title: MKO5

description:
SMART-Seq2
S11


title: Hs_11w4d_MS321_205

status: Public on Aug 01 2022

type: SRA

city: Kyoto

first: Yukihiro

last: Yabuta

state: Kyoto

address: Yoshida-Konoe-cho, Sakyo-ku

department: Anatomy and Cell Biology

country: Japan

institute: Kyoto University, Graduate school of medicine

description: cDNAs amiplified from 1 ng total RNA extracted from cells using SC3-seq technology

library_source: transcriptomic

data_processing:
The processing of the 3-prime-sequencing data for calculating the gene-expression levels was performed as described previously (Nakamura et al., 2015).  In brief, all reads were processed with cutadapt v1.9.1 to remove adaptors and poly-A sequences.
The reads of 30-bp or longer were mapped onto a mouse transcript reference, ref_GRCh38.p12, using Tophat v2.0.11/Bowtie2 v2.2.7.  Read counts per gene were calculated using HTSeq v0.9.1 with the default settings.  Read counts were converted to read counts per million mapped reads (RPM) and used for the analysis.
Genome_build: GRCh38.p12
Supplementary_files_format_and_content: Tab-delimited text files include RPM values for each sample.

molecule: polyA RNA

organism: Homo sapiens

source_name: An oocyte from 11w4d fetal ovary

extract_protocol:
Fetal ovaries collected from human fetus were carefully dissected out in chilled PBS.  Fetal ovaries at 8-14 wpf were minced with scissors in 0.05% trypsin/EDTA/PBS solution (GIBCO, 15400-054), then incubated for 15 min at 37℃ with gentle pipetting every 5 min.  Dissociation medium [DMEM (GIBCO, 10313021) containing 10% FBS, 2mM L-glutamine (GIBCO, 25030081), 1X penicillin/streptomycin (GIBCO, 15070063), and 100μg/mL DNaseI (MERCK, DN25)] was added for further dissociation.  For SC3-seq analysis, cells were picked under the inverted microscope (OLYMPUS, IX71) to exclude multiplet cells.   For dissociating human rOvaries, tissues were incubated with TryPLE Express (1X) (GIBCO, 12604021) for 5 min.  After adding dissociation medium, cells were processed for single-cell analyses. Dissociated germ cells were manually picked using glass capillaries under the stereo microscope (Leica, S8 APO).  Following cDNA synthesis and amplification methods were previously described (Nakamura et al., 2015).
The cDNA synthesis was performed according to the single-cell 3 prime RNA-seq (SC3-seq) method described previously (Nakamura et al., 2015).  The cDNA libraries for Nextseq500 (Illumina) were constructed as described previously (Ishikura et al., 2016).The sequencing was performed on an Illumina Nextseq500/550 platform with a Nextseq500/550 Hiqh output kit (Illumina).

tag: developmental stage

tag: in vitro culture period

tag: cell type

value: Germ cell


title: Hs_11w4d_MS285_211


title: Old dFOXO null_Biol rep 3

city: Waltham

first: Michael

middle: Thomas

last: Marr

state: Massachusetts

address: 415 South street

department: Biology

country: USA

institute: Brandeis University

data_processing:
RNA-seq fastq files were uploaded to the public server usegalaxy.org and processed at the Galaxy web platform.
The tools FASTQ Groomer and FastQC were used for library quality control. Parameters: Default.
To obtain gene counts, the RNA STAR aligner was used to map the sequencing data to both the D. melanogaster genome (dm6) and to a TE consensus FASTA file with 176 Drosophila TE. The per gene/transcript count output option was activated. Default parameters were used.
Assembly: Drosophila melanogaster genome (dm6) and Drosophila transposon consensus sequence FASTA file.
Supplementary files format and content: csv file contains matrix with raw counts for each sample.

molecule: total RNA

organism: Drosophila melanogaster

source_name: Whole fly

growth_protocol: Flies were kept at low density in mixed sex vials. Fly food was changed twice a week. Recipe per 100mL: Agar (0.75g), Dextrose (7.87g), Flaked yeast (2.44g), Cornmeal (5.71g), Methyl-4 (0.21g), Ethanol (1.11mL), Sodium Potassium (0.6g), Calcium Chloride (0.08g).

extract_protocol:
Total RNA from the whole body of 10 male wDAH and dFOXO deletion (wDAH ∆94) flies was extracted with TRI Reagent at 5 to 6 days and 30 to 31 days old according to the manufacturers protocol (Molecular Research Center, Inc., Cincinnati, OH).
To generate RNA-seq libraries, 1 μg of total RNA was used as input for the TruSeq RNA Library Prep Kit v2 (Illumina, Inc., San Diego, CA) and the manufacturer’s protocol was followed. Strandedness: Unstranded.

tag: Sex

value: Male

tag: tissue

value: Whole fly

tag: age

value: 31 days

tag: genotype

value: wDAH [delta]94

tag: condition

value: dFOXO deletion


title: MKO3

city: Boston

first: Yangpo

last: Cao

address: 320 Longwood Ave

institute: Boston Children's hospital

description:
SMART-Seq2
S9

data_processing:
alignment by STAR to mm10
generate reads count matrix with FeatureCount
Differential expression analysis by DESeq2

organism: Mus musculus

source_name: heart

extract_protocol:
PureLink RNA Micro Scale Kit
SMART Seq v4 kit, Nextera XT library construction kit
Dual indexed library, enzymatic

treatment_protocol: AAV treatment of mice, Langendorff isolation of cardiomyocytes, FACS purification of cardiomyocytes

value: heart

value: cardiomyocyte

tag: treatment

value: AAV-Myocd-sgRNA

value: 4 week old


title: Hs_11w4d_MS321_207


title: Old dFOXO null_Biol rep 1

value: 30 days


title: T47D_LTED_shcontrol_input_ChIPseq_rep2

city: Miami

first: Liliana

last: Garcia-Martinez

state: Florida

address: 1501 NW 10th Avenue

department: Human Genetics

institute: University of Miami

library_source: genomic

data_processing:
ATAC-seq: Paired-end ATAC-seq fastq files were processed using the ATAC-seq Pipeline from the Kundaje Lab (https://github.com/kundajelab/atac_dnase_pipelines). Reads were trimmed using cutadapt v1.9.1 using the following parameters: -m 5. Reads were aligned to the hg19 genome using Bowtie2 v2.2.6, and the SAMtools v1.7 was used to convert the output files into the BAM format using default parameters. Duplicate reads were removed using Picard tool v1.126 using default parameters. Shuf was used to subsample reads to 50 million. Peaks were called using MACS2 v2.1.0 using default parameters
Assembly: hg19
Supplementary files format and content: The processed data files are in bigWig format containing the indexed tag chromosome coordinates and score produced from MACS2 peak calling.

molecule: genomic DNA

growth_protocol: T47D were maintained at 37°C with 5% CO2 according to ATCC recommendations. Cells were culture in  estrogen deprived conditions for 9 months. Cell culture media: IMEM with no phenol red (Corning 10-026-CV supplemented with 10% charcoal stripped FBS(Gemini 100-119) and Penicillyn/Streptomycin (Thermo Fisher Scientific 15140-12, 10000U/mL.

extract_protocol:
Cells were grown to 80% confluence on 150cm plates and fixed with 11% solution containing formaldehyde (Thermo Fisher #28908), 50mM HEPES-KOH(pH 7.5), 100mM Nacl, 0.5M EDTA, 0.5M EGTA for 10 min. 1/20 of the plates volume equivalent of freshly prepared Glycine 2.5M(stock) was added to quench the reaction for 5min . The supernatant was aspirated, and the cells were washed twice with 5 ml PBS and harvested on ice using cell scrapers . Cells were pelleted at 2950 x g for 10 min at 4°C, washed 1x with 10 ml cold PBS, and pelleted once more. Cell pellets were flash freeze in liquid nitrogen and store at -80°C until processing day. Cell pellet collected from 6 plates of cells was resuspended in 5 ml of Lysis Buffer 1 (50 mM HEPES-KOH, pH 7.5; 140 mM NaCl, 1mM EDTA, 10% glycerol, 0.5% NP-40, 0.25% Triton X-100), allowed to rotate for 10 min at 4°C, and pelleted at 2950 x g for 10 min at 4°C. Cells were then resuspended in 5 ml of Lysis Buffer 2 (10 mM Tris-HCL pH 8.0, 200 mM NaCl, 1 mM EDTA, 0.5 mM EGTA), allowed to rotate for 10 min at room temperature, and pelleted at 2950 x g for 10 min at 4°C. Cells were then resuspended in 3 ml of Lysis Buffer 3 (10 mM Tris-HCl pH 8, 100 mM NaCl, 1 mM EDTA, 0.5 mM EGTA, 0.1% Na-Deoxycholate, 0.5% N-lauroylsarcosine) and then transfer to 15ml sonication tubes (Diagenode cat# C01020031). Cells were sonicated on a Bioruptor Pico at 4°C for 20min of 30’’ ON-OFF cycles with 1.2 g of sonication beads. After sonication, samples were transferred to 2 separate 1.5 ml microtube and centrifuged at 20,000 x g for 15 minutes at 4°C. The supernatant was transferred to new 15 ml tubes. To check sonication efficiency, 50µl of sonicated samples was transferred to a new microtube with 50µl PBS and incubated at 65°C for 3hr on an Eppendorf Thermomixer shaking at 1000rpm to decrosslink. DNA was purified with the QIAquick PCR Purification Kit (Qiagen cat# 28106) and eluted in 50µl of H20. 1 ug of decrosslinked chromatin from each sample were run in a 1% agarose gel at 100V for 45min and imaged on a Bio-rad ChemiDoc XRS+. DNA concentration in the sonicated decrosslinked samples were measured via qubit, and this concentration was then used to estimate the concentration of chromatin in the un-decrosslinked samples. 5ug of Antibody (see Key Resources table for list of antibodies) along with 1ug of Spike-in antibodies (Active Motif 61686) were bound to 50 ul of Dynabeads Protein G (Thermo Fisher 10004D), previously washed with 1 ml of 0.5% BSA in 1x PBS three times, and rotated end-to-end overnight at 4°C (Eppendorf cat# 0030108051). After binding of the antibodies , beads were washed with 1 ml of 0.5% BSA in 1x PBS three times. 30ug of chromatin and 50 ng of Spike-in chromatin (Active Motif 53083) were added to 1.5ml LoBind tube (Eppendorf cat# 0030108051) containing pre-bound antibodies beads and brought up to 500µl final volume with Lysis Buffer and rotated end-to-end overnight at 4°C.  5 µl was removed as input material (1%) and placed in a separate microtube at 4°C completing the volume to 50uL with Lysis Buffer 3.  Beads were washed 6 times with 1 ml Wash Buffer (50 mM Hepes-KOH, pH 7.6; 500 mM LiCl, 1 mM EDTA, 1% NP-40; 0.5% Na-Deoxycholate) and once with 1 ml TE containing 50 mM NaCl. Beads were dried after the last wash by removing as much TE as possible. 210 ul of Elution Buffer (50 mM Tris-HCl, pH 8; 10 mM EDTA, 1% SDS) was added to each sample and  150 ul elution buffer were added to each input sample that was previously set aside. To elute immunocomplexes from beads, samples were incubated at 65°C for 15 minutes on an Eppendorf Thermomixer shaking at 1000rpm. Tubes were centrifuged for 3min at 960xg at RT and 200µl of supernatant was transferred to a new tube and reverse crosslinked overnight at 65°C. Next day, 200 ul of TE was added to each decrosslinked sample and input, and the samples were treated sequentially with 0.2 mg/ml RNAse A (Sigma R5503) at 37°C and 0.2 ug/ml Proteinase K (NEB P8107S) at 55°C for two hours each. DNA purification was performed with the QIAquick PCR Purification Kit (Qiagen cat# 28106) and eluted in 50µl of H2O and quantified by Qubit.
Immunoprecipitated DNA was used to generate libraries using the NEBNext Ultra DNA Library Prep Kit for Illumina (New England Biolabs cat# 7370) following the manufacturer’s instructions.

value: Mammary gland ductal carcarcinoma epithelial cells derived from metastatic site

tag: cell source

value: ATCC #HTB-133

tag: modifications

value: Parental estrogen deprived for 9 months

tag: chip antibody


title: T47D_LTED_shcjun_cjun_ChIPseq_rep2

value: Parental estrogen deprived for 9 months, plko-shcjun estable SHCLNG-NM_002228 (TRCN0000010366) Sigma Aldrich

value: Anti-c-Jun antibody (ab31419)


title: T47D_LTED_shcontrol_cjun_ChIPseq_rep2


title: Wild type normal

city: Nashville

first: Scott

middle: Mattox

last: Haake

state: TN

address: 2220 Pierce Ave, PRB 606

department: Medicine, Division of Hematology/Oncology

institute: Vanderbilt University Medical Center

description: single cell

data_processing:
RTA (version 2.4.11; Illumina) was used for base calling and analysis was completed using 10X Genomics Cell Ranger software v2.1.1.
After alignment and demultiplexing, samples were jointly analyzed using a standard Seurat (2)/Scanpy (3) pipeline as previously described (4).
Briefly, cells containing fewer than 500 genes, <0.5% or >10% mitochondrial reads were filtered and excluded from downstream analysis.
Libraries were merged and jointly normalized and scaled using SCTransform (4) in Seurat v3.2 including “percent.mt” as a regression variable, followed by principal components analysis using variable genes and graph-based clustering.
Immune (Ptprc+), epithelial (Epcam+) and stromal (Pecam1+ or Col1a1+) cells were independently extracted, followed by recursive clustering, doublet exclusion (clusters containing nonphysiologic marker combinations), and cell-type annotation.
Genome_build: mm10
Supplementary_files_format_and_content: Contains detected cellular barcodes.

source_name: Lung

extract_protocol:
Tumors and normal adjacent tissue of two WT and two integrin b1-KO mice were macrodissected and disassociated using the Miltenyi Biotec gentleMacs dissociator and the mouse tumor dissociation kit (Miltenyi Biotec #130-096-730). 10,000 viable cells were captured for each tissue.
scRNA-seq libraries were prepared using the 10X Chromium Single Cell Platform (10X Genomics #1000006, 1000080, and 1000020) following the manufacturer's protocol.
Other:single cell RNA-seq

tag: tissue type

value: Normal adjacent lung

value: Type 2 alveolar epithelial cell integrin beta1 KO


title: MKO4

description:
SMART-Seq2
S10


title: Hs_11w4d_MS285_214

value: Germ cell


title: Young dFOXO null_Biol rep 3

value: Male

value: Whole fly

value: 6 days


title: MKO6

description:
SMART-Seq2
S12


title: T47D_LTED_LSD1_KO_2x_INPUT_ChIPseq_rep2

extract_protocol:
Cells were grown to 80% confluence on 150cm plates and fixed using dual crosslinking approach. First protein-protein interactions were preserved with ChIP-gold crosslinking reagent (Diagenode C01019027) according to manufacturer's instructions followed by  DNA-protein crosslink with 11% solution containing formaldehyde (Thermo Fisher #28908), 50mM HEPES-KOH(pH 7.5), 100mM Nacl, 0.5M EDTA, 0.5M EGTA for 10 min. 1/20 of the plates volume equivalent of freshly prepared Glycine 2.5M(stock) was added to quench the reaction for 5min . The supernatant was aspirated, and the cells were washed twice with 5 ml PBS and harvested on ice using cell scrapers . Cells were pelleted at 2950 x g for 10 min at 4°C, washed 1x with 10 ml cold PBS, and pelleted once more. Cell pellets were flash freeze in liquid nitrogen and store at -80°C until processing day. Cell pellet collected from 6 plates of cells was resuspended in 5 ml of Lysis Buffer 1 (50 mM HEPES-KOH, pH 7.5; 140 mM NaCl, 1mM EDTA, 10% glycerol, 0.5% NP-40, 0.25% Triton X-100), allowed to rotate for 10 min at 4°C, and pelleted at 2950 x g for 10 min at 4°C. Cells were then resuspended in 5 ml of Lysis Buffer 2 (10 mM Tris-HCL pH 8.0, 200 mM NaCl, 1 mM EDTA, 0.5 mM EGTA), allowed to rotate for 10 min at room temperature, and pelleted at 2950 x g for 10 min at 4°C. Cells were then resuspended in 3 ml of Lysis Buffer 3 (10 mM Tris-HCl pH 8, 100 mM NaCl, 1 mM EDTA, 0.5 mM EGTA, 0.1% Na-Deoxycholate, 0.5% N-lauroylsarcosine) and then transfer to 15ml sonication tubes (Diagenode cat# C01020031). Cells were sonicated on a Bioruptor Pico at 4°C for 20min of 30’’ ON-OFF cycles with 1.2 g of sonication beads. After sonication, samples were transferred to 2 separate 1.5 ml microtube and centrifuged at 20,000 x g for 15 minutes at 4°C. The supernatant was transferred to new 15 ml tubes. To check sonication efficiency, 50µl of sonicated samples was transferred to a new microtube with 50µl PBS and incubated at 65°C for 3hr on an Eppendorf Thermomixer shaking at 1000rpm to decrosslink. DNA was purified with the QIAquick PCR Purification Kit (Qiagen cat# 28106) and eluted in 50µl of H20. 1 ug of decrosslinked chromatin from each sample were run in a 1% agarose gel at 100V for 45min and imaged on a Bio-rad ChemiDoc XRS+. DNA concentration in the sonicated decrosslinked samples were measured via qubit, and this concentration was then used to estimate the concentration of chromatin in the un-decrosslinked samples.  10ug of Antibody (see Key Resources table for list of antibodies) along with 2ug of Spike-in antibodies (Active Motif 61686) were bound to 50 ul of Dynabeads Protein G (Thermo Fisher 10004D), previously washed with 1 ml of 0.5% BSA in 1x PBS three times, and rotated end-to-end overnight at 4°C (Eppendorf cat# 0030108051). After binding of the antibodies , beads were washed with 1 ml of 0.5% BSA in 1x PBS three times. 30ug of chromatin and 50 ng of Spike-in chromatin (Active Motif 53083) were added to 1.5ml LoBind tube (Eppendorf cat# 0030108051) containing pre-bound antibodies beads and brought up to 500µl final volume with Lysis Buffer and rotated end-to-end overnight at 4°C.  5 µl was removed as input material (1%) and placed in a separate microtube at 4°C completing the volume to 50uL with Lysis Buffer 3.  Beads were washed 6 times with 1 ml Wash Buffer (50 mM Hepes-KOH, pH 7.6; 500 mM LiCl, 1 mM EDTA, 1% NP-40; 0.5% Na-Deoxycholate) and once with 1 ml TE containing 50 mM NaCl. Beads were dried after the last wash by removing as much TE as possible. 210 ul of Elution Buffer (50 mM Tris-HCl, pH 8; 10 mM EDTA, 1% SDS) was added to each sample and  150 ul elution buffer were added to each input sample that was previously set aside. To elute immunocomplexes from beads, samples were incubated at 65°C for 15 minutes on an Eppendorf Thermomixer shaking at 1000rpm. Tubes were centrifuged for 3min at 960xg at RT and 200µl of supernatant was transferred to a new tube and reverse crosslinked overnight at 65°C. Next day, 200 ul of TE was added to each decrosslinked sample and input, and the samples were treated sequentially with 0.2 mg/ml RNAse A (Sigma R5503) at 37°C and 0.2 ug/ml Proteinase K (NEB P8107S) at 55°C for two hours each. DNA purification was performed with the QIAquick PCR Purification Kit (Qiagen cat# 28106) and eluted in 50µl of H2O and quantified by Qubit.
Immunoprecipitated DNA was used to generate libraries using the NEBNext Ultra DNA Library Prep Kit for Illumina (New England Biolabs cat# 7370) following the manufacturer’s instructions.

value: LSD1 KO parental cells generated with CRISPR/Cas9


title: Hs_11w4d_MS277_019


title: Hs_11w4d_MS285_208


title: Hs_11w4d_MS285_209


title: T47D_LTED_LSD1_KO_2x_INPUT_ChIPseq_rep1


title: MKO1

description:
SMART-Seq2
S7

value: heart


title: Ctrl6

description:
SMART-Seq2
S6

value: cardiomyocyte

value: AAV-Cre


title: Young dFOXO null_Biol rep 1

value: wDAH [delta]94

value: dFOXO deletion


title: Hs_11w4d_MS321_204


title: T47D_LTED_shcjun_input_ChIPseq_rep2

value: Mammary gland ductal carcarcinoma epithelial cells derived from metastatic site


title: Old dFOXO null_Biol rep 2

value: 31 days


title: T47D_LTED_LSD1_KO_1x_INPUT_ChIPseq_rep2

value: ATCC #HTB-133


title: Ctrl5

description:
SMART-Seq2
S5

value: 4 week old


title: Hs_11w4d_MS285_210


title: Old wild type control_Biol rep 3

value: 30 days

value: wDAH

value: wildtype control


title: MKO2

description:
SMART-Seq2
S8

value: AAV-Myocd-sgRNA


title: Ctrl4

description:
SMART-Seq2
S4


title: Young wild type control_Biol rep 1

value: Male


title: Old wild type control_Biol rep 1

value: Whole fly


title: Ctrl3

description:
SMART-Seq2
S3


title: Young dFOXO null_Biol rep 2

value: 6 days


title: Ctrl2

description:
SMART-Seq2
S2


title: Hs_11w4d_MS321_203

value: Germ cell


title: Hs_11w4d_MS285_206


title: T47D_LTED21M_Corin_H3K4me2_ChIPseq_rep2

growth_protocol: T47D were maintained at 37°C with 5% CO2 according to ATCC recommendations. Cells were culture in  estrogen deprived conditions for 21 months. Cell culture media: IMEM with no phenol red (Corning 10-026-CV supplemented with 10% charcoal stripped FBS(Gemini 100-119) and Penicillyn/Streptomycin (Thermo Fisher Scientific 15140-12, 10000U/mL. Cells were treated with 1uM Corin (MedChem Express HY-111048) for 48h.

value: Parental estrogen deprived for 21 months treated with Corin 1uM 48h

value: Histone H3K4me2 antibody (pAb) Active Motif 39141


title: Young wild type control_Biol rep 2


title: Old wild type control_Biol rep 2


title: TS/A_Emp_ColIII_rep01

city: Kanazawa

first: Soichiro

last: Sasaki

address: Kakuma-machi

institute: Cancer Research Institute, Kanazawa University

data_processing:
We used TruSeq RNA Library Preparation Kit v2.
Sequencing was performed by NextSeq 500/550, High output 75 bp.
Obtained fastq files were anaylzed by CLC Genomics Wrokbench (20.0.4).
Genome_build: Mus-musclus-2019-04-08-19-53
Supplementary_files_format_and_content: We used total gene reads for analysis which was generated by CLC Genomics Wrokbench (20.0.4)

source_name: Mouse breast cancer cell line

growth_protocol: RPMI supplemented with 10% FCS

extract_protocol:
Total RNAs were extracted by using RNeasy Mini Kit (Qiagen).
We used TruSeq RNA Library Preparation Kit v2.
bulk-RNA-Seq

treatment_protocol: 4T1 or TS/A cells which were transduced Gpr56/Adgrg1 expressing virus vector or empty vector

tag: strain

value: BALB/c

tag: transduced vector

value: Gpr56/Adgrg1 expressing vector


title: T47D_LTED_LSD1_KO_H3K4me2_ChIPseq_rep2

growth_protocol: T47D were maintained at 37°C with 5% CO2 according to ATCC recommendations. Cells were culture in  estrogen deprived conditions for 21 months. Cell culture media: IMEM with no phenol red (Corning 10-026-CV supplemented with 10% charcoal stripped FBS(Gemini 100-119) and Penicillyn/Streptomycin (Thermo Fisher Scientific 15140-12, 10000U/mL.

value: LSD1 KO parental cells generated with CRISPR/Cas9

value: Histone H3K4me2 antibody (pAb)   Active Motif 39141


title: T47D_LTED_LSD1_KO_SMARCC1_ChIPseq_rep2

value: Mammary gland ductal carcarcinoma epithelial cells derived from metastatic site

value: SMARCC1/BAF155 (D7F8S) Rabbit mAb #11956 Cell Signaling Technology


title: Ctrl1

description:
SMART-Seq2
S1

value: heart

value: cardiomyocyte

value: AAV-Cre

value: 4 week old


title: Hs_11w4d_MS277_008


title: TS/A_Emp_Control_rep02

value: Empty vector


title: Young wild type control_Biol rep 3

value: Male

value: 5 days

value: wDAH

value: wildtype control


title: T47D_LTED_LSD1_KO_cjun_ChIPseq_rep2

value: ATCC #HTB-133

value: Anti-c-Jun antibody (ab31419)


title: Hs_11w4d_MS321_202

value: Germ cell


title: Hs_11w4d_MS285_203


title: T47D_LTED21M_Corin_H3K4me2_ChIPseq_rep1

value: Parental estrogen deprived for 21 months treated with Corin 1uM 48h

value: Histone H3K4me2 antibody (pAb) Active Motif 39141


title: Hs_11w4d_MS285_204


title: T47D_LTED_LSD1_KO_SMARCC1_ChIPseq_rep1

value: LSD1 KO parental cells generated with CRISPR/Cas9


title: Hs_11w4d_MS321_201


title: Naive_primed_D30ASECRiAV_10XscATACseq

city: Leuven

first: Vincent

last: Pasque

address: Herestraat 49 bus 804

department: Development and Regeneration

country: Belgium

institute: KU Leuven

description: 10X Genomics scATACseq

data_processing:
Preprocessing of single-cell chromatin accessibility data was performed using the cellranger-atac pipeline (10X Genomics).
Read filtering, alignment, cell and peak calling, as well as cell-by-peak count matrix generation were performed using the “count” option (cellranger-atac count) with default parameters.
Downstream analysis was performed using Signac (Stuart et al. 2021). The count matrix was filtered for cells where at least 15% of all fragments fell within peaks, cells with less than 5% of fragments falling within blacklist regions, cells with less than 4% nucleosome signal, cells with greater than 2% enrichment for transcription start sites, and peaks with a minimum of 2500 fragments and a maximum of 20000 fragments.
Cell clusters were identified using the FindNeighbors function with parameters “reduction = ‘lsi’, dims = 2:30’ and FindClusters function with parameters “algorithm = 3, resolution = 0.5”. Upon initial clustering, cluster-specific peaks were called with MACS2 using the function CallPeaks with parameters “group.by = ‘idents’”. These peaks were used to generate a final SIGNAC object which was used for all downstream analyses.
Fragment counts were mapped to peaks using FeatureMatrix with parameters “process_n = 2000”.
Filtering and clustering were performed on the new object as described above. Peaks were then filtered for mean accessibility, keeping all peaks with greater than 0.1 mean accessibility across cells.
Genome_build: GRCh38
Supplementary_files_format_and_content: Processed data files containing the ATAC-seq peaks for each cell are provided in BED format, and metadata pertaining to the cluster of each cell is provided in a tab-deliminated file. A loom file is also provided.

source_name: Primed and naive hPSCs, hTSCs and EXMCs.

growth_protocol:
growth protocol for human primed hPSCs: Human primed pluripotent stem cells (H9 hESCs and Sigma hiPSCs) were grown with or without feeder conditions as per the cell lines at normoxia conditions (5% CO2) and under humidified conditions at 37° C. In feeder-free conditions cells were cultured in pre-coated geltrex tissue culture treated plates in complete E8Flex medium (Stem cell technology). Cells were dissociated into smaller clumps every 5-6 days using veresene incubating 5 min at room temperature.
growth protocol for human naive hPSCs: Naive hPSCs (H9 hESCs and Sigma hiPSCs) were cultured on mitotically inactivated MEF feeders in 5% O2 and 5% CO2 incubator under humidified conditions at 37°C. All naive hPSCs were cultured in PXGL medium (Bredenkamp/Guo Stem cell reports 2019 (PMID: 31708477) which consists of 1:1 DMEM/F12 and Neurobasal, 0.5% N2-supplement, 1% B27-Supplement, 2 mM L-Glutamine, 0.1 mM β-mercaptoethanol, 1x penicillin-streptomycin, 1 μM PD0325901 (Axon Medchem and Wellcome-MRC Cambridge Stem Cell Institute), 2 μM XAV939 (Sigma), 2 μM Gö6983 (Tocris), 20 ng/mL human LIF (PeproTech and Wellcome-MRC Cambridge Stem Cell Institute) and 10 μM Y-27632 (Tocris and Cell Guidance Systems). Naive hPSCs were passaged every 4-5 days in a ratio 1:2 or 1:3 by single-cell dissociation with Accutase followed by filtering through a 40 μm cell strainer.
growth protocol for human TSCs: Human naive to trophoblast and EXMC conversion were done using the following previously described protocol (Dong et al. 2020; Cinkornpumin et al. 2020). Naive hPSCs were seeded in such a way that expecting at least 90% confluency by Day 2 on mitomycin C treated MEFs after dissociating in single-cell using TryplE (for 15 min at 37°C) in their respective naive culture conditions supplemented with 10μM Y-27632. The very next day the media was switched from naive to trophoblast stem cell medium (Okae et al. 2018) consisting of DMEM/F12 (Gibco) supplemented with 0.3% BSA, 0.2% FBS, 1% Penicillin-streptomycin, 1% insulin-transferrin-selenium-ethanolamine-X 100 supplement, 1.5 μg/ml L-ascorbic acid, 0.5 μM A83-01, 1μM SB431542, 50ng/ml hEGF, 2uM CHIR99021, 0.8 mM Valproic acid, 0.1 mM β-mercapto-EtOH (Gibco) and 5 μM Y-27632. The medium was changed every two days supplemented with 5 μM Y-27632. From passage 1 onwards cultures were cultured and maintained on 5 μg/ml Collagen IV coated tissue culture treated plates in hypoxia conditions (5% O2 and 5% CO2) and passaged every 4 days in 1:3 or 1:6 splitting ratios.

extract_protocol:
Cells were washed with Phosphate-buffered Saline (PBS), dissociated from culture dishes by Accutase (7 min. at 37°C) in hypoxic condition for naive hPSCs and TSCs and in normoxic condition for primed hPSCs, and finally diluted with DMEM. Single cell suspensions were filtered through a 40µm cell strainer, centrifuged at 200 rcf for 5 min., resuspended in PBS with 0.04% Bovine Serum Albumin (BSA), and counted with a Luna-FL automated Fluorescence Cell Counter (Logos Biosystems). 100 000 – 1 000 000 cells were added to a 2-ml microcentrifuge tube and were centrifuged (300 rcf, 5min at 4°C). Supernatant was removed without disrupting the cell pellet and 100µl chilled lysis buffer (10 mM Tris-HCl (pH 7.4), 10 mM NaCl, 3 mM MgCl2, 0.1% Tween-20, 0.1% Nonidet P40 Substitute, 0.01% digitonin and 1% BSA) was added and mixed by pipetting 10 times. The microcentrifuge tube was incubated on ice for 4 min and then 1ml chilled wash buffer (10 mM Tris-HCl (pH 7.4), 10 mM NaCl, 3 mM MgCl2, 0.1% Tween-20 and 1% BSA) was added and mixed by pipetting 5 times. Nuclei were centrifuged (500 rcf, 5 min at 4°C) and the supernatant was removed without disrupting the nuclei pellet. Nuclei were resuspended in a chilled Diluted Nuclei Buffer (10x Genomics; 2000153) at 610-1540 nuclei per µl based on the starting number of cells. The nuclei concentration was determined using a NucleoCounter NC-100 (Chemometec). Nuclei were mixed using the following ratio (H9 primed: Sigma primed: H9 naive: Sigma naive: Sigma TSCs = 1:1:1:1:2) and were immediately used to generate scATAC-seq libraries as described in the methods below.
scATAC-seq libraries were prepared according to the Chromium Single Cell ATAC Reagent Kits User Guide (10x user guide; CG000168, Revision D). Nuclei were loaded onto the 10x Chromium Single Cell Platform (10x Genomics) at a concentration targeting 2000 nuclei, according to the manufacturer's protocol. Nuclei transposition, generation of gel beads in emulsion (GEMs), barcoding, GEM cleanup, and library construction were performed with the Chromium Single Cell ATAC Reagent Kits (v1 Chemistry) according to the manufacturer's protocol. Library Quality control and quantification was assessed using the Tapestation (Agilent) and Qubit 2.0 (ThermoFisher Scientific). The Library was sequenced on a Nextseq500 (Illumina) instrument using the MID output kit (Illumina) (20024904).

value: mix of human naive pluripotent stem cells (H9 and Sigma), primed pluripotent stem cells (H9 and Sigma), trophoblast stem cells (Sigma) and extraembryonic mesoderm cells (Sigma), all samples were pooled in one.

tag: cell line

value: H9 (primed, and naive hPSCs) and Sigma (primed hPSCs, naive hPSCs, day 30 ASECRiAV conversion (trophobast stem cells and extraembryonic mesoderm cells)


title: T47D_LTED21M_DMSO_H3K4me2_ChIPseq_rep2

growth_protocol: T47D were maintained at 37°C with 5% CO2 according to ATCC recommendations. Cells were culture in  estrogen deprived conditions for 21 months. Cell culture media: IMEM with no phenol red (Corning 10-026-CV supplemented with 10% charcoal stripped FBS(Gemini 100-119) and Penicillyn/Streptomycin (Thermo Fisher Scientific 15140-12, 10000U/mL. Cells were treated with DMSO 48h

value: Mammary gland ductal carcarcinoma epithelial cells derived from metastatic site

value: Parental estrogen deprived for 21 months treated with DMSO 48h


title: sc_C49: BY4741, WT, 36-hour

city: Beijing

first: Jincheng

last: Wang

address: yiheyuan road No5

country: China

institute: BIOPIC, Peking University

library_source: transcriptomic single cell

data_processing:
Paired-end reads were mapped to the S288c Saccharomyces cerevisiae genome R64 version (www.yeastgenome.org) with ERCC spike-in sequences added using HISAT2 (version 2.1.0).
Resulting bam files were sorted and indexed using samtools (version 1.1).
Final read counts mapped to genes were extracted using FeatureCounts.
Assembly: S288c Saccharomyces cerevisiae genome R64.
Supplementary files format and content: tab-delimited text files include log2-normalized read counts for each Sample

organism: Saccharomyces cerevisiae

growth_protocol: BY4742 WT were grown on SD solid agar plates.

extract_protocol:
Yeast cells were lysed with ~4µl lysis buffer, containing 0.5%Triton, 2.5µM oligo-dT, 2.5mM dNTP (Invitrogen,1959189), 8000 molecules of external RNA control consortium (ERCC) spike in, 3 x 10-2U/µl zymolyase(ZYMO, E1004-A), 1U/µl Recombinant RNase Inhibitor(TaKaRa, AI41189A), and RNA released.
RNA libraries for RNA-seq were prepared using Smart-seq2 following manufacturer's protocols.

value: WT

tag: time

value: 36 hour


title: sc_C48: BY4741, WT, 36-hour


title: TS/A_Emp_ColIII_rep02

value: BALB/c

value: Gpr56/Adgrg1 expressing vector


title: TS/A_Emp_Control_rep01

value: Empty vector


title: sc_C47: BY4741, WT, 36-hour


title: 4T1_Emp_ColIII_rep02


title: sc_C46: BY4741, WT, 36-hour


title: Starved3

city: Glasgow

first: Bashir

middle: Abdirahman

last: Mohamed

state: Glasgow

address: Switchback Road

country: United Kingdom

institute: Beatson Institute for Cancer Research

data_processing:
Base calling - bcl2fastq 2
Adapter trimming using cutadapt
Alignment - bowtie2
Counts obtained
Expression data was normalized and differential expression done in DESEQ2
Genome_build: GRCh38
Supplementary_files_format_and_content: Raw tRNA counts in a .txt file

source_name: BJ-5Ta cells

growth_protocol: Cells were maintained in Dulbecco’s modified Eagle’s medium (DMEM) supplemented with 10% FBS, 1% pen/step and 2 mM L-glutamine (hereafter called complete medium). All cell lines were incubated at 37°C in a humidified atmosphere with 5% CO2.

extract_protocol:
RNA was extracted using the mirVana Small enriched RNA extraction protocol
Library prepared using ALICE-tRNA-seq: 3' Adapter ligation, RT, Circularization, Relinearization, PCR amplification

treatment_protocol: For serum starvation growth conditions cells were seed one day prior to serum starvation. Cells were washed twice with PBS and the media replaced with either 10% or 0% FBS containing DMEM for 72 hours.

value: 72 hours FBS-

value: quiescent fibroblasts


title: T47D_LTED21M_DMSO_H3K4me2_ChIPseq_rep1

value: ATCC #HTB-133


title: Starved2


title: sc_C45: BY4741, WT, 36-hour


title: Starved1


title: sc_C43: BY4741, WT, 36-hour


title: 4T1_Emp_Control_rep01


title: 4T1_Emp_Control_rep02


title: Fed3

value: 72 hours FBS+

value: proliferating fibroblasts


title: T47D_LTED9M_FOXA1_ChIPseq_rep2

value: Parental estrogen deprived for 9 months.

value: Anti-FOXA1 antibody - ChIP Grade (ab23738)


title: 4T1_Emp_ColIII_rep01


title: T47D_LTED_LSD1_KO_FOXA1_ChIPseq_rep2

value: LSD1 KO parental cells generated with CRISPR/Cas9


title: T47D_LTED_LSD1_KO_FOXA1_ChIPseq_rep1


title: T47D_LTED21M_Corin_Input_ChIPseq_rep1

value: Parental estrogen deprived for 21 months treated with Corin 48h


title: Fed1


title: sc_C44: BY4741, WT, 36-hour


title: T47D_LTED21M_DMSO_Input_ChIPseq_rep2


title: Fed2


title: T47D_LTED9M_FOXA1_ChIPseq_rep1


title: RBKO1_2_CTRL_H3K4me_7

city: Charlestown

first: Ioannis

last: Sanidas

state: MA

department: Cancer Center

institute: Massachusetts General Hospital

data_processing:
Sequencing reads were trimmed using trim-galore, aligned to the human genome (build hg19) using bwa-mem, then duplicate-marked using sambamba, and filtered to remove reads that were duplicates, had mapping quality <30, had >2 mismatches, or had >1/3 of their sequence soft-clipped.
Filtered BAMs were sorted and indexed, and deeptools-bamCoverage was used to generate CPM-normalized bigwig files for peak visualization, and bedtools-coverage was used to count reads in 200bp genomic windows tiling the genome.
Genome_build: human genome (build hg19)
Supplementary_files_format_and_content: CPM-normalized bigwig files were generated using deeptools-bamCoverage

source_name: retina epithelial cells RPE1

growth_protocol: The telomerase-expressing non-transformed human retina epithelial cells RPE1 and skin fibroblasts BJ cells were cultured in Dulbecco’s Modified Eagle’s Medium (DMEM) supplemented with 5% Fetal Bovine Serum (FBS) and antibiotics (100 units/ml Penicillin and 100 μg/ml Streptomycin; P/S). The MCF10A human breast epithelial cell line were cultured in DMEM/Nutricient mixture F12 medium supplemented with 5% horse serum (Gibco, Cat. No. 16050122), 20 ng/ml human EGF (ThermoFisher, Cat. No. PHG0311), 0.5 µg/ml hydrocortisone (Sigma, Cat. No. H-0888), 100 ng/ml cholera toxin (Sigma, Cat. No. C-8052), 10 µg/ml insulin (Sigma, Cat. No. I-1882) and antibiotics (P/S). The human breast cancer cell line T47D was cultured in RPMI 1640 medium supplemented with 10% FBS and antibiotics (P/S).

extract_protocol:
ChIP assays were carried out on RPE1, BJ, MCF10A and T47D cell cultures using approximately 5 million cells per sample and per epitope. Cells were washed once with PBS and fixed with 1.5 mM ethylene glycol bis(succinimidyl succinate) (Thermo Scientific, Cat. No. 21565) for 30 min at 25oC and 1% formaldehyde (Sigma, Cat. No. F8775) for 10 min at 37oC. 20mM Tris-HCl pH7.5 and 150 mM Glycine were used to quench cross-linking. Cell pellets were washed twice with ice-cold PBS and lysates were prepared using 300 µl lysis buffer (50mM Tris-HCl pH 7.4, 1% SDS, 0.25% DOC) supplement with protease and phosphatase inhibitors cocktail (PPI) (Thermo Scientific, Cat. No. 78442) for 10 min on ice. Cell lysates were diluted with 900 µl ChIP dilution buffer (50mM Tris-HCl pH 7.4, 0.1% SDS, 150mM NaCl, 1.84% Triton-X) supplement with PPI and chromatin was fragmented to a size of 200 – 700 bases with Branson 250 sonifier. Solubilized chromatin was further diluted with 1800 µl ChIP dilution buffer and immunoprecipitated with the indicated antibodies overnight at 4°C. Anti-DYKDDDDK Tag (Cell Signaling, Cat. No. 14793) was used for FLAG-RB ChIP. Antibody-chromatin complexes were pulled down with protein G-Dynabeads (Life Technologies, Cat. No. 10004D) for 2 hours at 4°C and washed 3 times with RIPA 150 Wash Buffer (0.1% SDS, 0.1% DOC 1% Triton X-100, 1mM EDTA, 10mM Tris-HCl pH 8, 150mM NaCl), 3 times with RIPA 500 Wash Buffer (0.1% SDS, 0.1% DOC 1% Triton X-100, 1mM EDTA, 10mM Tris-HCl pH 8, 500mM NaCl), 3 times with LiCl wash buffer (10mM Tris-HCl, pH 8, 250mM LiCl, 0.5% Triton X-100, 0.5% DOC) and once with ice-cold 10mM Tris-HCl, pH 8. Chromatin complexes were eluted from the beads using elution buffer (10mM Tris-HCl pH 8, 0.1% SDS, 150mM NaCl, 5 mM DTT) for 1 hour at 65°C and treated with RNase A (Roche, Cat. No. 11119915001) for 30 min at 37oC. Proteinase K (Thermo Scientific, Cat. No. 25530049) treatment and reverse cross-linking was performed for 3 hours at 65°C. After crosslink reversal, immunoprecipitated DNA was extracted with AMP Pure XP beads (Beckman Coulter, Cat. No. A63881). ChIP DNA was quantified with Qubit dsDNA HS Assay kit (Thermo Scientific, Cat. No. Q32851).
1-2 ng ChIP DNA samples were used to prepare sequencing libraries, using the Ovation Ultralow System V2 kit (TECAN, Cat. No. 0344NB-A01).

treatment_protocol: RB1 knock down or replacement of the endogenous by exogenous RB protein in cells transduced with pINDUCER11 and/or pINDUCER20 constructs were induced by addition of 0.5 μg/ml DOX (Sigma, Cat. No. D9891) for the indicating time. Expression of the dominant negative A-FOS or the dominant negative DP1_aa127-410 in cells transduced with pLenti-CMVtight puro constructs were induced by addition of 0.5 μg/ml DOX for 48 hours. Contact inhibition experiments performed by maintaining RPE1 cells in confluent culture for 6 days.

value: sgRB1 (Nicolay et al. 2015)

value: anti-Histone H3 (Mono-Methyl-Lys4) (Abcam, ab8895)


title: CIH3

first: 唐

last: 伟评

address: 2 Anzhen Rd, Chaoyang District,

institute: Beijing Institute of Heart Lung and Blood Vessel Diseases

data_processing:
Illumina  software used for basecalling.
Raw data(raw reads) of fastq format were firstly processed through in-houseperl scripts.In this step,clean data(clean reads) were obtained by removing reads containing adapter,reads containing ploy-N and low quality reads from raw data.
Reference genome and gene model annotation files were downloaded from genome website directly.Index of the reference genome was built using HiSeq 2500 and paired-end clean reads were aligned to the reference genome using HiSeq 2500.
Feature Counts v1.5.0-p3 was used to count the reads numbers mapped to each gene.And then FPKM of each gene was calculated based on the length of the gene and readscount mapped to this gene.
Differential expression analysis of two conditions/groups was performed using the DESeq R package (1.8.3).
Assembly: Mus_musculus.GRCm38
Supplementary files format and content: tab_delimited text files include FPKM values for each Sample

organism: Mus

source_name: adipocytes

extract_protocol:
Total miRNA was collected from fat cells using mirVnanTM miRNA kit (AM1561, Thermo Fisher Scientific, USA) and purified using an RNeasy mini kit (Qiagen, Valencia, CA, USA). A miRNA Library was generated using the HiSeq 2500 system according to the manufacturer’s instructions. Quality controls were performed and miRNA libraries were then sequenced at 10pM on the Illumina HiSeq 2500 to generate 150bp paired-end reads.
miRNA-seq libraries were prepared RNeasy mini kit.

value: Chronic intermittent hypoxia

value: adipocytes


title: sc_C41: BY4741, WT, 36-hour

value: WT

value: 36 hour


title: sc_C42: BY4741, WT, 36-hour


title: CIH1


title: CIH2


title: sc_C40: BY4741, WT, 36-hour


title: T47D_LTED21M_Corin_Input_ChIPseq_rep2

value: Mammary gland ductal carcarcinoma epithelial cells derived from metastatic site


title: RBwt_CTRL_H3K4me_5

value: sgControl (Nicolay et al. 2015)


title: sc_C39: BY4741, WT, 36-hour


title: Ctrl3

value: control


title: T47D_LTED21M_Corin_cjun_ChIPseq_rep2

value: Parental estrogen deprived for 21 months treated with Corin 1uM 48h

value: Anti-c-Jun antibody (ab31419)


title: RBKO1_2_CTRL_JUN_11

value: anti-c-JUN (Cell Signaling Technology, 9165)


title: T47D_LTED21M_DMSO_Input_ChIPseq_rep1

value: ATCC #HTB-133

value: Parental estrogen deprived for 21 months treated with DMSO 48h


title: sc_C38: BY4741, WT, 36-hour


title: Ctrl1


title: T47D_LTED21M_DMSO_cjun_ChIPseq_rep2


title: Orobanche tubercle- small RNAs

city: Kermanshah

first: Hooman

last: Salari

address: Emam khomaini hyw.

department: Production Engineering and Plant Genetics

country: Iran

institute: Razi University

description: small RNA

data_processing:
sRNA libraries were trimmed using Trim Galore and first aligned to S. lycopersicum reference genome (SL 3.0) using Bowtie with “–v0” (https://plants.ensembl.org/) to remove possible host-originated reads. The filtered reads were then subjected to mapping against the available transcriptome reference of O. aegyptiaca (http://ppgp.huck.psu.edu/) using Bowtie with the command “–v2”.
Conserved miRNAs were identified using miRProf with only genome-matching sRNAs. New miRNAs were identified using miRCat and only considering new miRNAs with the following characteristics: 21-22 nt in length, detection of miRNA* complement, at least 10 reads in total and identification in multiple bioreplicates.
Genome_build: FASTA with miRNA transcript sequences is available on the series record.

organism: Phelipanche aegyptiaca

source_name: Orobanche tubercle

growth_protocol: O. aegyptiaca seeds collected in bulk from several emerged shoots in a highly infested tomato field were grown in the controlled research greenhouse (15h natural light, temperature of 25 oC, and humidity of 25%) of the Razi University-Kermanshah, Iran, on Solanum lycopersicum as host.

extract_protocol:
12 biological replicates each containing materials from three attachments were pooled for total RNA isolation using TRIzol reagent (Life Technologies).
Accurate detected samples were sent to the Novogene Co., Ltd. (Beijing Nuohe Zhiyuan Biological Information Technology, China) for downstream steps toward small RNA sequencing on an Illumina Hiseq 2500 platform.

treatment_protocol: Tissue samples were harvested when O. aegyptiaca was at the “stage 4.2” of its life cycle (“late post-vascular connection stage” or “spider stage”) described by Westwood, et al. (2012).

value: tubercle


title: sc_C37: BY4741, WT, 36-hour


title: Ctrl2


title: Orobanche-tomato interface- small RNAs

description: small RNA

source_name: Orobanche-tomato interface

value: Orobanche-tomato interface


title: W_CI_F

value: pINDUCER11-shRB1, pINDUCER20-FLAG-RBWT

value: anti-DYKDDDDK Tag (Cell Signaling Technology, 14793)

value: 6 days Contact Inhibition in doxycycline


title: T47D_LTED21M_DMSO_cjun_ChIPseq_rep1


title: sc_C35: BY4741, WT, 36-hour


title: sc_C36: BY4741, WT, 36-hour


title: sc_C34: BY4741, WT, 36-hour


title: T47D_LTED9M_SMARCC1_ChIPseq_rep2

growth_protocol: T47D were maintained at 37°C with 5% CO2 according to ATCC recommendations. Cells were culture in  estrogen deprived conditions for 9 months. Cell culture media: IMEM with no phenol red (Corning 10-026-CV supplemented with 10% charcoal stripped FBS(Gemini 100-119) and Penicillyn/Streptomycin (Thermo Fisher Scientific 15140-12, 10000U/mL

value: Parental estrogen deprived for 9 months

value: SMARCC1/BAF155 (D7F8S) Rabbit mAb #11956 Cell Signaling Technology


title: WT_GFP_E2F1_19

value: anti-E2F1 (Cell Signaling Technology, 3742)

value: 48 hours doxycycline


title: rv3249c mutant - rep 2

city: Portland

first: Georgiana

last: Purdy

state: OR

address: 3181 SW Sam Jackson Park Rd

department: Molecular Microbiology and Immunology

institute: Oregon Health & Science University

data_processing:
Illumina CASAVA 1.8 software used for basecalling
Sequence reads assessed for adapters and quality scores using FastQC.
For each sample, sequence reads aligned to the reference genome using salmon v1.4.0. Library type setting SF <-l SF>
Sequence reads quantified for each sample using salmon v1.4.0
Differential expression assessed using DESeq2 v1.32.0
Assembly: GCF_000195955.2
Supplementary files format and content: Count file aggregating data from raw files

organism: Mycobacterium tuberculosis

source_name: bacterial culture

growth_protocol: Middlebrook 7H9 liquid medium (Difco) with 0.05% Tween-80 supplemented with albumin dextrose salts (ADS) containing 8.1 mg/mL NaCl, 50 mg/mL bovine serum albumin (BSA), and 20 mg/mL dextrose.

extract_protocol:
bacteria were harvested by centrifugation and the pellet washed in GTC buffer (4 M guanidine thiocyanate, 0.5% Na N-lauryl sarcosine, 25 mM Na citrate, pH 7.0, 0.1 M beta-mercaptoethanol) followed by phosphate-buffered saline (PBS)–0.1% Tween 80. Bacteria were disrupted by treatment with lysozyme followed by bead beating in the presence of warm TRIzol. RNA was subsequently isolated using DirectZol RNA isolation columns (Zymo). RNA was eluted in 50 L RNase-free water and then treated with DNase (Ambion) to ensure removal of genomic DNA contamination.
Starting with total RNA, libraries were prepared according to Illumina instructions. Ribosomal RNA was removed using Ribo-Zero rRNA. Stranded RNA Library Prep was performed on a Wafergen Bio-systems Apollo 324 Robot. Libraries were quantified by qPCR.

value: rv3249c mutant


title: WT_DNDP1_E2F1_20


title: rv3249c mutant - rep 3


title: T47D_LTED9M_RNApol_ChIPseq_rep2

extract_protocol:
Cells were grown to 80% confluence on 150cm plates and fixed with 11% solution containing formaldehyde (Thermo Fisher #28908), 50mM HEPES-KOH(pH 7.5), 100mM Nacl, 0.5M EDTA, 0.5M EGTA for 10 min. 1/20 of the plates volume equivalent of freshly prepared Glycine 2.5M(stock) was added to quench the reaction for 5min . The supernatant was aspirated, and the cells were washed twice with 5 ml PBS and harvested on ice using cell scrapers . Cells were pelleted at 2950 x g for 10 min at 4°C, washed 1x with 10 ml cold PBS, and pelleted once more. Cell pellets were flash freeze in liquid nitrogen and store at -80°C until processing day. Cell pellet collected from 6 plates of cells was resuspended in 5 ml of Lysis Buffer 1 (50 mM HEPES-KOH, pH 7.5; 140 mM NaCl, 1mM EDTA, 10% glycerol, 0.5% NP-40, 0.25% Triton X-100), allowed to rotate for 10 min at 4°C, and pelleted at 2950 x g for 10 min at 4°C. Cells were then resuspended in 5 ml of Lysis Buffer 2 (10 mM Tris-HCL pH 8.0, 200 mM NaCl, 1 mM EDTA, 0.5 mM EGTA), allowed to rotate for 10 min at room temperature, and pelleted at 2950 x g for 10 min at 4°C. Cells were then resuspended in 3 ml of Lysis Buffer 3 (10 mM Tris-HCl pH 8, 100 mM NaCl, 1 mM EDTA, 0.5 mM EGTA, 0.1% Na-Deoxycholate, 0.5% N-lauroylsarcosine) and then transfer to 15ml sonication tubes (Diagenode cat# C01020031). Cells were sonicated on a Bioruptor Pico at 4°C for 20min of 30’’ ON-OFF cycles with 1.2 g of sonication beads. After sonication, samples were transferred to 2 separate 1.5 ml microtube and centrifuged at 20,000 x g for 15 minutes at 4°C. The supernatant was transferred to new 15 ml tubes. To check sonication efficiency, 50µl of sonicated samples was transferred to a new microtube with 50µl PBS and incubated at 65°C for 3hr on an Eppendorf Thermomixer shaking at 1000rpm to decrosslink. DNA was purified with the QIAquick PCR Purification Kit (Qiagen cat# 28106) and eluted in 50µl of H20. 1 ug of decrosslinked chromatin from each sample were run in a 1% agarose gel at 100V for 45min and imaged on a Bio-rad ChemiDoc XRS+. DNA concentration in the sonicated decrosslinked samples were measured via qubit, and this concentration was then used to estimate the concentration of chromatin in the un-decrosslinked samples. 2ug of Antibody (see Key Resources table for list of antibodies) along with 0.4ug of Spike-in antibodies (Active Motif 61686) were bound to 50 ul of Dynabeads Protein G (Thermo Fisher 10004D), previously washed with 1 ml of 0.5% BSA in 1x PBS three times, and rotated end-to-end overnight at 4°C (Eppendorf cat# 0030108051). After binding of the antibodies, beads were washed with 1 ml of 0.5% BSA in 1x PBS three times. 10ug of chromatin and 20 ng of Spike-in chromatin (Active Motif 53083) were added to 1.5ml LoBind tube (Eppendorf cat# 0030108051) containing pre-bound antibodies beads and brought up to 500µl final volume with Lysis Buffer and rotated end-to-end overnight at 4°C.  5 µl was removed as input material (1%) and placed in a separate microtube at 4°C completing the volume to 50uL with Lysis Buffer 3.  Beads were washed 6 times with 1 ml Wash Buffer (50 mM Hepes-KOH, pH 7.6; 500 mM LiCl, 1 mM EDTA, 1% NP-40; 0.5% Na-Deoxycholate) and once with 1 ml TE containing 50 mM NaCl. Beads were dried after the last wash by removing as much TE as possible. 210 ul of Elution Buffer (50 mM Tris-HCl, pH 8; 10 mM EDTA, 1% SDS) was added to each sample and  150 ul elution buffer were added to each input sample that was previously set aside. To elute immunocomplexes from beads, samples were incubated at 65°C for 15 minutes on an Eppendorf Thermomixer shaking at 1000rpm. Tubes were centrifuged for 3min at 960xg at RT and 200µl of supernatant was transferred to a new tube and reverse crosslinked overnight at 65°C. Next day, 200 ul of TE was added to each decrosslinked sample and input, and the samples were treated sequentially with 0.2 mg/ml RNAse A (Sigma R5503) at 37°C and 0.2 ug/ml Proteinase K (NEB P8107S) at 55°C for two hours each. DNA purification was performed with the QIAquick PCR Purification Kit (Qiagen cat# 28106) and eluted in 50µl of H2O and quantified by Qubit.
Immunoprecipitated DNA was used to generate libraries using the NEBNext Ultra DNA Library Prep Kit for Illumina (New England Biolabs cat# 7370) following the manufacturer’s instructions.

value: Mammary gland ductal carcarcinoma epithelial cells derived from metastatic site

value: Rpb1 NTD (D8L4Y) Rabbit mAb #14958 Cell Signaling Technology


title: W_CIR_F

value: 6 days Contact Inhibition (C.I.) in doxycycline and release from C.I. for 24 hours


title: T47D_LTED9M_H3K27ac_ChIPseq_rep2

extract_protocol:
Cells were grown to 80% confluence on 150cm plates and fixed with 11% solution containing formaldehyde (Thermo Fisher #28908), 50mM HEPES-KOH(pH 7.5), 100mM Nacl, 0.5M EDTA, 0.5M EGTA for 10 min. 1/20 of the plates volume equivalent of freshly prepared Glycine 2.5M(stock) was added to quench the reaction for 5min . The supernatant was aspirated, and the cells were washed twice with 5 ml PBS and harvested on ice using cell scrapers . Cells were pelleted at 2950 x g for 10 min at 4°C, washed 1x with 10 ml cold PBS, and pelleted once more. Cell pellets were flash freeze in liquid nitrogen and store at -80°C until processing day. Cell pellet collected from 6 plates of cells was resuspended in 5 ml of Lysis Buffer 1 (50 mM HEPES-KOH, pH 7.5; 140 mM NaCl, 1mM EDTA, 10% glycerol, 0.5% NP-40, 0.25% Triton X-100), allowed to rotate for 10 min at 4°C, and pelleted at 2950 x g for 10 min at 4°C. Cells were then resuspended in 5 ml of Lysis Buffer 2 (10 mM Tris-HCL pH 8.0, 200 mM NaCl, 1 mM EDTA, 0.5 mM EGTA), allowed to rotate for 10 min at room temperature, and pelleted at 2950 x g for 10 min at 4°C. Cells were then resuspended in 3 ml of Lysis Buffer 3 (10 mM Tris-HCl pH 8, 100 mM NaCl, 1 mM EDTA, 0.5 mM EGTA, 0.1% Na-Deoxycholate, 0.5% N-lauroylsarcosine) and then transfer to 15ml sonication tubes (Diagenode cat# C01020031). Cells were sonicated on a Bioruptor Pico at 4°C for 20min of 30’’ ON-OFF cycles with 1.2 g of sonication beads. After sonication, samples were transferred to 2 separate 1.5 ml microtube and centrifuged at 20,000 x g for 15 minutes at 4°C. The supernatant was transferred to new 15 ml tubes. To check sonication efficiency, 50µl of sonicated samples was transferred to a new microtube with 50µl PBS and incubated at 65°C for 3hr on an Eppendorf Thermomixer shaking at 1000rpm to decrosslink. DNA was purified with the QIAquick PCR Purification Kit (Qiagen cat# 28106) and eluted in 50µl of H20. 1 ug of decrosslinked chromatin from each sample were run in a 1% agarose gel at 100V for 45min and imaged on a Bio-rad ChemiDoc XRS+. DNA concentration in the sonicated decrosslinked samples were measured via qubit, and this concentration was then used to estimate the concentration of chromatin in the un-decrosslinked samples.  10ug of Antibody (see Key Resources table for list of antibodies) along with 2ug of Spike-in antibodies (Active Motif 61686) were bound to 50 ul of Dynabeads Protein G (Thermo Fisher 10004D), previously washed with 1 ml of 0.5% BSA in 1x PBS three times, and rotated end-to-end overnight at 4°C (Eppendorf cat# 0030108051). After binding of the antibodies , beads were washed with 1 ml of 0.5% BSA in 1x PBS three times. 30ug of chromatin and 50 ng of Spike-in chromatin (Active Motif 53083) were added to 1.5ml LoBind tube (Eppendorf cat# 0030108051) containing pre-bound antibodies beads and brought up to 500µl final volume with Lysis Buffer and rotated end-to-end overnight at 4°C.  5 µl was removed as input material (1%) and placed in a separate microtube at 4°C completing the volume to 50uL with Lysis Buffer 3.  Beads were washed 6 times with 1 ml Wash Buffer (50 mM Hepes-KOH, pH 7.6; 500 mM LiCl, 1 mM EDTA, 1% NP-40; 0.5% Na-Deoxycholate) and once with 1 ml TE containing 50 mM NaCl. Beads were dried after the last wash by removing as much TE as possible. 210 ul of Elution Buffer (50 mM Tris-HCl, pH 8; 10 mM EDTA, 1% SDS) was added to each sample and  150 ul elution buffer were added to each input sample that was previously set aside. To elute immunocomplexes from beads, samples were incubated at 65°C for 15 minutes on an Eppendorf Thermomixer shaking at 1000rpm. Tubes were centrifuged for 3min at 960xg at RT and 200µl of supernatant was transferred to a new tube and reverse crosslinked overnight at 65°C. Next day, 200 ul of TE was added to each decrosslinked sample and input, and the samples were treated sequentially with 0.2 mg/ml RNAse A (Sigma R5503) at 37°C and 0.2 ug/ml Proteinase K (NEB P8107S) at 55°C for two hours each. DNA purification was performed with the QIAquick PCR Purification Kit (Qiagen cat# 28106) and eluted in 50µl of H2O and quantified by Qubit.
Immunoprecipitated DNA was used to generate libraries using the NEBNext Ultra DNA Library Prep Kit for Illumina (New England Biolabs cat# 7370) following the manufacturer’s instructions.

value: Anti-Histone H3 (acetyl K27) antibody - ChIP Grade (ab4729)


title: RBwt_CTRL_JUN_9

value: sgControl (Nicolay et al. 2015)

value: anti-c-JUN (Cell Signaling Technology, 9165)


title: rv3249c mutant - rep 1


title: WT_DNDP1_FLAG_2_18

description: replicate 2


title: T47D_LTED21M_Corin_cjun_ChIPseq_rep1

value: ATCC #HTB-133

value: Parental estrogen deprived for 21 months treated with Corin 1uM 48h

value: Anti-c-Jun antibody (ab31419)


title: WT_DNDP1_FLAG_17

description: replicate 1


title: sc_C31: BY4741, WT, 36-hour

value: WT

value: 36 hour


title: H37Rv WT - rep 3

value: wildtype


title: WT_A-FOS_FLAG_2_14

description: replicate 2

value: anti-DYKDDDDK Tag (Cell Signaling Technology, 14793)


title: T47D_LTED9M_H3K4me2_ChIPseq_rep2

value: Parental estrogen deprived for 9 months

value: Histone H3K4me2 antibody (pAb)   Active Motif 39141


title: T47D_LTED9M_CoREST_ChIPseq_rep2

value: Anti-CoREST Antibody 07-455 Sigma Aldrich


title: H37Rv WT - rep 2


title: W_AFOS_F

description: replicate 1

value: 48 hours doxycycline


title: sc_C30: BY4741, WT, 36-hour


title: H37Rv WT - rep 1


title: sc_C29: BY4741, WT, 36-hour


title: sc_C33: BY4741, WT, 36-hour


title: T47D_LTED9M_cjun_ChIPseq_rep2

value: Mammary gland ductal carcarcinoma epithelial cells derived from metastatic site


title: WT_GFP_FLAG_2_13

description: replicate 2


title: T47D_LTED9M_2x_linked_input_ChIPseq_rep2


title: W_GFP_F

description: replicate 1


title: T47D_LTED9M_1x-linked_Input_ChIPseq_rep2


title: SCR_CTCF

value: anti-CTCF (Cell Signaling Technology, 3418)


title: sc_C28: BY4741, WT, 36-hour


title: D5_CTCF


title: sc_C26: BY4741, WT, 36-hour


title: sc_C27: BY4741, WT, 36-hour


title: T47D_FM_SMARCC1_ChIPseq_rep1

growth_protocol: T47D were maintained at 37°C with 5% CO2 according to ATCC recommendations and split 1:3 every 3 days. Culture media: RPMI-1640(Lonza 12-167Q) supplemented with 10% FBS(Gemini 100-106), 1x penicillin/streptomycin(Thermo Fisher Scientific 15140-12, 10000U/mL), and 1x glutamax( Life Technologies 35050-061).

value: Parental

value: SMARCC1/BAF155 (D7F8S) Rabbit mAb #11956 Cell Signaling Technology


title: sc_C32: BY4741, WT, 36-hour


title: sc_C25: BY4741, WT, 36-hour


title: T47D_FM_LSD1_ChIPseq_rep2

extract_protocol:
Cells were grown to 80% confluence on 150cm plates and fixed with 11% solution containing formaldehyde (Thermo Fisher #28908), 50mM HEPES-KOH(pH 7.5), 100mM Nacl, 0.5M EDTA, 0.5M EGTA for 10 min. 1/20 of the plates volume equivalent of freshly prepared Glycine 2.5M(stock) was added to quench the reaction for 5min . The supernatant was aspirated, and the cells were washed twice with 5 ml PBS and harvested on ice using cell scrapers . Cells were pelleted at 2950 x g for 10 min at 4°C, washed 1x with 10 ml cold PBS, and pelleted once more. Cell pellets were flash freeze in liquid nitrogen and store at -80°C until processing day. Cell pellet collected from 6 plates of cells was resuspended in 5 ml of Lysis Buffer 1 (50 mM HEPES-KOH, pH 7.5; 140 mM NaCl, 1mM EDTA, 10% glycerol, 0.5% NP-40, 0.25% Triton X-100), allowed to rotate for 10 min at 4°C, and pelleted at 2950 x g for 10 min at 4°C. Cells were then resuspended in 5 ml of Lysis Buffer 2 (10 mM Tris-HCL pH 8.0, 200 mM NaCl, 1 mM EDTA, 0.5 mM EGTA), allowed to rotate for 10 min at room temperature, and pelleted at 2950 x g for 10 min at 4°C. Cells were then resuspended in 3 ml of Lysis Buffer 3 (10 mM Tris-HCl pH 8, 100 mM NaCl, 1 mM EDTA, 0.5 mM EGTA, 0.1% Na-Deoxycholate, 0.5% N-lauroylsarcosine) and then transfer to 15ml sonication tubes (Diagenode cat# C01020031). Cells were sonicated on a Bioruptor Pico at 4°C for 20min of 30’’ ON-OFF cycles with 1.2 g of sonication beads. After sonication, samples were transferred to 2 separate 1.5 ml microtube and centrifuged at 20,000 x g for 15 minutes at 4°C. The supernatant was transferred to new 15 ml tubes. To check sonication efficiency, 50µl of sonicated samples was transferred to a new microtube with 50µl PBS and incubated at 65°C for 3hr on an Eppendorf Thermomixer shaking at 1000rpm to decrosslink. DNA was purified with the QIAquick PCR Purification Kit (Qiagen cat# 28106) and eluted in 50µl of H20. 1 ug of decrosslinked chromatin from each sample were run in a 1% agarose gel at 100V for 45min and imaged on a Bio-rad ChemiDoc XRS+. DNA concentration in the sonicated decrosslinked samples were measured via qubit, and this concentration was then used to estimate the concentration of chromatin in the un-decrosslinked samples.  5ug of Antibody (see Key Resources table for list of antibodies) along with 1ug of Spike-in antibodies (Active Motif 61686) were bound to 50 ul of Dynabeads Protein G (Thermo Fisher 10004D), previously washed with 1 ml of 0.5% BSA in 1x PBS three times, and rotated end-to-end overnight at 4°C (Eppendorf cat# 0030108051). After binding of the antibodies , beads were washed with 1 ml of 0.5% BSA in 1x PBS three times. 30ug of chromatin and 50 ng of Spike-in chromatin (Active Motif 53083) were added to 1.5ml LoBind tube (Eppendorf cat# 0030108051) containing pre-bound antibodies beads and brought up to 500µl final volume with Lysis Buffer and rotated end-to-end overnight at 4°C.  5 µl was removed as input material (1%) and placed in a separate microtube at 4°C completing the volume to 50uL with Lysis Buffer 3.  Beads were washed 6 times with 1 ml Wash Buffer (50 mM Hepes-KOH, pH 7.6; 500 mM LiCl, 1 mM EDTA, 1% NP-40; 0.5% Na-Deoxycholate) and once with 1 ml TE containing 50 mM NaCl. Beads were dried after the last wash by removing as much TE as possible. 210 ul of Elution Buffer (50 mM Tris-HCl, pH 8; 10 mM EDTA, 1% SDS) was added to each sample and  150 ul elution buffer were added to each input sample that was previously set aside. To elute immunocomplexes from beads, samples were incubated at 65°C for 15 minutes on an Eppendorf Thermomixer shaking at 1000rpm. Tubes were centrifuged for 3min at 960xg at RT and 200µl of supernatant was transferred to a new tube and reverse crosslinked overnight at 65°C. Next day, 200 ul of TE was added to each decrosslinked sample and input, and the samples were treated sequentially with 0.2 mg/ml RNAse A (Sigma R5503) at 37°C and 0.2 ug/ml Proteinase K (NEB P8107S) at 55°C for two hours each. DNA purification was performed with the QIAquick PCR Purification Kit (Qiagen cat# 28106) and eluted in 50µl of H2O and quantified by Qubit.
Immunoprecipitated DNA was used to generate libraries using the NEBNext Ultra DNA Library Prep Kit for Illumina (New England Biolabs cat# 7370) following the manufacturer’s instructions.

value: Anti-KDM1 / LSD1 antibody - ChIP Grade (ab17721)


title: T47D_FM_SMARCC1_ChIPseq_rep2


title: D5_cJUN

value: anti-c-JUN (Cell Signaling Technology, 9165)


title: sc_C24: BY4741, WT, 36-hour


title: SCR_cJUN


title: T47D_FM_RCOR1_ChIPseq_rep2

value: ATCC #HTB-133


title: dCDK_H3K4me3

value: anti-Histone H3 (Tri-Methyl-Lys4) (Cell Signaling Technology, 9751)


title: T47D_FM_2x_input_ChIPseq_rep2


title: sc_C22: BY4741, WT, 36-hour


title: Mf_15w-ivc_MS236_077

data_processing:
The processing of the 3-prime-sequencing data for calculating the gene-expression levels was performed as described previously (Nakamura et al., 2015).  In brief, all reads were processed with cutadapt v1.9.1 to remove adaptors and poly-A sequences.
The reads of 30-bp or longer were mapped onto a mouse transcript reference, ref_macFas5, annotation 101, with 3'-end of transcripts, extended by up to 10kb, using Tophat v2.0.11/Bowtie2 v2.2.7.  Read counts per gene were calculated using HTSeq v0.9.1 with the default settings.  Read counts were converted to read counts per million mapped reads (RPM) and used for the analysis.
Genome_build: macFas5
Supplementary_files_format_and_content: Tab-delimited text files include RPM values for each sample.

organism: Macaca fascicularis

source_name: An oocyte from 15w-ivc rOvary

extract_protocol:
Fetal ovaries collected from cy fetuses at 8-18 wpf were carefully dissected out in chilled PBS.  Fetal ovaries at 8-14 wpf were minced with scissors in 0.05% trypsin/EDTA/PBS solution (GIBCO, 15400-054), then incubated for 15 min at 37℃ with gentle pipetting every 5 min.  Dissociation medium [DMEM (GIBCO, 10313021) containing 10% FBS, 2mM L-glutamine (GIBCO, 25030081), 1X penicillin/streptomycin (GIBCO, 15070063), and 100μg/mL DNaseI (MERCK, DN25)] was added for further dissociation. For fetal ovaries at 16 and 18 wpf, minced tissues were incubated in Accumax (Innovative Cell Technologies, AM105) for 40 min at 37℃ with gentle pipetting every 5 min.  Dissociation medium was added for further dissociation.  For SC3-seq analysis, cells were picked under the inverted microscope (OLYMPUS, IX71) to exclude multiplet cells.   For dissociating cy rOvaries, tissues were incubated with TryPLE Express (1X) (GIBCO, 12604021) for 5 min.  After adding dissociation medium, cells were processed for single-cell analyses. Dissociated germ cells from in vivo monkey fetal ovaries and monkey rOvaries were manually picked using glass capillaries under the stereo microscope (Leica, S8 APO).  Following cDNA synthesis and amplification methods were previously described (Nakamura et al., 2015).
The cDNA synthesis was performed according to the single-cell 3 prime RNA-seq (SC3-seq) method described previously (Nakamura et al., 2015).  The cDNA libraries for Nextseq500 (Illumina) were constructed as described previously (Ishikura et al., 2016).The sequencing was performed on an Illumina Nextseq500/550 platform with a Nextseq500/550 Hiqh output kit (Illumina).

tag: developmental stage

tag: xenotransplantation period

value: 15 weeks

value: Germ cell


title: sc_C23: BY4741, WT, 36-hour

value: WT

value: 36 hour


title: T47D_FM_cjun_ChIPseq_rep1

tag: cell type

value: Anti-c-Jun antibody (ab31419)


title: WT_H3K4me3

value: pINDUCER11-shRB1, pINDUCER20-FLAG-RBWT


title: sc_C19: BY4741, WT, 36-hour


title: T47D_FM_cjun_ChIPseq_rep2


title: T47D_FM_2x_input_ChIPseq_rep1


title: sc_C20: BY4741, WT, 36-hour


title: dCDK_H3K4me

value: anti-Histone H3 (Mono-Methyl-Lys4) (Abcam, ab8895)


title: Mf_12w-ivc_MS236_215

source_name: An oocyte from 12w-ivc rOvary

tag: in vitro culture period

value: 12 weeks


title: dCDK_H3K27ac

value: anti-Histone H3 (Acetyl-Lys27) (Active Motif, 39034)

value: 48 hours doxycycline


title: sc_C21: BY4741, WT, 36-hour


title: sc_C17: BY4741, WT, 36-hour


title: WT_H3K27ac


title: Mf_12w-ivc_MS236_214


title: sc_C18: BY4741, WT, 36-hour


title: Mf_12w-ivc_MS236_213


title: WT_H3K4me


title: Mf_12w-ivc_MS233_118


title: sc_C16: BY4741, WT, 36-hour


title: W_E2F1

value: anti-E2F1 (Cell Signaling Technology, 3742)


title: T47D_FM_Input_ChIPseq_rep2

value: Mammary gland ductal carcarcinoma epithelial cells derived from metastatic site

value: Parental


title: T47D_FM_ER_ChIPseq_rep2

growth_protocol: T47D were maintained at 37°C with 5% CO2 according to ATCC recommendations and split 1:3 every 3 days. Culture media: RPMI-1640(Lonza 12-167Q) supplemented with 10% FBS(Gemini 100-106), 1x penicillin/streptomycin(Thermo Fisher Scientific 15140-12, 10000U/mL), and 1x glutamax( Life Technologies 35050-061 ).

value: Anti-Estrogen Receptor alpha antibody (ab3575)


title: T47D_FM_FOXA1_ChIPseq_rep2

value: Anti-FOXA1 antibody - ChIP Grade (ab23738)


title: Mf_12w-ivc_MS233_108


title: sc_C15: BY4741, WT, 36-hour


title: sc_C14: BY4741, WT, 36-hour


title: INPUT_FLAG_RB_WT

value: input


title: T47D_FM_RNApol_ChIPseq_rep2

value: ATCC #HTB-133

value: Rb monoclonal Rpb1 NTD CST 14958S


title: FLAG_RB_dCDK_T826

value: anti-DYKDDDDK Tag (Cell Signaling Technology, 14793)


title: T47D_LTED21m6d_Corin_rep2

description: T47D_LTED21m6d_Corin_rep.pooled.pval.signal.bigwig

growth_protocol: T47D were maintained at 37°C with 5% CO2 according to ATCC recommendations. Cells were were culture in estrogen deprived conditions for 21 months. Cell culture media: IMEM with no phenol red (Corning 10-026-CV supplemented with 10% charcoal stripped FBS(Gemini 100-119) and Penicillyn/Streptomycin (Thermo Fisher Scientific 15140-12, 10000U/mL).Cells were treated for 72h with 500nM of compound Corin (MedChem Express

extract_protocol:
ATAC-seq experiments were performed as previously described (Corces et al., 2017) with minor modifications. 50,000 viable cells were used to perform the transposition reaction.The cleanup of the transposed fragments was performed using Zymo DNA Clean and Concentrator-5 Kit (Zymo catalog number D4014) and DNA was eluted in 21 µL of provided elution buffer
The ATAC-seq library construction was performed as previously described (Corces et al, 2017). To determine the average fragment size of each library, samples were run through a high sensitivity DNA screentape (Agilent Technologies #5067–5584) following the manufacturer’s instructions on an Agilent Technologies 2200 TapeStation machine. Concentration of each library was measured using Qubit dsDNA high sensitivity reagents (Thermo Fisher Scientific #Q32851) following the manufacturer’s instructions on a Qubit 3 fluorometer. .

value: Parental cells estrogen deprived 21 months treated with Corin


title: sc_C13: BY4741, WT, 36-hour

value: WT

value: 36 hour


title: T47D_LTED21m6d_DMSO_rep1

description: T47D_LTED21m6d_DMSO_rep.pooled.pval.signal.bigwig

growth_protocol: T47D were maintained at 37°C with 5% CO2 according to ATCC recommendations. Cells were were culture in estrogen deprived conditions for 3 days. Cell culture media: IMEM with no phenol red (Corning 10-026-CV supplemented with 10% charcoal stripped FBS(Gemini 100-119) and Penicillyn/Streptomycin (Thermo Fisher Scientific 15140-12, 10000U/mL). Cells were treated with DMSO for 72h.

value: Parental cells estrogen deprived 21 months treated with DMSO


title: Mf_12w-ivc_MS233_101

value: Germ cell


title: Mf_12w-ivc_MS239_202

value: 12 weeks


title: FLAG_RB_dCDK_T821


title: T47D_LTED21m6d_Corin_rep1


title: FLAG_RB_dCDK_S811


title: hMSMCs_input

city: Guangzhou

first: Kaiyuan

last: Ji

address: No.9 Jinsui Road, Guangzhou, China

department: Guangzhou Women and Children’s Medical Center

institute: Guangzhou Medical University

data_processing:
Raw sequencing data was first filtered by Trimmomatic (version 0.36), low-quality reads were discarded and the reads contaminated with adaptor sequences were trimmed. The clean reads were used for protein binding site analysis.
ChIP-seq reads were aligned to the hg38 genome assembly using STAR software (version 2.5.3a) with default configurations.
The RSeQC (version 2.6) was used for reads distribution analysis.The MACS2 software (Version 2.1.1) was used for peak calling. The bedtools（Version 2.25.0）was used for peaks annotation and peak distribution analysis.
Genome_build: hg38
Supplementary_files_format_and_content: bigwig files

source_name: hMSMCs

growth_protocol: Myometrium from non-laboring pregnant women were collected and washed with cold PBS until there was no obvious blood stain. Endometrium and vessels, if present, also were weeded out. The sample was cut into small pieces (5mm × 5mm × 5mm) and inoculated onto several culture dishes. After a 14-day culture with DMEM containing 10% fetal bovine serum (#10099-141, Gibco) and 1% penicillin-streptomycin (10000 U/mL, #15140122, Gibco), cells were harvested and subcultured for further identification via immunofluorescence.

extract_protocol:
Lysates were clarified from sonicated nuclei and histone-DNA complexes were isolated with antibody.
The cells were fixed in 1% formaldehyde for 10 min at room temperature, followed by the addition of glycine to 0.125M final concentration for terminating the crosslinking reaction. Following 5 min of formaldehyde quenching, plates were placed on ice and washed three times with ice-cold PBS and the cells were then collected and frozen in liquid nitrogen.ChIP assay was performed by SeqHealth (Wuhan, China). The cells were treated with nucleus lysis buffer and sonicated to fragment chromatin DNA of ~200-1000 bp. The 10% lysis sonicated chromatin was stored and named “input”, and 80% was used in immunoprecipitation reactions with anti-HIF-1α antibody（#36169S, Cell signaling technology, Massachusetts, USA）and named “IP”, and 10% was incubated with rabbit IgG (Cell Signaling Technology) as a negative control and named “IgG”, respectively. The DNA of input and IP was extracted by phenol-chloroform method. The high-throughput DNA sequencing libraries were prepared by using VAHTS Universal DNA Library Prep Kit for Illumina V3 (Catalog NO. ND607, Vazyme). The library products corresponding to 200-500 bps were enriched, quantified and finally sequenced on Novaseq 6000 sequencer (Illumina) with PE150 model.
ChIP assay was performed by SeqHealth (Wuhan, China). The cells were treated with nucleus lysis buffer and sonicated to fragment chromatin DNA of ~200-1000 bp. The 10% lysis sonicated chromatin was stored and named “input”, and 80% was used in immunoprecipitation reactions with anti-HIF-1α antibody

treatment_protocol: Once the cellular fusion degree reached more than 90%, hMSMCs were sent into the hypoxic workstation (H35, Don Whitley Scientific, United Kingdom) under the set condition: 3%O2, 5%CO2, 92%N2 for 2 hours.

value: human uterine smooth muscle cells (hMSMCs)

tag: tissue

value: uterine

value: hypoxia


title: Mf_12w-ivc_MS233_099


title: FLAG_RB_dCDK_S788

value: 48 hours doxycycline


title: INPUT_FLAG_RB_dCDK


title: sc_C9: BY4741, WT, 36-hour


title: Mf_12w-ivc_MS233_100


title: sc_C12: BY4741, WT, 36-hour


title: T47D_FM_H3K27ac_ChIPseq_rep2

value: Mammary gland ductal carcarcinoma epithelial cells derived from metastatic site

value: Parental

value: Anti-Histone H3 (acetyl K27) antibody - ChIP Grade (ab4729)


title: T47D_LTED21m6d_DMSO_rep2

growth_protocol: T47D were maintained at 37°C with 5% CO2 according to ATCC recommendations. Cells were were culture in estrogen deprived conditions for 3 days. Cell culture media: IMEM with no phenol red (Corning 10-026-CV supplemented with 10% charcoal stripped FBS(Gemini 100-119) and Penicillyn/Streptomycin (Thermo Fisher Scientific 15140-12, 10000U/mL).Cells were treated with DMSO for 72h.


title: sc_C11: BY4741, WT, 36-hour


title: FLAG_RB_dCDK_S795


title: Mf_12w-ivc_MS233_098


title: FLAG_RB_dCDK_S807


title: sc_C7: BY4741, WT, 36-hour


title: hMSMCs_HIF1A


title: Mf_12w-ivc_MS238_220


title: sc_C10: BY4741, WT, 36-hour


title: FLAG_RB_dCDK_S780


title: Mf_12w-ivc_MS233_095


title: Mf_12w-ivc_MS233_097


title: sc_C5: BY4741, WT, 36-hour


title: FLAG_RB_dCDK_S608


title: poly-aden.un1

city: Parkville

first: Viacheslav

last: Kriachkov

state: VIC

address: 30 Flemington Road

department: Department of Biochemistry and Pharmacology

country: Australia

institute: University of Melbourne

data_processing:
Real Time Analysis (RTA) v.3.4.4 was used for basecalling. Then the Illumina bcl2fastq 2.20.0.422 pipeline was used to generate the sequence data.
Reads were trimmed for adaptor sequence using Cutadapt (v.3.4)
Trimmed reads were mapped to hg38 human genome using HISAT2 (v.2.1.0). Aligned reads were counted using featureCount tool
Differential gene expression analysis was performed using DESeq2 (v.1.30.1). The comparisons were made between the following groups of samples: 102xPR.IND vs ChFP.IND, poly-aden.IND vs ChFP.IND
Genome_build: hg38
Supplementary_files_format_and_content: Matrix table with raw gene counts for every gene and every sample
Supplementary_files_format_and_content: Tab-delimited text files with the analysis for differentially expressed genes (output of DESeq2)

source_name: Human embryonic kidney cells

growth_protocol: HEK293T cells, obtained originally from the American Type Culture Collection (ATCC, Manassas, Virginia), were maintained in complete DMEM (Dulbecco’s modified Eagle medium (DMEM) supplemented with 10% v/v fetal calf serum and 1% GlutaMAX).  Cells were cultured in a humidified incubator with 5% v/v atmospheric CO2 at 37 °C.

extract_protocol:
Cell were collected in ice-cold lysis buffer (20 mM Tris pH 7.4, 150 mM NaCl, 5 mM MgCl2, 1 mM DTT, 100 μg/ml cycloheximide, 1% v/v Triton X-100 and 25 U/ml Turbo DNase I), triturated ten times through a 26-gauge needle and centrifuged for 10 min at 20,000 g at 4 °C. Total RNA was isolated by Zymo Research Direct-zol RNA MiniPrep Kit.
RNA libraries were prepared using TruSeq mRNA Library Prep with polyA selection. Libraries were sequenced on the Illumina NovaSeq 6000 (100bp Single Read, 20M reads) following the manufacturer's protocols.

treatment_protocol: For the induced samples, cells were treated with 500 ng/μl doxycycline for 48 h prior to harvesting.

value: Human embryonic kidney cells

tag: construct

value: poly-adenylate

tag: construct expression

value: uninduced


title: human fetal ovary, 11w4d

description: Single cell transcriptome of human fetal ovary

data_processing:
Raw data were demultiplexed with the mkfastq command in Cell Ranger (v5.0.1) with default setting to generate fastq files. The fastq files were processed with count command in Cell Ranger with default setting to map to the reference genome for human genome, GRCh38.p12 (PAR of chrY is hard masked).
Cell Ranger count command were also performed with transcript reference including long non-coding RNAs (data marked with "_include_lncRNA").
Genome_build: GRCh38.p12
Supplementary_files_format_and_content: Cell Ranger output files

source_name: fetal ovary

extract_protocol:
The human fetal ovary was collected after informed consent from participants undergoing elective abortion or gynecological procedures.  All experimental procedures were approved by the Institutional Review Board at Kyoto University (G1047 and G1192).  The fetal ovary were carefully dissected out in chilled PBS and minced with scissors in 0.05% trypsin/EDTA/PBS solution (GIBCO, 15400-054), then incubated for 15 min at 37℃ with gentle pipetting every 5 min.  Dissociation medium [DMEM (GIBCO, 10313021) containing 10% FBS, 2mM L-glutamine (GIBCO, 25030081), 1X penicillin/streptomycin (GIBCO, 15070063), and 100μg/mL DNaseI (MERCK, DN25)] was added for further dissociation.  Dissociated cells  were stained with DRAQ7 to distinguish dead cells.  All DRAQ7(-) live cells were collected by using FACS (BD Biosciences, FACS Aria III).
The cDNA library for NGS was conducted with Chromium Next GEM Single Cell 3' Kit v3.1 (Single Index) according to the manufacturer’s instructions.
scRNA-seq

value: fetal

tag: age

value: ovary

tag: scrna-seq protocol


title: Mf_12w-ivc_MS233_094


title: sc_C4: BY4741, WT, 36-hour

tag: genotype

value: WT


title: sc_C6: BY4741, WT, 36-hour

value: 36 hour


title: Mf_9w-ivc_MS236_223

source_name: An oocyte from 9w-ivc rOvary

value: 9 weeks


title: sc_C3: BY4741, WT, 36-hour


title: Mf_12w-ivc_MS239_201

value: Germ cell


title: FLAG_RB_dCDK_S612

value: anti-DYKDDDDK Tag (Cell Signaling Technology, 14793)

tag: treatment


title: poly-aden.un3


title: sc_C8: BY4741, WT, 36-hour


title: poly-aden.un2


title: Mf_9w-ivc_MS236_222


title: FLAG_RB_dCDK_T373

tag: chip antibody


title: sc_C1: BY4741, WT, 36-hour


title: FLAG_RB_dCDK_T356


title: FLAG_RB_dCDK_S249


title: poly-aden.IND2

description:
A2_HFWFNDRXY_CGATGCGGTT-ACCAAGCAGG
poly-aden_deseq2.txt

value: induced


title: poly-aden.IND3

description:
A3_HFWFNDRXY_TCCGGACTAG-ATTGTTCGTC
poly-aden_deseq2.txt


title: poly-aden.IND1

description:
A1_HFWFNDRXY_CTAGATTGCG-CGCCATATCT
poly-aden_deseq2.txt


title: FLAG_RB_dCDK_T252


title: Mf_9w-ivc_MS236_220


title: sc_C2: BY4741, WT, 36-hour


title: 102xPR.un3


title: FLAG_RB_dCDK_S230


title: Mf_9w-ivc_MS236_219


title: Mf_9w-ivc_MS236_221


title: sc_B49: BY4741, WT, 16-hour

value: 16 hour


title: 102xPR.un2


title: FLAG_RB_dCDK


title: sc_B48: BY4741, WT, 16-hour


title: Mf_9w-ivc_MS236_218


title: sc_B46: BY4741, WT, 16-hour


title: 102xPR.IND3

description:
PR3_HFWFNDRXY_TTAACCTTCG-TGTCTGGCCT
102xPR_deseq2.txt


title: Mf_9w-ivc_MS231_147


title: notag_RB_dCDK


title: sc_B47: BY4741, WT, 16-hour


title: 102xPR.un1


title: Mf_9w-ivc_MS236_217


title: 102xPR.IND2

description:
PR2_HFWFNDRXY_CGTGTACCAG-AACTTATCCT
102xPR_deseq2.txt


title: 102xPR.IND1

description:
PR1_HFWFNDRXY_ACTTGTTATC-TCTAGGCGCG
102xPR_deseq2.txt


title: ChFP.un3

value: ChFP


title: sc_B45: BY4741, WT, 16-hour


title: notag_RB_WT

value: pINDUCER11-shRB1, pINDUCER20-notag-RBWT

value: 48 hours doxycycline


title: sc_B44: BY4741, WT, 16-hour


title: sc_B43: BY4741, WT, 16-hour


title: FLAG_RB_WT

value: pINDUCER11-shRB1, pINDUCER20-FLAG-RBWT


title: Mf_6w-ivc_MS231_092

source_name: An oocyte from 6wks-ivc rOvary

value: 6 weeks


title: ChFP.un2

value: Human embryonic kidney cells


title: sc_B42: BY4741, WT, 16-hour


title: Mf_6w-ivc_MS238_215


title: ChFP.un1


title: Mf_6w-ivc_MS263_229


title: T47D_FM_RNApol_ChIPseq

data_processing:
ChIP-seq: Single-end ChIP-seq fastq files were processed using the ENCODE Transcription Factor and Histone ChIP-Seq processing pipeline (https://github.com/ENCODE-DCC/chip-seq-pipeline2). Reads were trimmed using cutadapt v2.5. Reads were aligned to the hg19 genome using Bowtie2 v2.3.4.3, and the SAMtools v1.9 was used to convert the output file to the BAM format. Duplicates were removed using Picard Tools v2.20.7. Peaks were called/signal tracks were generated using MACS2 v2.2.4, and peaks with fold change > 3.5 compared to input and a FDR < 0.05 were used for subsequent analysis. Only default parameters were used.
RNA-seq: Single-end RNA-seq fastq files were processed using the LSF-RNAseq Pipeline from Diderote (https://github.com/diderote/LSF-RNAseq). Reads were trimmed using cutadapt v2.3 with the following parameters: -j 4 --netseq-trim=20 -m 18. Reads were aligned to the hg19 genome using STAR v2.6.1a along default parameters, and RSEM v1.3.0 was used to obtain the expected gene counts against the human refseq also along default parameters.
RNA-seq differential expression was determined using DESeq2 v1.18.1 and R (version 3.4.1) with q-value <0.05. Ruvseq v1.12.0 was used for factor analysis to remove count based on ERCC spike-in. GSEA analysis was performed using gsea-3.0.jar. These additional analysis steps were included in the Diderote LSF-RNAseq Pipeline.
Bedtools v2.29.0 intersect was used to determine peak overlaps and assign target genes.
Homer v4.11 was used for peak annotation and motif analysis (http://homer.ucsd.edu/homer/data/software/homer.v4.11.1.zip)
Genome_build: hg19
Supplementary_files_format_and_content: The processed data files are in bigWig format containing the indexed tag chromosome coordinates and score produced from MACS2 peak calling.

extract_protocol:
Cells were grown to 80% confluence on 150cm plates and fixed with 11% solution containing formaldehyde (Thermo Fisher #28908), 50mM HEPES-KOH(pH 7.5), 100mM Nacl, 0.5M EDTA, 0.5M EGTA for 10 min. 1/20 of the plates volume equivalent of freshly prepared Glycine 2.5M(stock) was added to quench the reaction for 5min . The supernatant was aspirated, and the cells were washed twice with 5 ml PBS and harvested on ice using cell scrapers . Cells were pelleted at 2950 x g for 10 min at 4°C, washed 1x with 10 ml cold PBS, and pelleted once more. Cell pellets were flash freeze in liquid nitrogen and store at -80°C until processing day. Cell pellet collected from 6 plates of cells was resuspended in 5 ml of Lysis Buffer 1 (50 mM HEPES-KOH, pH 7.5; 140 mM NaCl, 1mM EDTA, 10% glycerol, 0.5% NP-40, 0.25% Triton X-100), allowed to rotate for 10 min at 4°C, and pelleted at 2950 x g for 10 min at 4°C. Cells were then resuspended in 5 ml of Lysis Buffer 2 (10 mM Tris-HCL pH 8.0, 200 mM NaCl, 1 mM EDTA, 0.5 mM EGTA), allowed to rotate for 10 min at room temperature, and pelleted at 2950 x g for 10 min at 4°C. Cells were then resuspended in 3 ml of Lysis Buffer 3 (10 mM Tris-HCl pH 8, 100 mM NaCl, 1 mM EDTA, 0.5 mM EGTA, 0.1% Na-Deoxycholate, 0.5% N-lauroylsarcosine) and then transfer to 15ml sonication tubes (Diagenode cat# C01020031). Cells were sonicated on a Bioruptor Pico at 4°C for 20min of 30’’ ON-OFF cycles with 1.2 g of sonication beads. After sonication, samples were transferred to 2 separate 1.5 ml microtube and centrifuged at 20,000 x g for 15 minutes at 4°C. The supernatant was transferred to new 15 ml tubes. To check sonication efficiency, 50µl of sonicated samples was transferred to a new microtube with 50µl PBS and incubated at 65°C for 3hr on an Eppendorf Thermomixer shaking at 1000rpm to decrosslink. DNA was purified with the QIAquick PCR Purification Kit (Qiagen cat# 28106) and eluted in 50µl of H20. 1 ug of decrosslinked chromatin from each sample were run in a 1% agarose gel at 100V for 45min and imaged on a Bio-rad ChemiDoc XRS+. DNA concentration in the sonicated decrosslinked samples were measured via qubit, and this concentration was then used to estimate the concentration of chromatin in the un-decrosslinked samples. 5ug of Antibody (see Key Resources table for list of antibodies) along with 1ug of Spike-in antibodies (Active Motif 61686) were bound to 50 ul of Dynabeads Protein G (Thermo Fisher 10004D), previously washed with 1 ml of 0.5% BSA in 1x PBS three times, and rotated end-to-end overnight at 4°C (Eppendorf cat# 0030108051). After binding of the antibodies , beads were washed with 1 ml of 0.5% BSA in 1x PBS three times. 30ug of chromatin and 50 ng of Spike-in chromatin (Active Motif 53083) were added to 1.5ml LoBind tube (Eppendorf cat# 0030108051) containing pre-bound antibodies beads and brought up to 500µl final volume with Lysis Buffer and rotated end-to-end overnight at 4°C.  5 µl was removed as input material (1%) and placed in a separate microtube at 4°C completing the volume to 50uL with Lysis Buffer 3.  Beads were washed 6 times with 1 ml Wash Buffer (50 mM Hepes-KOH, pH 7.6; 500 mM LiCl, 1 mM EDTA, 1% NP-40; 0.5% Na-Deoxycholate) and once with 1 ml TE containing 50 mM NaCl. Beads were dried after the last wash by removing as much TE as possible. 210 ul of Elution Buffer (50 mM Tris-HCl, pH 8; 10 mM EDTA, 1% SDS) was added to each sample and  150 ul elution buffer were added to each input sample that was previously set aside. To elute immunocomplexes from beads, samples were incubated at 65°C for 15 minutes on an Eppendorf Thermomixer shaking at 1000rpm. Tubes were centrifuged for 3min at 960xg at RT and 200µl of supernatant was transferred to a new tube and reverse crosslinked overnight at 65°C. Next day, 200 ul of TE was added to each decrosslinked sample and input, and the samples were treated sequentially with 0.2 mg/ml RNAse A (Sigma R5503) at 37°C and 0.2 ug/ml Proteinase K (NEB P8107S) at 55°C for two hours each. DNA purification was performed with the QIAquick PCR Purification Kit (Qiagen cat# 28106) and eluted in 50µl of H2O and quantified by Qubit.
Immunoprecipitated DNA was used to generate libraries using the NEBNext Ultra DNA Library Prep Kit for Illumina (New England Biolabs cat# 7370) following the manufacturer’s instructions.

value: Mammary gland ductal carcarcinoma epithelial cells derived from metastatic site

tag: cell source

value: ATCC #HTB-133

tag: modifications

value: Parental

value: Rpb1 NTD (D8L4Y) Rabbit mAb #14958 Cell Signaling Technology


title: T47D_FM_H3K27ac_ChIPseq

value: Anti-Histone H3 (acetyl K27) antibody - ChIP Grade (ab4729)


title: T47D_FM_FOXA1_ChIPseq

value: Anti-FOXA1 antibody - ChIP Grade (ab23738)


title: Mf_9w-ivc_MS236_216

value: 9 weeks


title: sc_B41: BY4741, WT, 16-hour

tag: strain

value: WT


title: ChFP.IND3

tag: cell line


title: Mf_6w-ivc_MS231_121

value: Germ cell


title: sc_B40: BY4741, WT, 16-hour

tag: time


title: ChFP.IND2

value: induced


title: sc_B39: BY4741, WT, 16-hour

value: 16 hour


title: Mf_6w-ivc_MS236_207

tag: developmental stage


title: Mf_3w-ivc_MS238_209

source_name: An oocyte from 3wks-ivc rOvary

tag: xenotransplantation period

value: 3 weeks


title: ChFP.IND1

tag: cell type

value: ChFP


title: sc_B38: BY4741, WT, 16-hour


title: Mf_3w-ivc_MS236_205

tag: in vitro culture period


title: sc_B37: BY4741, WT, 16-hour


title: T47D_LTED1m_Rep2_RNAseq

growth_protocol: T47D were maintained at 37°C with 5% CO2 according to ATCC recommendations. Cells were culture in  estrogen deprived conditions for 1 month. Cell culture media: IMEM with no phenol red (Corning 10-026-CV supplemented with 10% charcoal stripped FBS(Gemini 100-119) and Penicillyn/Streptomycin (Thermo Fisher Scientific 15140-12, 10000U/mL)

extract_protocol:
Total RNA was isolated from frozen cell pellets using the TRIzol reagent (Thermo Fisher Scientific cat# 15596018) according to manufacturer's instructions. RNA quantity and quality was measured on a nanodrop.2 ug of RNA were treated using the TURBO DNA-free Kit (Thermo Fisher AM1907) to remove DNA contamination and later on DNAse and divalent cations from the sample. 1ug of the treated RNA was used for library preparation. Quality was checked using a Bioanalyzer
Library preparation was perfromed on Dnase-treated RNA using the Illumina TruSeq Stranded Total RNA Library Prep Gold (Illumina 20020598). TruSeq RNA UD Indexes (Illumina 20022371) were used for the adapter ligation step. Samples were normalized to a concentration of ranging from 100-250ng for input, and 1 ul of ERCC spike-in Mix 1 (Thermo Fisher 4456740) diluted 1:200 was added to the samples.

value: Parental estrogen deprived for 1 month


title: Mf_3w-ivc_MS238_208


title: Mf_3w-ivc_MS263_204


title: T47D_LTED3m_Rep1_RNAseq

growth_protocol: T47D were maintained at 37°C with 5% CO2 according to ATCC recommendations. Cells were culture in  estrogen deprived conditions for 3 months. Cell culture media: IMEM with no phenol red (Corning 10-026-CV supplemented with 10% charcoal stripped FBS(Gemini 100-119) and Penicillyn/Streptomycin (Thermo Fisher Scientific 15140-12, 10000U/mL)

extract_protocol:
Total RNA was isolated from frozen cell pellets using the TRIzol reagent (Thermo Fisher Scientific cat# 15596018) according to manufacturer's instructions. RNA quantity and quality was measured on a nanodrop. 2 ug of RNA were treated using the TURBO DNA-free Kit (Thermo Fisher AM1907) to remove DNA contamination and later on DNAse and divalent cations from the sample. 1ug of the treated RNA was used for library preparation. Quality was checked using a Bioanalyzer
Library preparation was perfromed on Dnase-treated RNA using the Illumina TruSeq Stranded Total RNA Library Prep Gold (Illumina 20020598). TruSeq RNA UD Indexes (Illumina 20022371) were used for the adapter ligation step. Samples were normalized to a concentration of ranging from 100-250ng for input, and 1 ul of ERCC spike-in Mix 1 (Thermo Fisher 4456740) diluted 1:200 was added to the samples.

value: Parental estrogen deprived for 3 months


title: WT-Caki1, rep3

city: Seoul

first: Sunghyouk

last: Park

state: Not US or Canada

address: Bldg 143, Room 302 College of Pharmacy Seoul National University, Gwanak 1

department: Natural Products

country: South Korea

institute: Seoul National University

data_processing:
Data quality was checked using FastQC v0.11.7 (http://www.bioinformatics.babraham.ac.uk/projects/fastqc/).
Low quality ends with <Q30 were trimmed and reads shorter than 36 bases after trimming were discarded.
Further adapter trimming was performed using Trimmomatic 0.38.
Raw sequencing data were mapped to the human reference genome (GRCh37.73) with HISAT2 version 2.1.0 (https://ccb.jhu.edu/software/hisat2/index.shtml).
The raw counts of genes were normalized by Deseq2 package (R ver. 4.1.1) and further analyzed to generate the differentially expressed genes (DEGs).
Genome_build: human reference genome (GRCh37.73)
Supplementary_files_format_and_content: results_GPx8KO_vs_WT_caki1.csv (Normalized raw count)
Supplementary_files_format_and_content: Caki_1_GPx8_KO_200721.csv (Unnormalized raw count)

source_name: Clear cell renal cell carcinoma (ccRCC) cell

extract_protocol:
Total RNA was extracted using the RNA extraction kit (Intron, Easy-spin).
RNA libraries were prepared for sequencing using standard Illumina protocols
RNA-Seq, paired-end reads

value: Wild type


title: T47D_LTED4m_Rep1_RNAseq

growth_protocol: T47D were maintained at 37°C with 5% CO2 according to ATCC recommendations. Cells were culture in  estrogen deprived conditions for 4 months. Cell culture media: IMEM with no phenol red (Corning 10-026-CV supplemented with 10% charcoal stripped FBS(Gemini 100-119) and Penicillyn/Streptomycin (Thermo Fisher Scientific 15140-12, 10000U/mL)

extract_protocol:
Total RNA was isolated from frozen cell pellets using the TRIzol reagent (Thermo Fisher Scientific cat# 15596018) according to manufacturer's instructions. RNA quantity and quality was measured on a nanodrop.   2 ug of RNA were treated using the TURBO DNA-free Kit (Thermo Fisher AM1907) to remove DNA contamination and later on DNAse and divalent cations from the sample. 1ug of the treated RNA was used for library preparation. Quality was checked using a Bioanalyzer
Library preparation was perfromed on Dnase-treated RNA using the Illumina TruSeq Stranded Total RNA Library Prep Gold (Illumina 20020598). TruSeq RNA UD Indexes (Illumina 20022371) were used for the adapter ligation step. Samples were normalized to a concentration of ranging from 100-250ng for input, and 1 ul of ERCC spike-in Mix 1 (Thermo Fisher 4456740) diluted 1:200 was added to the samples.

value: Parental estrogen deprived for 4 months


title: GPX8-KO-Caki1, rep1

value: GPX8 knockout

value: Clear cell renal cell carcinoma (ccRCC) cell


title: GPX8-KO-Caki1, rep2


title: T47D_LTED4m_Rep2_RNAseq

growth_protocol: T47D were maintained at 37°C with 5% CO2 according to ATCC recommendations. Cells were culture in  estrogen deprived conditions for 4 months. Cell culture media: IMEM with no phenol red (Corning 10-026-CV supplemented with 10% charcoal stripped FBS(Gemini 100-119) and (Thermo Fisher Scientific 15140-12, 10000U/mL

value: Mammary gland ductal carcarcinoma epithelial cells derived from metastatic site

value: ATCC #HTB-133


title: T47D_LTED3m_Rep2_RNAseq


title: T47D_LTED1m_Rep1_RNAseq

growth_protocol: T47D were maintained at 37°C with 5% CO2 according to ATCC recommendations. Cells were culture in  estrogen deprived conditions for 1 month. Cell culture media: IMEM with no phenol red (Corning 10-026-CV supplemented with 10% charcoal stripped FBS (Gemini 100-119) and Penicillyn/Streptomycin (Thermo Fisher Scientific 15140-12, 10000U/mL)


title: T47D_LTED3d_Rep1_RNAseq

growth_protocol: T47D were maintained at 37°C with 5% CO2 according to ATCC recommendations. Cells were culture in estrogen deprived conditions for 3 days. Cell culture media: IMEM with no phenol red (Corning 10-026-CV supplemented with 10% charcoal stripped FBS(Gemini 100-119) and Penicillyn/Streptomycin(Thermo Fisher Scientific 15140-12, 10000U/mL)

value: Parental estrogen deprived for 3 days


title: sc_B35: BY4741, WT, 16-hour

growth_protocol: BY4742 WT were grown on SD solid agar plates.

extract_protocol:
Yeast cells were lysed with ~4µl lysis buffer, containing 0.5%Triton, 2.5µM oligo-dT, 2.5mM dNTP (Invitrogen,1959189), 8000 molecules of external RNA control consortium (ERCC) spike in, 3 x 10-2U/µl zymolyase(ZYMO, E1004-A), 1U/µl Recombinant RNase Inhibitor(TaKaRa, AI41189A), and RNA released.
RNA libraries for RNA-seq were prepared using Smart-seq2 following manufacturer's protocols.


title: Mf_6w-ivc_MS231_017

value: 6 weeks


title: sc_B34: BY4741, WT, 16-hour