_HFD-2


title: CC013_F_HFD-1


title: CC013_F_Ctr-3


title: CC019_M_HFD-1


title: HSD BEACON1 mouse embryo#8;BEACON2:PMS rep c

city: Shanghai

first: Li

last: Yang

address: 131 Dong-An Road

department: Institutes of Biological Sciences

country: China

institute: Fudan University

data_processing:
Library strategy: DNA-Seq
Basecalling using Illumina Casava1.8.2 software.
High-throughput sequencing reads were separated according to 6-nt experimental barcodes.
Reads were aligned against the GRCh38/hg38 human or GRCm38/mm10 mouse reference genome using BWA-MEM 0.7.9a.
Genome_build: hg38 for human samples and mm10 for mouse samples
Supplementary_files_format_and_content: Excel for Indels and Base Subtitutions

organism:
Homo sapiens
Mus musculus

source_name: HEK293T cells and mouse embryos

growth_protocol: HEK293T and U2OS cells from ATCC were maintained in DMEM (10566, Gibco/Thermo Fisher Scientific) + 10% FBS (16000-044, Gibco/Thermo Fisher Scientific) and regularly tested to exclude mycoplasma contamination.

extract_protocol:
For DNA, after 72 hr, the genomic DNA was extracted from the cells with QuickExtractTM DNA Extraction Solution (QE09050, Epicentre) or the cells were lysed in 2× SDS loading buffer for western blot.
For DNA, DNA-seq libraries were prepared with NEB Next Ultra™ II  FS DNA Library Prep Kit. Targeted deep sequencing was performed on Illumina Hiseq X ten or NextSeq 500 at CAS-MPG Partner Institute for Computational Biology Omics Core, Shanghai, China.

treatment_protocol: For base editing in genomic DNA, HEK293T and U2OS cells were seeded in a 24-well plate at a density of 1.6 × 105 per well and transfected with 200 μl serum-free Opti-MEM that contained 5.04 μl LIPOFECTAMINE LTX (Life, Invitrogen), 1.68 μl LIPOFECTAMINE plus (Life, Invitrogen), 1 μg dCpf1-BE expression vector (or hA3A-dCas12a-BE, hA3B-dCas12a-BE, hAID-dCas12a-BE, hA3AW98A-dCas12a-BE, hA3AW98Y-dCas12a-BE, hA3AW104A-dCas12a-BE, hA3AW104Y-dCas12a-BE, hA3AP134A-dCas12a-BE, hA3AP134Y-dCas12a-BE, hA3AW98Y/W104A-dCas12a-BE, hA3AW98Y/P134Y-dCas12a-BE, hA3AW104A/P134Y-dCas12a-BE, hA3AW98Y/W104A/P134Y-dCas12a-BE, hA3AW104A-dCas12a-BE-op, hA3AW104A/Y130F-dCas12a-BE-op, hA3AW104A/Y132D-dCas12a-BE-op, hA3AW98Y/W104A-dCas12a-BE-op, hA3AW98Y/W104A/Y130F-dCas12a-BE-op, hA3AW98Y/W104A/Y132D-dCas12a-BE-op, hA3AW104A/P134Y-dCas12a-BE-op, hA3AW104A/P134Y/Y130F-dCas12a-BE-op, hA3AW104A/P134Y/Y132D-dCas12a-BE-op and hA3AW98Y/W104A/Y130F/Y132D-dCas12a-BE-op expression vector) and 0.68 μg crRNA or sgRNA expression vector. After 72 hr, the genomic DNA was extracted from the cells with QuickExtractTM DNA Extraction Solution (QE09050, Epicentre) or the cells were lysed in 2× SDS loading buffer for western blot.

tag: cell line

value: mouse embryos

value: BEACON1 and BEACON2

tag: 5'-end trimming of r1

tag: 3'-end trimming of r1


title: HSD BEACON1 mouse embryo#6;BEACON1:S6, S4, RUN, DYR, CFT, CDK rep b;BEACON2:PMS, PAF, CHD rep b

tag: treatment

value: BEACON1 and BEACON1 and BEACON2


title: CC013_F_Ctr-1


title: CC011_M_HFD-1


title: CC011_M_HFD-3


title: CC011_M_Ctr-3


title: HSD BEACON1 mouse embryo#5;BEACON1:S6, S4, RUN, DYR, CFT, CDK rep a;BEACON2:PMS, PAF, CHD rep a


title: CC013_M_Ctr-2


title: CC011_F_HFD-3


title: hA3A(W104A/Y130F)-dCas12a-BE-op:CFT, CDK rep c; HSD BEACON1 mouse embryo#4;rA1-dCas12a-BE:BLM rep c

value: hA3A(W104A/Y130F)-dCas12a-BE-op and BEACON1 and rA1-dCas12a-BE


title: CC011_M_Ctr-1


title: hA3A(W104A/Y130F)-dCas12a-BE-op:CFT,S6,S4,RUN,DYR; rA1-dCas12a-BE:BLM, SPA, PMS, PDE, PAF, CHD, BMR;HSD BEACON1 embryo#3

value: hA3A(W104A/Y130F)-dCas12a-BE-op and BEACON1 and hA3A(W104A/Y130F)-dCas12a-BE-op and rA1-dCas12a-BE and rA1-dCas12a-BE


title: hA3A(W104A/Y130F)-dCas12a-BE-op:CFT,S6,S4,RUN,DYR,CDK; rA1-dCas12a-BE:SPA, PMS, PDE, PAF, CHD, BMR;HSD BEACON1 embryo#2

value: hA3A(W104A/Y130F)-dCas12a-BE-op and BEACON1 and hA3A(W104A/Y130F)-dCas12a-BE-op and rA1-dCas12a-BE


title: CC011_F_Ctr-3


title: CC011_F_HFD-1


title: CC011_F_Ctr-2


title: HSD BEACON1 embryo#1;hA3A(W104A/Y130F)-dCas12a-BE-op:S6,S4,RUN,DYR,CDK;rA1-dCas12a-BE:SPA,PMS,PDE,PAF,CHD,BMR,BLM

value: BEACON1 and hA3A(W104A/Y130F)-dCas12a-BE-op and rA1-dCas12a-BE


title: CC011_M_Ctr-2


title: hA3A(W104A)-dCas12a-BE:CDK rep d;S6 rep e; AncBE4max: SPA, PMS, PDE, PAF, BMR rep a; HSD BEACON1 mouse kindey#7

source_name: HEK293T cells and mouse kindey

value: mouse kindey

value: hA3A(W104A)-dCas12a-BE and BEACON1 and hA3A(W104A)-dCas12a-BE and AncBE4max


title: CC013_F_Ctr-2


title: CC011_M_HFD-2


title: CC003_M_HFD-3


title: hA3A(W104A)-dCas12a-BE:CDK rep e;AncBE4max:CHD, BLM rep a;SPA,PMS,PDE,PAF,BMR rep b;DYR rep d;HSD BEACON1 mouse brain#7

source_name: HEK293T cells and mouse brain

value: mouse brain

value: hA3A(W104A)-dCas12a-BE and hA3A(W104A)-dCas12a-BE and BEACON1 and AncBE4max and AncBE4max


title: HSD BEACON1 mouse lung#7;  BE3:SPA, PMS, PDE, PAF, CHD rep c

source_name: HEK293T cells and mouse lung

value: mouse lung

value: BEACON1 and BE3


title: HSD BEACON1 mouse embryo#7;BEACON1:S6, S4, RUN, DYR, CFT, CDK rep c;BEACON2:PAF, CHD rep c


title: CC011_F_HFD-2

value: Liver

value: Female

value: 12 weeks


title: CC003_M_HFD-2

value: Male

value: HF/HS diet-2


title: HSD rA1-dCas12a-BE mouse embryo#9; hA3A(W104A)-dCas12a-BE:S6, DYR, CDK rep f; AncBE4max:CHD, BLM rep c

value: mouse embryos

value: rA1-dCas12a-BE and hA3A(W104A)-dCas12a-BE and AncBE4max


title: CC003_M_HFD-1

value: HF/HS diet-1


title: CC011_F_Ctr-1


title: CC003_M_Ctr-3


title: HSD BEACON1 mouse spleen#7; BE3:BMR, BLM BE3 rep c; SPA, PMS, PDE, PAF, CHD rep b

source_name: HEK293T cells and mouse spleen

value: mouse spleen

value: BEACON1 and BE3 and BE3


title: HSD BEACON1 mouse liver#7; BE3:BMR, BLM BE3 rep b; SPA, PMS, PDE rep a

source_name: HEK293T cells and mouse liver

value: mouse liver


title: hA3A(W104A)-dCas12a-BE:DYR rep e;AncBE4max:CHD, BLM rep b; SPA, PMS, PDE, PAF, BMR rep c; HSD BEACON1 mouse testis#7

source_name: HEK293T cells and mouse testis

value: mouse testis

value: hA3A(W104A)-dCas12a-BE and BEACON1 and AncBE4max and AncBE4max


title: CC003_M_Ctr-2


title: HSD BEACON1 mouse heart#7; BE3:PAF, CHD, BMR, BLM rep a

source_name: HEK293T cells and mouse heart

value: mouse heart


title: CC003_M_Ctr-1


title: CC003_F_HFD-3

value: HF/HS diet-3


title: HSD rA1-dCas12a-BE mouse embryo#6; hA3A(W104A/Y132D/P134Y)-dCas12a-BE-op: S6, S4, RUN, DYR, CFT, CDK rep c


title: CC003_F_Ctr-3


title: HSD rA1-dCas12a-BE mouse embryo#5; hA3A(W104A/Y130F/P134Y)-dCas12a-BE-op:S6, S4, RUN, DYR, CFT rep c


title: HSD rA1-dCas12a-BE mouse embryo#4; hA3A(W98Y/W104A/Y132D)-dCas12a-BE-op:S6, S4, RUN, DYR, CFT, CDK rep c


title: HSD rA1-dCas12a-BE mouse embryo#3; BEACON2:S4, RUN, CFT rep c

value: rA1-dCas12a-BE and BEACON2


title: CC003_F_Ctr-2


title: CC003_F_Ctr-1


title: HSD rA1-dCas12a-BE mouse embryo#1; hA3A(W98Y/W104A)-dCas12a-BE-op:S4, DYR, CFT, CDK rep c


title: CC003_F_HFD-2


title: HSD BEACON1 mouse tail#24; hA3A(W98Y/W104A/Y132D)-dCas12a-BE-op:S6, S4, RUN, DYR , CFT, CDK rep b

source_name: HEK293T cells and mouse tail

value: mouse tail


title: HSD BEACON1 mouse tail#22; hA3A(W104A/P134Y)-dCas12a-BE-op:RUN, DYR rep c; S6, S4, CFT, CDK rep b

value: BEACON1 and hA3A(W104A/P134Y)-dCas12a-BE-op and hA3A(W104A/P134Y)-dCas12a-BE-op


title: KC-N_InputForATF3_ChIP_rep2

source_name: Sorted hepatic macrophages

tag: cell type

value: Sorted hepatic macrophages

value: F4/80-Hi Tim4-Pos Kupffer cells

value: NASH diet


title: HSD BEACON1 mouse tail#23; BEACON2:S4, RUN, CFT rep b;S6, DYR, CDK rep c

value: BEACON1 and BEACON2 and BEACON2


title: KC-N_H3K27ac_ChIP_rep2

value: H3K27ac (Active Motif 39133, RRID:AB_2561016)


title: hA3A(W98Y/W104A/Y130F/Y132D)-dCas12a-BE-op:CDK rep b; S6, S4, RUN, DYR, CFT rep a; HSD BEACON1 mouse tail#15

value: hA3A(W98Y/W104A/Y130F/Y132D)-dCas12a-BE-op and BEACON1 and hA3A(W98Y/W104A/Y130F/Y132D)-dCas12a-BE-op


title: KC-N_InputForATF3_ChIP_rep1


title: KC-N_ATF3_ChIP_rep2

description:
ATF3_ChIP
processed data file: KC-N_ATF3_IDR.txt


title: KC-H_InputForATF3_ChIP_rep2

value: Control diet


title: KC-N_H3K27ac_ChIP_rep1


title: hA3A(W104A/Y132D/P134Y)-dCas12a-BE-op:DYR  rep b;S6, S4, RUN, CFT rep a; HSD BEACON1 mouse tail#14

value: hA3A(W104A/Y132D/P134Y)-dCas12a-BE-op and BEACON1 and hA3A(W104A/Y132D/P134Y)-dCas12a-BE-op


title: HSD rA1-dCas12a-BE mouse embryo#7; hA3A(W98Y/W104A/Y130F/Y132D)-dCas12a-BE-op: S6, S4, RUN, DYR, CFT rep c

value: mouse embryos


title: hA3A(W104A/Y130F/P134Y)-dCas12a-BE-op:CDK rep b;S6, S4, RUN, DYR, CFT; HSD BEACON1 mouse tail#13

value: hA3A(W104A/Y130F/P134Y)-dCas12a-BE-op and BEACON1 and hA3A(W104A/Y130F/P134Y)-dCas12a-BE-op


title: CC003_F_HFD-1

value: Liver

value: Female

value: HF/HS diet-1

value: 12 weeks


title: HSD rA1-dCas12a-BE mouse embryo#8


title: HSD BEACON1 mouse tail#21; hA3A(W98Y/W104A)-dCas12a-BE-op:S4, DYR, CFT, CDK rep b;S6, RUN rep c

value: BEACON1 and hA3A(W98Y/W104A)-dCas12a-BE-op and hA3A(W98Y/W104A)-dCas12a-BE-op


title: KC-H_InputForATF3_ChIP_rep1


title: HSD rA1-dCas12a-BE mouse embryo#2; hA3A(W104A/P134Y)-dCas12a-BE-op:S6, S4, CFT, CDK rep c


title: HSD BEACON1 mouse tail#12; hA3A(W98Y/W104A/Y132D)-dCas12a-BE-op:S4, RUN, DYR, CFT rep a

value: mouse tail


title: HSD BEACON1 mouse tail#10; hA3A(W104A/P134Y)-dCas12a-BE-op:RUN, DYR rep b;S6, CDK rep a

value: BEACON1 and hA3A(W104A/P134Y)-dCas12a-BE-op and hA3A(W104A/P134Y)-dCas12a-BE-op


title: hA3A(W98Y/W104A)-dCas12a-BE-op:DYR, CFT, CDK rep a;S6, RUN rep b; HSD BEACON1 mouse tail#9;

value: hA3A(W98Y/W104A)-dCas12a-BE-op and BEACON1 and hA3A(W98Y/W104A)-dCas12a-BE-op


title: HSD BEACON1 mouse tail#11; BEACON2:S4, RUN, CFT  rep a;S6, DYR, CDK rep b


title: KC-H_ATF3_ChIP_rep1

description:
ATF3_ChIP
processed data file: KC-H_ATF3_IDR.txt

value: Sorted hepatic macrophages

value: F4/80-Hi Tim4-Pos Kupffer cells


title: KC-N_ATF3_ChIP_rep1

value: NASH diet


title: ATAC_KC-N_rep1

description: processed data file: KC-N_ATAC_IDR.txt

data_processing:
FASTQ files from sequencing experiments were mapped to the mouse mm10 genome. Bowtie2 with default parameters was used to map ATAC-seq experiments (Langmead and Salzberg, 2012). HOMER was used to convert aligned reads into “tag directories” for further analysis (Heinz et al., 2010).
IDR analysis: ATAC-seq experiments were performed in replicate. Peaks were called with HOMER for each tag directory with parameters -L 0 -C 0 -fdr 0.9 -minDist 200 - size 200. IDR (Li et al., 2011) was used to test for reproducibility between replicates, and only peaks with IDR < 0.05 were used for downstream analysis. The pooled tag directory from two replicates was used for track visualization.
To quantify transcription factor chromatin accessibility, peak files were merged with HOMER’s mergePeaks and annotated with raw tag counts with HOMER’s annotatePeaks using parameters -noadj, -size given. To annotate H3K27ac signal around ATAC-seq peaks the parameter -size 2000 was used. Subsequently, DESeq2 (Love et al., 2014) was used to identify the differential H3K27ac signal or chromatin accessibility with > 2 fold-change and p-adj < 0.05, unless stated otherwise in the text.
To identify motifs enriched in peak regions over the background, HOMER’s motif analysis (findMotifsGenome.pl) including known default motifs and de novo motifs was used. The background sequences were either from random genome sequences or from peaks from the comparing condition indicated in the main text and in the figure legends.
Genome_build: mm10 (GRCm38)
Supplementary_files_format_and_content: *_IDR.txt: Tab-delimited text files include IDR peak files for each set of replicates (as described above) as well as peak files on each individual replicate with HOMER with parameters -minDist 200 - size 200.

extract_protocol:
Hepatic macrophages: Mice were humanely euthanized by exposure to CO2 and liver non-parenchymal cells processed for fluorescence activated cell sorting of Kupffer cells, with modifications from published methodology (Mederacke et al., 2015; Muse et al., 2018; Seki et al., 2007). In brief, livers were retrograde perfused for 3 min at a rate of 5-7 ml/min through the inferior vena cava with HBSS without Ca++ or Mg++ supplemented with 0.5 mM EGTA, 0.5 mM EDTA, and 20 mM HEPES. Perfusions were then switched to 40 ml of a digestion buffer, held at 37C, comprised of HBSS with Ca++ and Mg++ supplemented with 0.033 mg/ml of Liberase TM (Roche), 20 μg/ml DNaseI (Worthington), and 20 mM HEPES. Livers were then excised, minced, and digested for an additional 20 minutes in vitro at 37C with gentle rotation in 20 ml of fresh digestion buffer. The perfusion and digestion steps were performed in the presence of 1 μM flavopiridol to offset transcriptional changes associated with digestion. After tissue digestion, cells were passed through a 70-micron cell strainer and hepatocytes removed by 2 low-speed centrifugation steps at 50 X G for 2 min. Nonparenchymal cells in the supernatant were further separated from debris by pelleting for 15 min at 600 X G in 50 ml of 20% isotonic Percoll (Sigma Aldrich) at room temperature. Cells were then washed from Percoll containing buffer and suspended in 10 ml 28% OptiPrep (Sigma Aldrich) and carefully underlaid beneath 3 ml of wash buffer. The resulting gradient was centrifuged at 1,400 X G for 25 minutes at 4C with no break and cells enriched at the interface were saved and subjected to isotonic erythrocyte lysis. Enriched non-parenchymal cells were then washed, suspended in PBS, then stained for 10 minutes with Zombie NIR (BioLegend) and purified anti-CD16/32 (93, BioLegend) to label dead cells and block Fc receptors. Cells were then immunolabeled with specific antibodies of interest, washed, and sorted using a Beckman Coulter MoFlo Astrios EQ configured with spatially separated 355 nm, 405 nm, 488 nm, 561 nm, and 642 nm lasers.
Peripheral monocytes: Mice were humanely euthanized by exposure to CO2. Blood was collected from mice via cardiac puncture into K3EDTA tubes and subjected to RBC lysis (eBioscience). Cells were pelleted by centrifugation at 350 X G for 10 minutes at 4°C and washed, suspended in PBS, then stained for 10 minutes with Zombie NIR (BioLegend) and purified anti-CD16/32 (93, BioLegend) to label dead cells and block Fc receptors. Next, cells were stained in wash buffer for an additional 20 minutes with the antibodies listed in the Key Resource Table. Stained cells were washed twice and strained through a 30 μm strainer, then subjected to cell sorting using a Beckman Coulter MoFlo Astrios EQ configured with 355 nm, 405 nm, 488 nm, 561 nm, and 642 nm lasers. Each cell population was hierarchically gated using Beckman Coulter Summit software. Ly6CHi monocytes were defined as CD45PosCD11bHiCD115PosCD19NegCD90.2NegLy6GNegLy6CHi. Ly6CHi monocytes were further restricted to single particles by comparing height and area side scatter pulses, and dead cells were excluded by detecting the integration of the live/dead dye (Zombie NIR).
Approximately 50,000 sorted cells were washed once with PBS and once with 10mM Tris pH 7.4, 10 mM NaCl, 3 mM MgCl2, and 0.1% Igepal by centrifugation at 500 X G at 4C. Cells were then suspended in 50 μl reaction buffer comprised of 25 μl Tagment DNA buffer, 2.5 μl Tagment DNA enzyme, and 22.5 μl nuclease free water, using reagents sourced from Illumina Nextera DNA Library Prep Kit (Buenrostro et al., 2013). Transposase reactions were carried out at 37C for 30 minutes and immediately DNA was purified using Zymo ChIP Clean & Concentrate columns. Resulting DNA was PCR amplified for 14 cycles using barcoding primers and resulting libraries were size selected by gel excision to 175-225 bp as described in (Link et al., 2018). Library DNA was purified, and single end sequenced using a HiSeq 4000 or a NextSeq 500.


title: ATAC_KN-RM_rep1

description: processed data file: KN-RM_ATAC_IDR.txt

value: F4/80-Hi Tim4-Neg Kupffer cells


title: hA3A(W104A)-dCas12a-BE-op:DYR rep c; S4, CFT, CDK rep b; HSD BEACON1 mouse tail#19

value: hA3A(W104A)-dCas12a-BE-op and BEACON1 and hA3A(W104A)-dCas12a-BE-op


title: hA3A(W104A)-dCas12a-BE-op:CFT rep a;S6, RUN, DYR rep b; HSD BEACON1 mouse tail#7


title: HSD BEACON1 mouse tail#5; hA3A-dCas12a-BE:S6 rep g

value: BEACON1 and hA3A-dCas12a-BE


title: ATAC_KN-RM_rep2

description: processed data file: KN-RM_ATAC_IDR.txt


title: Sample 14: CD105_AIA

city: Zagreb

first: Nina

last: Lukač

address: Šalata 11

department: Department of Anatomy

country: Croatia

institute: University of Zagreb School of Medicine

data_processing:
Reads were trimmed with cutadapt (Martin, EMBnet Journal, 2011) to remove Illumina and SmartSeq adapter sequences (CTGTCTCTTATACACATCT and AAGCAGTGGTATCAACGCAGAGT).
Sequence alignment was done using HISAT2 (Kim, Nat Methods, 2015) with default parameters.
Transcripts were assembled and quantified using Stringtie (Pertea, Nat Biotecnol, 2015).
Count matrices were normalized with trimmed mean of M values normalization (TMM) in egdeR (Robinson, Bioinformatics, 2010).
Genome_build: GRCm38 - mm10
Supplementary_files_format_and_content: "gene_count_matrix84.csv" is a comma separated file which contains raw count values for each gene (output from StringTie). "gene_count_matrix84 normalized filtered.csv"  is a comma separated file which contains normalized count values (edgeR, TMM normalization) for each gene after genes with sum of counts in all samples lower than 50 and those without ensable annotation and ERCC controls were excluded. This count matrix was used for differential expression and GSEA analysis. Gene IDs in both files are from Endembl.

source_name: sorted synovial CD31-TER119-CD45-CD51+CD200-CD105+ cells from mice with arthritis

growth_protocol: Antigen-induced arthritis (AIA) was induced by immunization of mice with methylated bovine serum albumin (mBSA) in complete Freund’s adjuvant (CFA) and subsequent intra-articular injection of mBSA. Nonimmunized control mice (NI) were injected with phosphate buffered saline (PBS) at all timepoints. Ten days after intra-articular injection knee joints were harvested.

extract_protocol:
Using Smartseq v4 Ultra® Low Input RNA Kit for Sequencing (TakaRa), RNA was reversely transcribed to first strand cDNA which was amplified. ERCC RNA Spike-In Mix (Invitrogen) was added to lysed cell samples, so that final ERCC dilution in RT reaction was 1:65.000 and final dilution in PCR mix was 1:100.000. Amplicons were purified using Agencourt AMPure XP Kit (Beckman Coulter). Quality of cDNA amplicons was assessed using High sensitivity cDNA chip (Agilent) and Bioanalyzer instrument (Agilent).
1 ng of each cDNA amplicon was used for library preparation using Nextera XT DNA Library Preparation Kit (Illumina).

treatment_protocol: Synovial cells were released from joints by collagenase type IV and labeled with anti-mouse CD90.2-FITC, CD200-PE, CD105-PECy7, CD31-APC, CD45-APC, TER119-APC, CD51-biotin, streptavidin-APCeF780 and DAPI. 4 samples of TER119–CD31–CD45-CD51+CD200+CD105- cells from nonimmunized mice, 5 samples of TER119–CD31–CD45-CD51+CD200+CD105- cells from mice with arthritis, and 5 samples of TER119–CD31–CD45-CD51+CD200-CD105+ from mice with arthritis were sorted using BD FASCAria IIu. For each sample, 200-500 live (DAPI-) cells of described phenotypes were sorted directly into cell lysis buffer (Smartseq v4 Ultra® Low Input RNA Kit for Sequencing, TakaRa) according to the manufacturer’s instructions.

value: 16 weeks

tag: disease

value: arthritits


title: hA3A(W104A/Y130F/P134Y)-dCas12a-BE-op:CDK rep a; HSD BEACON1 mouse tail#1


title: HSD rA1-dCas12a-BE mouse tail#6; S4, hA3A(W104A/P134Y)-dCas12a-BE-op:RUN, DYR, CFT rep a


title: hA3A(W104A/Y132D/P134Y)-dCas12a-BE-op:DYR, CDK rep a; HSD BEACON1 mouse tail#2


title: hA3A(W98Y/W104A/Y130F/Y132D)-dCas12a-BE-op:CDK rep a; HSD BEACON1 mouse tail#3


title: HSD rA1-dCas12a-BE mouse tail#3; hA3A(W104A)-dCas12a-BE-op:S6, S4, RUN, DYR, CDK rep a


title: Sample 12: CD105_AIA

value: sorted synovial CD31-TER119-CD45-CD51+CD200-CD105+ cells

tag: strain


title: Sample 13: CD105_AIA

tag: tissue


title: Sample 4: CD200_NI

source_name: sorted synovial CD31-TER119-CD45-CD51+CD200+CD105- cells from nonimunized mice

value: nonimmunized


title: KC-H_ATF3_ChIP_rep2

value: Control diet


title: Sample 3: CD200_NI

value: sorted synovial CD31-TER119-CD45-CD51+CD200+CD105- cells


title: Sample 9: CD200_AIA

source_name: sorted synovial CD31-TER119-CD45-CD51+CD200+CD105- cells from mice with arthritis


title: Sample 6: CD200_AIA


title: Sample 10: CD105_AIA


title: Sample 8: CD200_AIA


title: Sample 7: CD200_AIA


title: Sample 2: CD200_NI


title: Sample 11: CD105_AIA


title: KC-N_LXR_ChIP_rep1

description:
LXR_ChIP
processed data file: KC-N_LXR_IDR.txt


title: KC-N_LXR_ChIP_rep2


title: Sample 1: CD200_NI


title: HSD rA1-dCas12a-BE mouse tail#8; hA3A(W98Y/W104A/Y132D)-dCas12a-BE-op:S6, CDK  rep a


title: KC-N_InputForLXR-P300_ChIP_rep1


title: HSD rA1-dCas12a-BE mouse embryo#11

source_name: Mouse embryos

value: mouse embryos


title: KC-N_P300_ChIP_rep2

description:
P300_ChIP
processed data file: KC-N_P300_IDR.txt


title: HSD rA1-dCas12a-BE mouse tail#5; hA3A(W98Y/W104A)-dCas12a-BE-op:S6, S4, RUN rep a


title: Sample 5: CD200_AIA


title: HSD BEACON1 mouse tail#8

source_name: Mouse tail


title: KC-N_P300_ChIP_rep1


title: KC-H_inputForLXR_ChIP_rep1


title: KC-H_inputForLXR_ChIP_rep2


title: HSD BEACON1 mouse tail#20


title: HSD rA1-dCas12a-BE mouse embryo#10


title: HSD rA1-dCas12a-BE mouse tail#7; BEACON2:S6, DYR, CDK rep a

value: rA1-dCas12a-BE and BEACON2


title: KC-H_LXR_ChIP_rep1

description:
LXR_ChIP
processed data file: KC-H_LXR_IDR.txt


title: HSD rA1-dCas12a-BE mouse tail#4

value: mouse tail


title: HSD BEACON1 mouse tail#18


title: KC-H_P300_ChIP_rep2

description:
P300_ChIP
processed data file: KC-H_P300_IDR.txt


title: HSD BEACON1 mouse tail#17


title: KC-H_InputForLXR-P300_ChIP_rep2


title: HSD BEACON1 mouse tail#4


title: KC-H_InputForLXR-P300_ChIP_rep1


title: KC-H_P300_ChIP_rep1


title: KC-H_LXR_ChIP_rep2


title: HSD rA1-dCas12a-BE mouse tail#1


title: KC-N_InputForLXR-P300_ChIP_rep2


title: KC-H_H3K27ac_ChIP_rep1

value: H3K27ac (Active Motif 39133, RRID:AB_2561016)


title: HSD BEACON1 mouse tail#16-tail


title: KC-H_H3K27ac_ChIP_rep2


title: HSD BEACON1 mouse tail#6


title: KPC248_mouse tumor

city: Grand Rapids

first: Ian

last: Beddows

state: MI

address: 333 Bostwick Ave NE

institute: Van Andel Institute

data_processing:
Base calling was done by Illumina NextSeq Control Software (NCS) v2.0 and output of NCS was demultiplexed and converted to FastQ format with Illumina Bcl2fastq v1.9.0.
Trim adapters with trim_galore
Align reads with STAR to mm10 genome with option --quantMode GeneCounts
Genome_build: mm10
Supplementary_files_format_and_content: Tab-delimited files with raw counts grom STAR --quantMode GeneCounts

source_name: Bl6 mouse tumor

growth_protocol: For cell_line samples:  whole tumor was dissected from symptomatic animals. About 1 gr of non-necrotic tumor tissue was used as starting material for single cell dissociation. Briefly, dissociated cells were cultured in growth medium: a 1:1 ratio of RPMI 1640 + L-glutamine (Gibco 11875) and DMEM (Gibco 11995) media supplemented with 10% Fetal Bovine Serum (FBS, Sigma F0926-500ML) and 100 units/ml Penicillin and 100 ?g/ml Streptomycin (Life Technologies 15140-122) at 37°C and 5% CO2 in a humidified environment. Cells were passaged at least 8 times to outgrow fibroblast contamination and select only cancer cells, which was confirmed by genotyping of recombined K-Ras and p53 alleles (which included loss of heterozygosity). All experiments were carried with established cell lines between passage 10 and 14. Cell lines were cultured as described above, but with 5% FBS in 10-cm plates each and collected when 60-80% confluent

extract_protocol:
RNA was collected from cell lines or snap-frozen tumors using TRIzol reagent according to the manufacturer’s instructions
Libraries were prepared by the Van Andel Genomics Core from 500g of total RNA using the KAPA Stranded mRNA-Seq Kit (v4.16) (Kapa Biosystems, Wilmington, MA USA). RNA was sheared to 300-400 nts. Prior to PCR amplification, cDNA fragments were ligated to Bio Scientific NEXTflex Adapters (Bio Scientific, Austin, TX, USA). Quality and quantity of the finished libraries were assessed using a combination of Agilent DNA High Sensitivity chip (Agilent Technologies, Inc.), QuantiFluor® dsDNA System (Promega Corp., Madison, WI, USA), and Kapa Illumina Library Quantification qPCR assays (Kapa Biosystems). Individually indexed libraries were pooled and 75 bp, single-end sequencing was performed on an Illumina NextSeq 500 sequencer using a 75 bp HO sequencing kit (v2) (Illumina Inc., San Diego, CA, USA) to a minimum read depth of 30M reads per library.
Quality and quantity of the finished libraries were assessed using a combination of Agilent DNA High Sensitivity chip (Agilent Technologies, Inc.), QuantiFluor® dsDNA System (Promega Corp., Madison, WI, USA), and Kapa Illumina Library Quantification qPCR assays (Kapa Biosystems). Individually indexed libraries were pooled and 75 bp, single-end sequencing was performed on an Illumina NextSeq 500 sequencer using a 75 bp HO sequencing kit (v2) (Illumina Inc., San Diego, CA, USA) to a minimum read depth of 30M reads per library.

treatment_protocol: Neither animals nor cell line samples were subjected to treatment. The main comparison is differential expression between genotype of the samples.

tag: genotype

value: WT


title: ATAC_Ly6CLo-RM_rep1

description: processed data file: Ly6CLo-RM_ATAC_IDR.txt

value: Sorted hepatic macrophages

tag: cell subtype

value: Cd11b-Hi Ly6CLo recruited liver macrophages


title: ATAC_KC-N_rep2

description: processed data file: KC-N_ATAC_IDR.txt

value: F4/80-Hi Tim4-Pos Kupffer cells

value: NASH diet


title: ATAC_Ly6CHi-RM_rep1

description: processed data file: Ly6CHi-RM_ATAC_IDR.txt

value: Cd11b-Hi Ly6CHi recruited liver macrophages


title: KPC202_mouse tumor


title: KPC2194_mouse tumor

value: miR21 KO


title: KPC190_mouse tumor


title: HSD rA1-dCas12a-BE mouse tail#2


title: KPC2190_mouse tumor


title: KPC2192_mouse tumor


title: Adar1 (flox/flox) // Nkx2.5-Cre (Cre/WT) E18.5 embryo #54

city: Copenhagen SV

first: Shizuka

last: Uchida

email: heart.lncrna@gmail.com, suc@dcm.aau.dk

address: Frederikskaj 10B, 2. (building C)

department: Department of Clinical Medicine

country: Denmark

institute: Aalborg University

data_processing:
Fastq files were trimmed using fastp version 0.20.0 and the following prameters: --cut_by_quality5 --cut_by_quality3 --detect_adapter_for_pe  --overrepresentation_analysis --correction  --trim_front1 7 --trim_front2 7
Alignment was performed using STAR version 020201 and the following parameters: --runMode alignReads --runThreadN 8  --outSAMstrandField intronMotif --outSAMmode Full --outSAMattributes All {params.stranded} --outSAMattrRGline {params.readgroup} --outSAMtype BAM SortedByCoordinate --limitBAMsortRAM 45000000000 --quantMode GeneCounts --outReadsUnmapped Fastx  -outSAMunmapped Within {params.keepPairs}
Differential expression analysis was perfomred using edgeR version 3.10 . The RLE method was used to normalize the data.
Genome_build: GRCm38.90
Supplementary_files_format_and_content: tab-delimited text file including CPM values and statistics based outputted by DESeq.

source_name: Heart

extract_protocol:
Trizol extraction of total RNA was performed according to the manufacturer's instruction.
mRNA RNA-seq was performed by Novogene (California, U.S.A.) using NEBNext Ultra II Directional with PolyA Selection kit and sequenced with Illumina HiSeq 4000 (pair-end, 2 × 150 bp).

value: C57BL/6 background

value: Embryo (E18.5)

value: Heart

tag: genotype/variation

value: Adar1 wildtype (KO) embryo


title: KPC2188_mouse tumor


title: ATAC_Ly6CLo-RM_rep2


title: KPC2157_mouse tumor


title: ATAC_Ly6CHi-RM_rep2


title: KPC2183_mouse tumor


title: KPC21167_mouse tumor


title: Adar1 (flox/flox) // Nkx2.5-Cre (WT/WT) E18.5 embryo #47

value: Adar1 wildtype (WT) embryo


title: Adar1 (flox/flox) // Nkx2.5-Cre (WT/WT) E18.5 embryo #50


title: KPC21171_mouse tumor


title: KPC204_pancreatic cell line

source_name: pancreatic cell line


title: KPC232_mouse tumor


title: KPC247_pancreatic cell line


title: ATAC_KC-H_rep2

description: processed data file: KC-H_ATAC_IDR.txt

value: Control diet


title: ATAC_Blood-Ly6CHi-NASH_rep1

description: processed data file: Blood-Ly6CHi-NASH_ATAC_IDR.txt

source_name: Sorted peripheral monocytes

value: Sorted peripheral monocytes

value: Ly6C-Hi blood monocytes


title: ATAC_KC-H_rep1

description: processed data file: KC-H_ATAC_IDR.txt


title: ATAC_Blood-Ly6CHi-NASH_rep2


title: Adar1 (flox/flox) // Nkx2.5-Cre (Cre/WT) E18.5 embryo #51


title: KPC202_pancreatic cell line


title: KPC2156_pancreatic cell line


title: KPC21171_pancreatic cell line


title: KPC2157_pancreatic cell line


title: KPC2154_pancreatic cell line


title: KPC2153_pancreatic cell line


title: KPC232_pancreatic cell line


title: hA3A(W104A/Y132D/P134Y)-dCas12a-BE-op:S6, S4, RUN, CFT, CDK rep b

source_name: HEK293T cells


title: hA3A(W98Y/W104A/Y130F/Y132D)-dCas12a-BE-op:CDK rep c; S6, S4, RUN, DYR, CFT rep b


title: NT: SPA, PMS, PDE, PAF, CHD, BMR, BLM rep a

value: NT


title: hFAST SHAPE-MaP DMSO

description: hFAST SHAPE-MaP DMSO sample

data_processing:
Basecalling using Illumina bcl2fastq v2.17.1.14 software.
High-throughput sequencing reads were separated according to 6-nt experimental barcodes.
SHAPE reactivity profiles were created from deep sequencing datasets for hFAST by using ShapeMapper (version 1.2) with default parameters (Smola et al., 2015). Briefly, low quality score sequence reads were trimmed. Paired-end reads without overlapping sequences were analyzed as two single reads, while paired-end reads with overlapped sequences were merged into single reads. Pre-processed reads were then mapped to target sequences by Bowtie2 (version 2.1.0, parameters: -p 24 -L 15 -D 20 -R 3 --local --sensitive-local --ma 2 --mp 6,2 --rdg 5,1 --rfg 5,1 --dpad 100 --maxins 800 --ignore-quals --no-unal). Mutation rate (MutR) of each position was calculated by dividing the total number of mismatches and indels (deletions and insertions) by the number of all mapped reads to the position. SHAPE reactivities of all positions were calculated by [(ModifiedMutR – UntreatedMutR)/DenaturedMutR] and normalized by a model-free box-plot approach (Deigan et al., 2009). More than 80% positions with at least 1000-read depth in three (modified, untreated and denatured) samples (Mustoe et al., 2018) were used for the hFAST secondary structure modeling.
Genome_build: hg19
Supplementary_files_format_and_content: TXT files include the read depths, standard errors and SHAPE reactivity of each base.

source_name: H9 cells

growth_protocol: H9 cells were cultured on 10 cm dishes for two days to reach 1e6 cells

extract_protocol:
Isolated RNAs were treated with DNase I (Ambion, DNA-freeTM kit) to remove possible DNA contamination. About 50-100 ng of RNAs were obtained under each treatment, and were then used for SHAPE-MaP reverse transcription by adding 1 μL (200 U/μL) of SuperScript II (Invitrogen), 6 mM Mn2+ and gene-specific primers for hFAST. Mn2+ was removed using G-25 micro-spin columns (GE Healthcare) after SHAPE-MaP reverse transcription. Second-strand synthesis was performed with Q5 hot start high-fidelity DNA polymerase and nested PCR was performed to further improve DNA yield. The resulting PCR products were further isolated with PureLink micr spin columns (Life Technologies).
SHAPE-MaP libraries were prepared from 1 ng of DNAs, and size-selected with AmpureXP beads (Agencourt) with a 1:1 (bead to sample) ratio to obtain library DNA products spanning 100-400 bp in length. Final libraries were quantified using Agilent Bioanalyzer 2100 and QuBit high-sensitivity dsDNA assay. Deep sequencing was performed by Illumina NextSeq 500 at CAS-MPG Partner Institute for Computational Biology Omics Core, Shanghai, China.
SHAPE-MaP

treatment_protocol: After washed with DPBS, H9 cells were incubated with 900 μL of cell culture medium and 100 μL of 10x SHAPE Chemical in DMSO with the final concentration of NAI (EMD Millipore) at 100 mM for 10 minutes at 37°C. After removed probing medium, RNAs were isolated by 1 mL Trizol reagent (Invitrogen) according to the manufacturer’s protocol. The same procedure was also performed in parallel for the untreated control samples, but with the addition of only DMSO. In the denaturing control (DC) reaction, RNA was suspended in a denaturing buffer containing 50% formamide and was incubated at 95°C before modification with SHAPE reagents.

value: H9 cells

tag: type

value: cDNA

value: DMSO samples only treated with DMSO


title: hFAST SHAPE-MaP DC

description: hFAST SHAPE-MaP DC sample

value: DC samples denatured by formamide at 95℃


title: hFAST SHAPE-MaP NAI

description: hFAST SHAPE-MaP NAI sample

value: NAI samples modified by NAI in DMSO.


title: NSC_20_ATAC_Seq

city: Evanston

first: Jubao

last: Duan

state: IL

address: 1001 University Place

department: Center for Psychiatric Genetics

institute: NorthShore University HealthSystem

data_processing:
ATAC-Seq data: All raw sequence reads generated by Illumina HiSeq 2000 have been de-complexed at the University of Minnesota Genomic Center and provided as 2x50 bp pair-end fastq files (100 M reads per sample). Adapter remnants, low-quality reads, and low QSEQ short sequences near either end of reads were processed by Trimmomatic (ILLUMINACLIP:NexteraPE-PE.fa:2:30:7, SLIDINGWINDOW:3:18, MINLENGTH:26). The survived sequences were separated into pair-end and single-end fastq files per sample, respectively. The pair-end and single-end fastq files were individually mapped against the human genome reference file including decoy sequences (GRCh38p7/hg38, 1000 Genome Project) using bowtie2 (-x 2000, -mm --qc-filter –met 1 –sensitive –no-mixed -t) and subsequently merged and sorted as BAM-formatted files using samtools, only uniquely mapped reads (MAPQ > 30) were collected. Picard tools MarkDuplicate was then used to remove all PCR and optical duplicated reads from the BAM file.
RNA-Seq data: RNA sequencing files were provided in the format of 2x150 bp pair-end (25 M reads per sample) fastq files. Briefly, libraries were prepared using the NEB Nextera kit with customised adapters. Since the sequencing facility had performed pre-cleaning on raw reads, no Trimmomatic action was performed. The files were subsequently mapped to the human hg38 genome (GRCh38p7) using STAR v 2.6.0 (--outSAMtype BAM SortedByCoordinate --quantMode GeneCounts --outSAMattrIHstart 0 --outSAMstrandField intronMotif --outSAMmultNmax 1 --outFilterIntronMotifs RemoveNoncanonical --outBAMcompression 10 --outBAMsortingThreadN 20 --outBAMsortingBinsN 20 --outFilterMultimapNmax 1 --outFilterMismatchNmax 1 --outSJfilterReads Unique --limitBAMsortRAM 10000000000 --alignSoftClipAtReferenceEnds No --quantTranscriptomeBAMcompression 10 10 ).
Genome_build: GRCh38.p7

source_name: Neural Progenitor Cell

extract_protocol:
For ATAC-seq, we harvested 50,000 cells by centrifugation at 500 g x 5 min at 4C. We washed cells once with PBS buffer and resuspended the cell pellets in 50 uL of cold lysis buffer (10 mM Tris-HCl, pH7.4, 10 mM NaCl, 3 mM MgCl2, 0.1% IGEPAL CA- 630). We collected cell pellets, discarded the supernatants, and immediately continued to transposition reaction in 50 uL transposition reaction mix. We incubated the transposition reaction at 37C for 30 min, immediately followed by purification through a QIAGEN MinElute Kit. We stored the purified DNAs at  20 C until generating sequencing library.
The library was generated and sequenced at University of Minnesota Genomic Center.We have followed the protocol as previously described (Buenrostro et al., 2013) with minor modifications. Specifically, we thawed the transposed and purified DNAs and carried out PCR in 50 uL of reaction with NEBNext High Fidelity 2X Master mix and 5 ul of 25 mM Forward/Reverse ATAC-seq index, i.e., standard barcoded primers of Nextera kit (FC-121-1030), for each sample. After reaction, we purified samples with SPRI beads and eluted the PCR products in 12 uL elution buffer. We checked the DNA library by picogreen quantification and the size of the fragments with an Agilent High Sensitivity DNA chip. Libraries were diluted to 2 nM and pooled for downstream sequencing.
The library was generated and sequenced at University of Minnesota Genomic Center.We have followed the protocol as previously described (Buenrostro et al., 2013) with minor modifications: (1) using the standard Nextera primer set for PCR amplification of the transposed DNA fragments, (2) monitoring qPCR reaction to determine the number of additional cycles for full PCR amplification, and (3) using Agilent High Sensitivity DNA chip for library size validation. Specifically, we thawed the transposed and purified DNAs and carried out an initial 5-cycle PCR (1 cycle: 72 C for 5 min; 1 cycle: 98 C for 30 s; then 5 cycles: 98 C for 10 s, 63 C for 30 s, 72 C for 1 min) in a 50 mL of reaction including NEBNext High Fidelity 2X Master mix and 5 mL of 25 mMForward/Reverse ATAC-seq index, i.e., standard barcoded primers of Nextera kit (FC-121-1030), for each sample.We then used qPCR to determine the number of additional cycles of PCR amplification that were required, by using the cycle number that corresponds to 1/3 of the maximum fluorescent intensity (typically +5, +6, +7) shown on real-time PCR plots on ABI7900 (Applied Biosystems). After the reactions were subjected to additional PCR cycles, we purified each sample with SPRI beads and eluted the PCR products in 12 mL elution buffer. We checked the DNA library by picogreen quantification and the size of the fragments with an Agilent High Sensitivity DNA chip (the library should be sized between 200-700 bp). Libraries were diluted to 2 nM and pooled for downstream sequencing. For the initial OCR profiling, we pooled all the 6 libraries (2 for iPSC, 1 for N-d27, 1 for N-d33, and 2 for N-d41) and sequenced on a HiSeq 2500 using a PE 2x50 bp flow cell in a single sequencing lane. For the additional ATAC-seq on the unedited and edited iNs, we followed the same procedures as described above. We achieved > 120 million passing-filter reads for the lane and the average quality scores for all libraries are all above Q30 for both R1 and R2.

treatment_protocol: NPC are prepared using PSC Neural Induction Medium (Thermofisher) following vendor’s protocol with modification. Briefly, hiPSCs were replated as clumps in mTeSR media (STEMCELL) supplemented with 5 µM ROCK inhibitor (R&D Systems) on Day 0. From Day 1 to Day 9, cells were treated in PSC Neural Induction Medium with routinely removal of differentiated non-neuron cells. PSC Neural Induction Medium was prepared as 490ml Neurobasal Medium (Thermofisher) and 10 ml Neural Induction Supplement (Thermofisher). On Day 10, P0 NPCs were ready for passaging. From P0 to P3, NPCs were passaged using Rosette Selection Reagent (STEMCELL) and expanded in Neural Expansion Medium. Neural Expansion Medium was prepared as 245 ml Neurobasal Medium, 245 ml Advanced DMEM⁄F-12 media (Thermofisher) and 10ml Neural Induction Supplement. From P4, NPCs were passaged as single cells using Accutase (Thermofisher). 5 µM ROCK inhibitor were supplemented into media on the day of passage and withdrawn 24 hr post replating.

value: iPS-derived Neural Progenitor Cell


title: NT: SPA, PMS, PDE, PAF, CHD, BMR, BLM rep b


title: NSC_19_ATAC_Seq


title: NSC_21_ATAC_Seq


title: NT: SPA, PMS, PDE, PAF, CHD, BMR, BLM rep c


title: hA3A(W104A/Y130F/P134Y)-dCas12a-BE-op:CDK  rep c; S6, S4, RUN, DYR, CFT rep b


title: KPC190_pancreatic cell line

value: WT


title: NSC_18_ATAC_Seq


title: NSC_17_ATAC_Seq


title: NSC_16_ATAC_Seq


title: NSC_10_ATAC_Seq


title: NSC_13_ATAC_Seq


title: NSC_12_ATAC_Seq


title: hA3A(W104A)-dCas12a-BE-op: S6, S4, RUN, CFT, CDK rep c


title: NSC_14_ATAC_Seq


title: NSC_09_ATAC_Seq


title: NSC_07_ATAC_Seq


title: hA3A-dCas12a-BE:CDK rep g; PD1 rep d; S6, DYR rep h

value: hA3A-dCas12a-BE and hA3A-dCas12a-BE and hA3A-dCas12a-BE


title: NSC_08_ATAC_Seq


title: NSC_06_ATAC_Seq


title: NSC_05_ATAC_Seq


title: NSC_15_ATAC_Seq


title: NSC_11_ATAC_Seq