title: Sample1_3_S51

data_processing:
STAR 2.7.5a alignment
htseq-count 0.6.1 tabulation
edgeR
Assembly: mm10
Supplementary files format and content: count data

source_name: Itaconate, RAW264.7, 24 hours

growth_protocol: RAW264.7 were exposed to Itaconate for 24 hrs and then gene expression profiling was performed using 3 prime RNA sequencing

extract_protocol:
RNA was extracted using TRIZOL
Lexogen QuantSeq 3'mRNAseq Library Prep FWD

treatment_protocol: RAW264.7 were exposed to Itaconate for 24 hrs.

value: Macrophages

value: Itaconate


title: MO_MCC_026.1, bulk RNA

status: Public on Jul 24 2024

type: SRA

city: Chicago

first: Jaehyuk

last: Choi

state: Illinois

address: 303 E. Superior St. 5-220

department: Dermatology

country: USA

institute: Northwestern University

library_source: transcriptomic

data_processing: Sequence reads were trimmed and filtered using Trim Galore! (v0.6.7) to remove low-quality reads, adapter sequences, and bases with low Phred scores. Reads were then aligned to the reference human genome (GRCh38.p13 primary assembly ) augmented with the Merkel cell polyomavirus transcriptome, using the GENCODE v32 transcript annotation with STAR (v2.7.10b) on two-pass mode.  The dataset was quality-controlled to remove significant outliers. Differential gene expression analysis was performed using DESeq2 (v1.28.1) or other appropriate software. The limma package was used to fit differential expression to a generalized linear model. Pathway analysis was conducted with GSVA and GSEA. ClusterProfiler4.0 was used to visualize resulting enrichment plots.

molecule: polyA RNA

organism: Homo sapiens

source_name: Merkel Cell Carcinoma (MCC)

extract_protocol:
Formalin-fixed paraffin-embedded (FFPE) tissue blocks were sectioned at a thickness of 5 μm. RNA was extracted from the FFPE tissue sections using the RNeasy FFPE Kit (Qiagen) according to the manufacturer's instructions. The extracted RNA was treated with RNase-Free DNase Set (Qiagen) to remove any genomic DNA contamination. RNA quality and quantity were assessed using a Bioanalyzer (Agilent) and Qubit fluorometer (Thermo Fisher Scientific), respectively.
RNA sequencing libraries were prepared using the TruSeq Stranded mRNA LT Sample Prep Kit (Illumina) according to the manufacturer's instructions. Briefly, 500 ng of total RNA was used for poly(A) mRNA selection, fragmentation, and cDNA synthesis. The resulting cDNA was end-repaired, adenylated, and ligated with Illumina-specific adapters. PCR amplification was then performed to enrich the adapter-ligated fragments. The final library was purified and validated for size distribution using a Bioanalyzer (Agilent) and quantified by qPCR using a Kapa Library Quantification Kit (Kapa Biosystems). Libraries were pooled at equimolar concentrations and sequenced on an Illumina NovaSeq 6000.

tag: cell type

value: Merkel Cell Carcinoma (MCC)

tag: library type

value: bulk RNA


title: MO_MCC_027, bulk RNA


title: MO_MCC_025, bulk RNA


title: Sample1_2_S50

city: Detroit

first: Nikhlesh

last: Singh

state: MI

address: 6135  Woodward Avenue

institute: Wayne State University School of Medicine

molecule: total RNA

organism: Mus musculus

tag: cell line

tag: treatment


title: Sample1_1_S49


title: MO_MCC_019, bulk RNA


title: Control_3_S48

source_name: Control, RAW264.7

value: No treatment


title: T4,tumor-PTEN

city: shanghai

first: jiaming

last: shen

address: SHANGHAI

country: China

institute: shanghai jiaotong university

data_processing:
Samples are sequenced on the platform to get image files, which are transformed by the software of the sequencing platform, and the original data in FASTQ format (Raw Data) is generated. Sequencing data contains a number of connectors, low-quality Reads, so we use fastp(0.22.0) software to filter the sequencing data to get high quality sequence (Clean Data) for further analysis
The reference genome and gene annotation files were downloaded from genome website. The filtered reads were mapping to the reference genome using HISAT2 (v2.1.0)
we used HTSeq (v0.9.1) statistics to compare the Read Count values on each gene as the original expression of the gene, and then used FPKM to standardize the exp
Supplementary files format and content: tab-delimited text file includes raw counts for each Sample

source_name: tumor

extract_protocol:
Total RNA was isolated using the Trizol Reagent (Invitrogen Life Technologies), after which the concentration, quality and integrity were determined using a NanoDrop spectrophotometer (Thermo Scientific). Three micrograms of RNA were used as input material for the RNA sample preparations. Sequencing libraries were generated according to the following steps. Firstly, mRNA was purified from total RNA using poly-T oligo-attached magnetic beads. Fragmentation was carried out using divalent cations under elevated temperature in an Illumina proprietary fragmentation buffer. First strand cDNA was synthesized using random oligonucleotides and Super Script II. Second strand cDNA synthesis was subsequently performed using DNA Polymerase I and RNase H. Remaining overhangs were converted into blunt ends via exonuclease/polymerase activities and the enzymes were removed. After adenylation of the 3′ ends of the DNA fragments, Illumina PE adapter oligonucleotides were ligated to prepare for hybridization. To select cDNA fragments of the preferred 400-500 bp in length, the library fragments were purified using the AMPure XP system (Beckman Coulter,Beverly, CA, USA). DNA fragments with ligated adaptor molecules on both ends were selectively enriched using Illumina PCR Primer Cocktail in a 15 cycle PCR reaction. Products were purified (AMPure XP system) and quantified using the Agilent high sensitivity DNA assay on a Bioanalyzer 2100 system (Agilent). The sequencing library was then sequenced on NovaSeq 6000 platform (Illumina)
RNA libraries for RNA-seg were prepared using SMARTER mRNA-Seq Library Prep Kit following manufacturer's protocols.

tag: tissue

value: tumor


title: MO_MCC_011, bulk RNA


title: MO_MCC_012, bulk RNA


title: Control_1_S46


title: T2,tumor-PTEN


title: MO_MCC_016, bulk RNA


title: WT_ZT12_Rep2

city: Planegg-Martinsried

first: Axel

last: Imhof

address: Großhadernerstr. 9

country: Germany

institute: BioMedical center

description: 10x Visium Spatial Transcriptomics

data_processing:
De-multiplexed reads were processed and mapped to the mouse genome (refdata-gex-mm10-2020-A), filtering and UMI counting was performed to obtain gene transcript counts per cell (gene barcode matrix) using Space Ranger (version 2.0.1) count function.
Assembly: mm10
Supplementary files format and content: tab separated values files and matrix files
Library strategy: Spatial transcriptomics

source_name: Brain

extract_protocol:
Fresh frozen tissues were cut into 10µm sections using a cryostat
Spatial transcriptome libraries were constructed with 10X Visium Spatial gene expression Slides and Kit according to the manufactures instructions

value: Brain

tag: strain

tag: genotype

value: wildtype


title: MO_MCC_014, bulk RNA


title: MO_MCC_010, bulk RNA


title: MO_MCC_009, bulk RNA


title: T1,tumor-PTEN


title: WT_ZT12_Rep1

description: 10x Visium Spatial Transcriptomics


title: MO_MCC_007.2, bulk RNA


title: Floxed control TIM-3 KO sample #2

city: Freiburg

first: Geoffroy

last: Andrieux

address: Breisacherstr 153

institute: University clinics Freiburg

library_source: transcriptomic single cell

data_processing:
Paired fastq files were processed with cellRanger (6.1.2) with the transcriptome index from Mouse mm10 (2020-A) downloaded from 10xgenomics website.
Assembly: mm10
Supplementary files format and content: Tab-separated values files and matrix files

source_name: spleen

extract_protocol:
Spleens were isolated on day 23 post BMT, and cells were smashed through a 75 µm cell strainer to obtain cell suspension. T cells were then isolated using pan-T cell isolation kit (Miltenyi). T cells were finally stained with H2K-b (donor haplotype) antibody (Biolegend, reference 116603) labelled with an oligotag (TotalSeq™-B0952, Biolegend, reference 405287), allowing scRNA-seq analysis of selected engrafted donor T cells. Cells were handled according to 10xGenomics Cell preparation guidelines though the whole process.
Samples were processed using the Chromium Next GEM Single Cell 3’ Reagent Kit v3.1 (Dual Index – 10x Genomics) and 3’ Feature Barcoding (10X genomis, 16 rxns PN-1000262) following manufactures protocol. In short: GEM´s were generated using a Chromium Controller (10x Genomics) at a target cell recovery rate of 10.000. GEM-RT and subsequent clean-up was performed. cDNA was amplified and 3’ Gene Expression Dual Index Library Construction was performed. Illumina paired-end, dual indexing was performed. For all amplification steps a Veriti 96 Well Thermal Cycler (appledbiosystems) and for all quality control steps a 2200 TapeStation (Agilent Technologies) was used.

treatment_protocol: Mice received lethal total body irradiation and subsequent AML cells (10 000 cells) and bone marrow transplantation (5.106 BM cells). On day 2, mice were injected with 200 000 donor T cells isolated from the spleen of donor mice.

value: spleen

value: T cells

value: TBI + BM/Tc transplantation


title: Floxed control TIM-3 KO sample #1


title: MO_MCC_007.1, bulk RNA


title: MO_MCC_004, bulk RNA


title: mESC, Flag-MBD6, Input, rep2

city: Shanghai

first: Xiaoyang

last: Dou

address: 320 Yue Yang Road

institute: CAS Center for Excellence in Molecular Cell Science, CAS

data_processing:
Low quality reads were filtered using ‘fastq_quality_filter’, and adapter were clipped using ‘fastx_clipper’, then adapter-free were collapsed to remove PCR duplicates by using ‘fastx_collapser’ and finally reads longer than 15 nt were retained for further analysis (http://hannonlab.cshl.edu/fastx_toolkit/).
Reads from rRNA were removed. The preprocessed reads were mapped to mouse genome (mm10) using bowtie (version 1.0.0) with ‘-v 3 -m 10 -k 1 –best –strata’ parameters.
Mapped reads were separated by strands with samtools (version 1.16.1) and peaks on each strand were called using MACS2 (version 2) with parameter ‘-nomodel, --keep-dup 5, -g 1.3e8, -extsize 150’ separately. Significant peaks with q < 0.01 identified by MACS2 were considered.
Assembly: mm10
Supplementary files format and content: narrowPeaks (excecp for input samples)

source_name: embryos

growth_protocol: All mESCs were kept in DMEM (Gibco, 11995065) supplemented with 15% Stem Cell Qualified Fetal Bovine Serum, Heat Inactivated (Gemini Bio Products, 100-525), 1 × L-glutamine (Gibco, 25030081), NEAA (Gibco, 25030081), LIF (MilliporeSigma, ESG1107), 1 × β-mercaptoethanol (Gibco, 21985023), 3 μM CHIR99021 (STEMCELL Technologies, 72052) and 1 μM PD0325901 (STEMCELL Technologies, 72182) at 37 °C and 5% CO2. The medium was replaced every day. ES cells were passaged on gelatin-coated plates twice to clear feeder cells before experiments.

extract_protocol:
Pellets were lysed by rotating at 4 °C for 15 minutes after passing through a 26 G needle (BD Biosciences). Embryo suspensions were sonicated on a bioruptor (Diagenode) with 30 s on/30 s off for 5 cycles. Lysates were cleared by centrifugation at 21,000 g for 15 minutes at 4 °C on a benchtop centrifuge. Supernatants were applied to MBD6 antibody (LSbio, LS-C158106-400) conjugated protein A beads (Invitrogen, 1001D) and left overnight at 4 °C on an end-to-end rotor. Beads were washed extensively with 1 ml wash buffer (50 mM HEPES pH 7.5, 300 mM KCl, 0.05% (v/v) NP-40, 1 × Halt™ Protease and Phosphatase Inhibitor Cocktail, 1 × RNaseOUT Recombinant Ribonuclease Inhibitor) at 4 °C for 5 times. Protein-RNA complex conjugated to the beads were treated by 8 U/μL RNase T1 (Thermo Scientific, EN0541) at 22 °C for 10 minutes with shaking. Input samples are digested in parallel. Then input and IP samples were separated on an SDS-PAGE gel and gel slices at corresponding size ranges were treated by proteinase K (Invitrogen, 25530049) elution. RNA was recovered with TRIZol reagent (Invitrogen, 15596026). Then T4 PNK (Thermo Scientific, EK0031) end repair was performed.
Library constructions were performed with NEBNext® Small RNA Library Prep Set for Illumina® (NEB, E7330S)

treatment_protocol: Flag-MBD5 or Flag-MBD6 stably overexpression cell lines were constructed as follows: the coding sequence for human MBD5 or MBD6 were inserted into pPB-3XFlag-Hyg vector, which is a gift from Dr. Jianlong Wang lab at Columbia University. The expression plasmid was delivered into WT mESCs by lentivirus. The resulting cell lines were kept in mESC medium supplemented with 100 mg/L Hygromycin B (Gibco, 10687010).

value: embryos

value: mESC

value: mouse embronic stem cell

value: Flag tagged MBD6

tag: antibody


title: CD8 TIM-3 KO sample #1


title: MO_MCC_003, bulk RNA


title: mESC, Flag-MBD6, CLIP, rep1

value: Flag (Abcam, #ab205606)


title: mESC, Flag-MBD5, Input, rep2

value: Flag tagged MBD5


title: Control_2_S47

value: Macrophages


title: mESC, Flag-MBD6, Input, rep1


title: mESC, Flag-MBD6, CLIP, rep2


title: NU_Pool04_RNA

description: pool 4, RNA

data_processing:
scRNA-seq Pre-processing: The GRCh38.p13 primary assembly from Ensembl Release 98 and the GENCODE v32 primary assembly were modified as follows. The sequence headers in the Ensembl FASTA were modified to match the primary assembly from GENCODE, by replacing metadata with original contig names, appending "chr" to names of autosomes and sex chromosomes, and handle the mitochondrial chromosome. The GTF file was modified to remove the version suffixes from transcript, gene, and exon IDs to match Previous CellRanger references. The GTF file was further filtered to include genes passing one or more of allowable gene_type, allowable transcript_type, no "PAR" tag, or no "readthrough_transcript" tag. Cellranger 6.1.0 was used to create a reference for scRNA-seq alignment. The MCPyV sequence was added to the primary assembly and to the final GTF file.
FASTQ files containing gene exPression and feature barcodes were aligned to the human hg38 genome and MCPyV genome using CellRanger 6.1.0. Ambient RNA was removed using the CellBender pipeline. Processed read counts for each sample were imported as a Seruat object after which TCR-seq data was added. We then filtered the data based on mitochondrial reads (25% cutoff) and minimum RNA genes (200 genes). Doublets were removed using DoubletFinder, and cells were annotated with ProjectTILS and SingleR, using Monaco and HPCA reference datasets. The data were merged into a single Seurat object. We performed standard Seurat (4.0) clustering, Harmony batch correction, and manual cell cluster annotation based on marker genes, including transcription factors, effector molecules, and Previously annotated cell types. The data were subset by major cell types and subclustered. Contaminating cells were filtered out based on marker gene exPression. Differential gene exPression, gene enrichment, and GSVA were performed, followed by comparison of cell type proportions, analysis of TCR clonotype exPression, and TCR linkage analysis.

source_name: blood

extract_protocol: CryoPreserved peripheral blood mononuclear cells (PBMCs) were thawed, labeled, and Prepared for 10x droplet encapsulation following a modified protocol adapted from 10x and BioLegend TotalSeq C protocols. The PBMCs were thawed in a 37°C water bath, and the cell number and viability were assessed using Trypan Blue staining. Cells were blocked using Human TruStain FcX™ Fc blocking reagent and stained with TotalSeq™ C antibodies and hashing antibodies. Live cells were sorted using fluorescence-activated cell sorting (FACS) with the gates set for CD45+ and CD3+ cells. The sorted cells were combined with CD3- cells to maintain a 1:3 ratio and cryoPreserved in Bambacker Preservation medium with SUPERase•In™ RNase Inhibitor. Equal volumes of five hashed samples were combined, ensuring that samples stained with the same hashing antibody were not mixed. The cells were slow-frozen at -80°C before being transferred to liquid nitrogen for long-term storage. Thawing and final sample Preparation for encapsulation were performed immediately before loading onto the 10x 5’ v2 Chromium platform. Sequencing parameters were set for 5' cDNA, a/b VDJ, g/d VDJ, and ADT libraries according to the targeted cell number for single-cell RNA sequencing.

treatment_protocol: Tissue dissociation: Human skin tumor biopsies (4mm punch) were dissociated into single-cell suspensions using the Miltenyi Biotec human tumor dissociation kit. Tissue punches were received on ice in biopsy transport media (PBS + FBS + antibiotics). Enzymes H, R, and A were reconstituted following the manufacturer's instructions and mixed with Pre-warmed RPMI 1640 medium. Biopsies were minced into 1-2mm pieces, transferred to enzyme mix, and incubated at 37°C for 30 minutes at 800 rpm in a thermomixer. Following incubation, samples were pipetted to break up clumps and filtered through a 70µm cell strainer into a 50ml conical tube. Tubes were centrifuged at 1500 rpm for 10 minutes at 4°C, and cells were resuspended in cold PBS. Cell viability and concentration were assessed using trypan blue and a cell counting slide. Cells were pelleted again and resuspended in a 1:1 mixture of FBS and FBS + 20% DMSO for cryoPreservation. Cryovials were stored at -80°C overnight before transferring to liquid nitrogen storage.

value: RNA

value: blood

value: PBMC


title: NU_Pool04_ADT

description: pool 4, ADT

value: ADT


title: NU_Pool04_gdVDJ

description: pool 4, gdVDJ

value: gd VDJ


title: NU_Pool04_abVDJ

description: pool 4, abVDJ

value: ab VDJ


title: NU_Pool03_RNA

description: pool 3, RNA


title: mESC, Flag-MBD5, Input, rep1


title: mESC, Flag-MBD5, CLIP, rep2


title: mESC, Flag-MBD5, CLIP, rep1


title: mESC, siNsun2, caRNA, rep3

data_processing:
Raw reads were trimmed with Trimmomatic (version 0.39), then aligned to mouse (mm10) or human (hg38) genome and their corresponding transcriptome using HISAT2 (version 2.2.1).
Annotation files (version M19 for mouse, and version v29 for human in gtf format) were obtained from GENCODE database (https://www.gencodegenes.org/). Reads were counted for each GENCODE annotated gene using HTSeq (version 0.12.4).
Assembly: mm10 (mouse) or hg38 (human)
Supplementary files format and content: raw counts for each Sample

growth_protocol: shCtrl or shMBD6 K-562 cells were kept in RPMI-1640 (Gibco, 61870036) with 10% fetal bovine serum (FBS, Gibco) and 1 μg/mL puromycin (Gibco, A1113803) at 37 °C and 5% CO2. All mESCs were kept in DMEM (Gibco, 11995065) supplemented with 15% Stem Cell Qualified Fetal Bovine Serum, Heat Inactivated (Gemini Bio Products, 100-525), 1 × L-glutamine (Gibco, 25030081), NEAA (Gibco, 25030081), LIF (MilliporeSigma, ESG1107), 1 × β-mercaptoethanol (Gibco, 21985023), 3 μM CHIR99021 (STEMCELL Technologies, 72052) and 1 μM PD0325901 (STEMCELL Technologies, 72182) at 37 °C and 5% CO2. The medium was replaced every day. ES cells were passaged on gelatin-coated plates twice to clear feeder cells before experiments.

extract_protocol:
Fractionation of mESCs and K-562 was performed following the published protocol with the optimized concentration of NP-40 (Sigma) for each cell line. In brief, 5-10 million cells were harvested and washed with 1 mL cold PBS/1 mM EDTA buffer, then centrifuged at 4 °C with 500 g to collect the cell pellet. 200 μL ice-cold lysis buffer (10 mM Tris-HCl, pH = 7.4, 0.05% NP-40, 150 mM NaCl) were added to the cell pellet and incubated on ice for 5 minutes, then gently pipetted up the cell lysate over 2.5 volumes of chilled sucrose cushion (24% RNase-free sucrose in lysis buffer) and centrifuged at 4 °C with 15,000 g for 10 minutes. All the supernatant was collected as cytoplasmic fraction and the nuclei pellet was washed once by gently adding 200 μL ice-cold PBS/1 mM EDTA to the nuclei pellet without dislodging the pellet. The nuclei pellet was resuspended in 200 μL prechilled glycerol buffer (20 mM Tris-HCl, pH = 7.4, 75 mM NaCl, 0.5 mM EDTA, 0.85 mM DTT, 0.125 mM PMSF, 50% glycerol) with gentle flicking of the tube. Then equal volume of cold nuclei lysis buffer (10 mM HEPES, pH = 7.6, 1 mM DTT, 7.5 mM MgCl2, 0.2 mM EDTA, 0.3 M NaCl, 1 M urea, 1% NP-40) was added, followed by vigorous vertexing for 5 seconds twice. The nuclei pellet mixtures were incubated for 2 minutes on ice, then centrifuged at 4 °C with 15,000 g for 2 minutes. The supernatant was collected as the soluble nuclear fraction (nucleoplasm). The pellet was gently rinsed with cold PBS/1 mM EDTA without dislodging and then collected as the chromosome-associated fraction.
Libraries were constructed with SMARTer® Stranded Total RNA-Seq Kit v2 - Pico Input Mammalian (TaKaRa Bio, 634411) according to the manufacturer’s instructions.

treatment_protocol: siRNA transfections in mESCs were performed according to the standard protocol of Lipofectamine™ RNAiMAX Transfection Reagent (Invitrogen, 13778150)

value: Nsun2 knockdown


title: NU_Pool03_ADT

description: pool 3, ADT


title: NU_Pool03_gdVDJ

description: pool 3, gdVDJ


title: mESC, siNsun2, caRNA, rep2


title: CD8 TIM-3 KO sample #2

value: spleen


title: NU_Pool02_RNA

description: pool 2, RNA


title: NU_Pool02_gdVDJ

description: pool 2, gdVDJ


title: mESC, siNsun2, caRNA, rep1


title: NU_Pool02_ADT

description: pool 2, ADT


title: mESC, siControl, caRNA, rep1

value: control


title: NU_Pool03_abVDJ

description: pool 3, abVDJ


title: MO_MCC_002, bulk RNA

value: Merkel Cell Carcinoma (MCC)

value: bulk RNA


title: mESC, siControl, caRNA, rep2

value: embryos

value: mESC


title: mESC, siControl, caRNA, rep3

value: mouse embronic stem cell


title: mESC, Pspc1 KO, caRNA, rep3

value: Pspc1 knockout


title: NU_Pool02_abVDJ

description: pool 2, abVDJ


title: NU_Pool01_RNA

description: pool 1, RNA


title: mESC, Pspc1 KO, caRNA, rep2


title: NU_Pool01_gdVDJ

description: pool 1, gdVDJ


title: mESC, Pspc1 KO, caRNA, rep1


title: NU_Pool01_ADT

description: pool 1, ADT


title: NU_Pool01_abVDJ

description: pool 1, ab VDJ


title: mESC, Pscp1 WT, caRNA, rep3

value: wildtype


title: PBMC10 - RNA

description: peripheral blood mononuclear cells 10


title: mESC, Pspc1 WT, caRNA, rep2


title: mESC, Pspc1 WT, caRNA, rep1


title: mESC, Tet2 KO, caRNA, rep2

value: Tet2 knockout


title: PBMC10 - gd VDJ

description: peripheral blood mononuclear cells 10


title: PBMC10 - ADT

description: peripheral blood mononuclear cells 10


title: PBMC10 - ab VDJ

description: peripheral blood mononuclear cells 10


title: mESC, Tet2 KO, caRNA, rep1


title: mESC, Tet2 WT, caRNA, rep2


title: mESC, Tet2 WT, caRNA, rep3


title: PBMC09 - RNA

description: peripheral blood mononuclear cells 9

value: RNA


title: mESC, Tet2 WT, caRNA, rep1


title: PBMC09 - gd VDJ

description: peripheral blood mononuclear cells 9

value: blood


title: PBMC08 - RNA

description: peripheral blood mononuclear cells 8

value: PBMC


title: PBMC09 - ab VDJ

description: peripheral blood mononuclear cells 9


title: PBMC09 - ADT

description: peripheral blood mononuclear cells 9

value: ADT


title: mLK, ASO IAP, totalRNA, rep3

data_processing:
Raw reads were trimmed with Trimmomatic (version 0.39), then aligned to mouse (mm10) or human (hg38) genome and their corresponding transcriptome using HISAT2 (version 2.2.1).
Annotation files (version M19 for mouse, and version v29 for human in gtf format) were obtained from GENCODE database (https://www.gencodegenes.org/).
Reads were counted for each GENCODE annotated gene using HTSeq (version 0.12.4).
Assembly: mm10 for mouse and hg38 for human
Supplementary files format and content: raw counts for each Sample

source_name: bone marrow

growth_protocol: shNC and shMBD6 K-562 cell lines were kept in RPMI-1640 (Gibco, 61870036) with 10% fetal bovine serum (FBS, Gibco) and 1 μg/mL puromycin (Gibco, A1113803) at 37 °C and 5% CO2. For hematopoietic stem and progenitor Lin-c-Kit+ (LK) cell selection, magnetic-activated cell sorting was applied with autoMACS® Pro Separator (Miltenyi Biotec). Briefly, the lineage-positive cells (Lin+) were firstly depleted from total BM cells of 6–8 weeks old mice using the Direct Lineage Cell Depletion Kit (Miltenyi Biotec, 130-110-470), and then the lineage-negative cells (Lin-) were sorted with c-Kit (CD117) MicroBeads (Miltenyi Biotec, 130-091-224). The purity of selected cells was analyzed by flow cytometry. The LK cells were also incubated in suspension culture containing 30% FBS and 2% BSA in complete RPMI-1640 medium supplemented with 100 ng/mL mSCF, 10 ng/mL mIL-3, 50 ng/mL mTPO and 10 ng/ml mGM-CSF.

extract_protocol:
Total RNA from cells were extracted by TRIzol reagent (Invitrogen, 15596026) according to the standard protocol
Libraries were constructed with SMARTer® Stranded Total RNA-Seq Kit v2 - Pico Input Mammalian (TaKaRa Bio, 634411) according to the manufacturer’s instructions.

treatment_protocol: All the ASOs and plasmids were transfected into LK cells using electroporation with P3 Primary Cell 4D-Nucleofector™ X Kit S (Lonza Bioscience, Cat#V4XP-3032) by program CV-137.

value: bone marrow

value: LK

value: IAP ASO


title: mLK, ASO NC, totalRNA, rep3

value: hematopoietic stem and progenitor Lin-c-Kit+ (LK) cell

value: control ASO


title: mLK, ASO IAP, totalRNA, rep2


title: PBMC08 - gd VDJ

description: peripheral blood mononuclear cells 8

value: gd VDJ


title: mLK, ASO IAP, totalRNA, rep1


title: PBMC08 - ADT

description: peripheral blood mononuclear cells 8


title: PBMC07 - RNA

description: peripheral blood mononuclear cells 7


title: mLK, ASO NC, totalRNA, rep1


title: PBMC07 - gd VDJ

description: peripheral blood mononuclear cells 7


title: mLK, Tet2 KO, totalRNA, rep3


title: PBMC07 - ADT

description: peripheral blood mononuclear cells 7


title: mLK, Tet2 KO, totalRNA, rep2


title: mLK, Tet2 KO, totalRNA, rep1


title: PBMC07 - ab VDJ

description: peripheral blood mononuclear cells 7

value: ab VDJ


title: PBMC06 - RNA

description: peripheral blood mononuclear cells 6


title: mLK, Tet2 WT, totalRNA, rep3


title: mLK, Tet2 WT, totalRNA, rep1


title: mLK, Tet2 WT, totalRNA, rep2


title: PBMC06 - ADT

description: peripheral blood mononuclear cells 6


title: PBMC06 - gd VDJ

description: peripheral blood mononuclear cells 6


title: mESC, Tet2 KO, nascentRNA, 40m, rep3

data_processing:
Raw reads were trimmed with Trimmomatic (version 0.39), and then aligned to mouse genome and transcriptome (mm10, version M19) as well as external RNA Control Consortium (ERCC) RNA spike-in control (Thermo Fisher Scientific) using HISAT2 (version 2.2.1).
Annotation files (version M19 for mouse) were obtained from GENCODE database (https://www.gencodegenes.org/). Reads on each GENCODE annotated gene were counted using HTSeq (version 0.12.4) .
Assembly: mm10
Supplementary files format and content: raw counts for each Sample

growth_protocol: mESCs were kept in DMEM (Gibco, 11995065) supplemented with 15% Stem Cell Qualified Fetal Bovine Serum, Heat Inactivated (Gemini Bio Products, 100-525), 1 × L-glutamine (Gibco, 25030081), NEAA (Gibco, 25030081), LIF (MilliporeSigma, ESG1107), 1 × β-mercaptoethanol (Gibco, 21985023), 3 μM CHIR99021 (STEMCELL Technologies, 72052) and 1 μM PD0325901 (STEMCELL Technologies, 72182) at 37 °C and 5% CO2. The medium was replaced every day. ES cells were passaged on gelatin-coated plates twice to clear feeder cells before experiments.

treatment_protocol: WT and Tet2-/- mESCs were seed to 6-cm dishes at the same density in three replicates. After 42 hours, cells were treated with 1 mM 5-ethynyl uridine (5-EU) for 10 minutes, 20 minutes, and 40 minutes before RNA harvest with TRIzolTM Reagent (Invitrogen, 15596026). Ribosomal RNA was depleted from total RNA preps prior to click reaction with biotin azide (PEG4 carboxamide-6-azidohexanyl biotin). Biotinylated RNA was enriched by Dynabeads™ MyOne™ Streptavidin T1 (Invitrogen, 65601). ERCC RNA Spike-In Mix (Invitrogen, 4456740) was added to eluted RNA with the amount proportional to total RNA to each sample before rRNA depletion. Spiked-RNAs were used as input for RNA-seq library constructions with SMARTer® Stranded Total RNA-Seq Kit v2 - Pico Input Mammalian (TaKaRa Bio, 634411) according to the manufacturer’s instructions. Libraries were sequenced on a NovaSeq 6000 sequencer

value: embryos

tag: time point

tag: agent

value: 1 mM 5-ethynyl uridine (5-EU)


title: PBMC06 - ab VDJ

description: peripheral blood mononuclear cells 6


title: mESC, Tet2 KO, nascentRNA, 40m, rep2

value: mESC

value: mouse embronic stem cell


title: PBMC05 - gd VDJ

description: peripheral blood mononuclear cells 5


title: PBMC05 - RNA

description: peripheral blood mononuclear cells 5


title: PBMC05 - ADT

description: peripheral blood mononuclear cells 5


title: mESC, Tet2 KO, nascentRNA, 20m, rep3


title: mLK, ASO NC, totalRNA, rep2


title: mESC, Tet2 KO, nascetnRNA, 40m, rep1


title: PBMC08 - ab VDJ

description: peripheral blood mononuclear cells 8


title: PBMC04 - RNA

description: peripheral blood mononuclear cells 4


title: mESC, Tet2 KO, nascentRNA, 20m, rep1


title: PBMC05 - ab VDJ

description: peripheral blood mononuclear cells 5


title: mESC, Tet2 KO, nascentRNA, 20m, rep2


title: PBMC04 - gd VDJ

description: peripheral blood mononuclear cells 4


title: mESC, Tet2 KO, nascentRNA, 10m, rep3


title: PBMC04 - ab VDJ

description: peripheral blood mononuclear cells 4


title: mESC, Tet2 KO, nascentRNA, 10m, rep1


title: PBMC03 - RNA

description: peripheral blood mononuclear cells 3


title: PBMC03 - gd VDJ

description: peripheral blood mononuclear cells 3


title: mESC, Tet2 WT, nascentRNA, 40m, rep2

value: wildtype


title: mESC, Tet2 WT, nascentRNA, 40m, rep1


title: mESC, Tet2 WT, nascentRNA, 20m, rep3


title: PBMC03 - ab VDJ

description: peripheral blood mononuclear cells 3


title: mESC, Tet2 WT, nascentRNA, 20m, rep2


title: mESC, Tet2 WT, nascentRNA, 20m, rep1


title: mESC, Tet2 WT, nascentRNA, 10m, rep2


title: PBMC02 - gd VDJ

description: peripheral blood mononuclear cells 2


title: PBMC01 - ADT

description: peripheral blood mononuclear cells 1


title: PBMC01 - RNA

description: peripheral blood mononuclear cells 1


title: PBMC01 - gd VDJ

description: peripheral blood mononuclear cells 1


title: mESC, Tet2 KO, nascentRNA, 10m, rep2

value: Tet2 knockout


title: mESC, WT, caRNA, m5C-IP, rep3

data_processing:
Raw reads were trimmed with Trimmomatic (version 0.39), then aligned to mouse (mm10) or human (hg38) genome and transcriptome using HISAT2 (version 2.1.0).
Annotation files (version M19 for mouse, and version v29 for human in gtf format) were downloaded from GENCODE database (https://www.gencodegenes.org/).
Mapped reads were separated by strands with samtools (version 1.16.1) and m5C peaks on each strand were called using MACS2 (version 2) with parameter ‘--nomodel, --keep-dup all, -g 7e8, -extsize 150’ separately. In this regard, the genome size was estimated based on reads coverage obtained from input samples. Significant peaks with q < 0.01 identified by MACS2 were considered.
Assembly: mm10 for mouse and hg38 for human
Supplementary files format and content: raw counts for each Sample, peaks in mESCs (except for input samples)
Library strategy: MeRIP-Seq

growth_protocol: WT K-562 cells were kept in RPMI-1640 (Gibco, 61870036) with 10% fetal bovine serum (FBS, Gibco) and 1 μg/mL puromycin (Gibco, A1113803) at 37 °C and 5% CO2. mESCs were kept in DMEM (Gibco, 11995065) supplemented with 15% Stem Cell Qualified Fetal Bovine Serum, Heat Inactivated (Gemini Bio Products, 100-525), 1 × L-glutamine (Gibco, 25030081), NEAA (Gibco, 25030081), LIF (MilliporeSigma, ESG1107), 1 × β-mercaptoethanol (Gibco, 21985023), 3 μM CHIR99021 (STEMCELL Technologies, 72052) and 1 μM PD0325901 (STEMCELL Technologies, 72182) at 37 °C and 5% CO2. The medium was replaced every day. ES cells were passaged on gelatin-coated plates twice to clear feeder cells before experiments.

extract_protocol:
Total RNAs from whole cell or the chromatin-associated fractions were randomly fragmented by incubation at 94 °C for 4 minutes using 1× fragmentation buffer (NEB E6186A). Fragmentation was stopped by adding 1× Stop Solution. Spike-in RNAs were added to each sample. Four microgram anti-m5C antibody (Diagenode, MAb-081-100) was conjugated with 30 μL of protein G beads (Invitrogen, 1003D) in 300 μL IP buffer (10 mM Tris-HCl pH 7.5, 150 mM NaCl, 0.05% Triton X-100 (v/v)) for two hours at 4 °C on a rotating wheel. The same procedure was performed for a control reaction using mouse IgGs (Abcam ab37355). Bead-antibody complexes were washed three times with IP buffer and finally brought to 250 μL with IP buffer. After heat denaturation and quick chill on ice, 10-μg samples of RNA were added to the bead-antibody complexes and incubated with 1 μL SUPERase•In™ RNase Inhibitor (Invitrogen, AM2694) overnight at 4 °C on a rotating wheel. After several washes with IP buffer, RNA was incubated in 100 μL elution buffer (5 mM Tris-HCl pH 7.5, 1 mM EDTA, 0.05% SDS, and 200 μg Proteinase K (Invitrogen 25530049)) for 1 hour at 50 °C. Beads were removed by centrifugation in a microcentrifuge, and the supernatant was purified with RCC-5 without size selection.
Libraries were constructed with SMARTer® Stranded Total RNA-Seq Kit v2 - Pico Input Mammalian (TaKaRa Bio, 634411) according to the manufacturer’s instructions.


title: mESC, WT, caRNA, Input, rep3


title: PBMC04 - ADT

description: peripheral blood mononuclear cells 4


title: NU_MCC_054, adrenal - ADT

source_name: adrenal

extract_protocol: CryoPreserved digested MCC tumor samples were thawed and processed following a modified version of the 10x Demonstrated Protocol "Thawing Dissociated Tumor Cells for Single Cell RNA Sequencing" and BioLegend TotalSeq C protocol. The protocol aimed to maintain high cell viability, decrease background, and reduce costs. Samples were thawed, labeled with TotalSeq C antibodies, and stained with 7-AAD viability staining solution. Cells were then subjected to FACS to sort CD45+ and CD45- cell populations for target enrichment of 2/3 CD45+ cells. After sorting, the final cell Preparation was resuspended in RPMI + 10% FBS + RNase Inhibitor, filtered, and counted to ensure the optimal concentration of 1e6 cells/mL. Finally, the cell suspension was processed for encapsulation and reverse transcription using the 10x 5’ v2 Chromium platform according to manufacture instructions.

value: adrenal

value: Merkel Cell Carcinoma (MCC) cell


title: mESC, Tet2 WT, nascentRNA, 40m, rep3


title: mESC, WT, caRNA, m5C-IP, rep2


title: NU_MCC_054, adrenal - gd VDJ


title: PBMC02 - ab VDJ

description: peripheral blood mononuclear cells 2

value: blood

value: PBMC


title: mESC, WT, caRNA, m5C-IP, rep1


title: mESC, Tet2 WT, nascentRNA, 10m, rep3


title: PBMC01 - ab VDJ

description: peripheral blood mononuclear cells 1


title: PBMC03 - ADT

description: peripheral blood mononuclear cells 3

value: ADT


title: PBMC02 - RNA

description: peripheral blood mononuclear cells 2

value: RNA


title: NU_MCC_054, adrenal - ab VDJ


title: mLK, Tet2 KO, H3K27me3, CUT&Tag, rep2

library_source: genomic

data_processing:
Raw reads were trimmed with Trimmomatic (version 0.39) and then mapped to mouse genome (mm10) or human genome (hg38) using bowtie2 (version 2.4.1) with default mode, where multiple alignments are searched and the best one is reported.
Peaks were called using HOMER (version 4.9) in histone mode with reads deduplicated by using ‘-tbp 1’ parameter.
Assembly: mm10 for mouse and hg38 for human
Supplementary files format and content: Peaks
Library strategy: CUT&Tag

molecule: genomic DNA

growth_protocol: K-562 cell lines were kept in RPMI-1640 (Gibco, 61870036) with 10% fetal bovine serum (FBS, Gibco) and 1 μg/mL puromycin (Gibco, A1113803) at 37 °C and 5% CO2. For hematopoietic stem and progenitor Lin-c-Kit+ (LK) cell selection, magnetic-activated cell sorting was applied with autoMACS® Pro Separator (Miltenyi Biotec). Briefly, the lineage-positive cells (Lin+) were firstly depleted from total BM cells of 6–8 weeks old mice using the Direct Lineage Cell Depletion Kit (Miltenyi Biotec, 130-110-470), and then the lineage-negative cells (Lin-) were sorted with c-Kit (CD117) MicroBeads (Miltenyi Biotec, 130-091-224). The purity of selected cells was analyzed by flow cytometry. The LK cells were also incubated in suspension culture containing 30% FBS and 2% BSA in complete RPMI-1640 medium supplemented with 100 ng/mL mSCF, 10 ng/mL mIL-3, 50 ng/mL mTPO and 10 ng/ml mGM-CSF. All mESCs were kept in DMEM (Gibco, 11995065) supplemented with 15% Stem Cell Qualified Fetal Bovine Serum, Heat Inactivated (Gemini Bio Products, 100-525), 1 × L-glutamine (Gibco, 25030081), NEAA (Gibco, 25030081), LIF (MilliporeSigma, ESG1107), 1 × β-mercaptoethanol (Gibco, 21985023), 3 μM CHIR99021 (STEMCELL Technologies, 72052) and 1 μM PD0325901 (STEMCELL Technologies, 72182) at 37 °C and 5% CO2. The medium was replaced every day. ES cells were passaged on gelatin-coated plates twice to clear feeder cells before experiments.

extract_protocol:
N/A
CUT&Tag libraries were constructed following the manufacturer's instructions of CUT&Tag-IT™ Assay Kit (Active Motif, 53160).

treatment_protocol: siRNA transfections in K-562 was performed according to the standard protocols of Lonza Walkersville SF Cell Line 4D-Nucleofector X Kit L (Lonza, V4XC2024) with program FF-120.

value: bone marrow

value: LK


title: NU_MCC_054, adrenal - RNA


title: NU_MCC_053, skin - ADT

source_name: skin

value: skin


title: mESC, WT, caRNA, Input, rep2


title: mLK, Tet2 WT, H3K27me3, CUT&Tag, rep2

value: hematopoietic stem and progenitor Lin-c-Kit+ (LK) cell


title: mLK, Tet2 KO, H3K27me3, CUT&Tag, rep1


title: PBMC02 - ADT

description: peripheral blood mononuclear cells 2


title: NU_MCC_053, skin - gd VDJ

value: gd VDJ


title: mLK, Tet2 WT, H3K27me3, CUT&Tag, rep1


title: NU_MCC_053, skin - ab VDJ

value: ab VDJ


title: NU_MCC_053, skin - RNA


title: NU_MCC_044Post1_ADT,skin


title: mLK, Tet2 WT, H2AK119ub, CUT&Tag, rep2


title: mESC, Tet2 WT, nascentRNA, 10m, rep1

value: embryos

value: 1 mM 5-ethynyl uridine (5-EU)


title: mLK, Tet2 KO, H2AK119ub, CUT&Tag, rep1


title: NU_MCC_044Post1_gdVDJ,skin


title: NU_MCC_044Post1_abVDJ,skin


title: mESC, WT, caRNA, Input, rep1

value: mESC

value: mouse embronic stem cell


title: mESC, Tet2 WT, H3K27me3, CUT&Tag, rep2

value: wildtype


title: NU_MCC_044Post2_gdVDJ,skin


title: mESC, Tet2 KO, H3K27me3, CUT&Tag, rep2

value: Tet2 knockout


title: NU_MCC_044Post2_ADT,skin

value: Merkel Cell Carcinoma (MCC) cell


title: mESC, Tet2 KO, H3K27me3, CUT&Tag, rep1


title: mESC, Tet2 WT, H3K27me3, CUT&Tag, rep1


title: NU_MCC_044Post1_RNA,skin


title: NU_MCC_044Post2_abVDJ,skin


title: mLK, Tet2 WT, H2AK119ub, CUT&Tag, rep1


title: NU_MCC_043, lymph node - gd VDJ

source_name: lymph node

value: lymph node


title: mESC, Tet2 WT, H2AK119ub, CUT&Tag, rep1


title: mESC, Tet2 KO, H2AK119ub, CUT&Tag, rep2


title: NU_MCC_043, lymph node - ab VDJ


title: mESC, Tet2 KO, H2AK119ub, CUT&Tag, rep1


title: NU_MCC_043, lymph node - RNA


title: mESC, Tet2 WT, H2AK119ub, CUT&Tag, rep2


title: mLK, Tet2 KO, H2AK119ub, CUT&Tag, rep2


title: mLK, Tet2 KO, ATAC, rep2

data_processing:
Raw reads were trimmed with Trimmomatic (version 0.39) and then mapped to mouse genome (mm10) using bowtie2 (version 2.4.1) with default mode, where multiple alignments are searched and the best one is reported.
Reads mapped to the mitochondrial genome were discard and then the remaining reads were deduplicated using a Picard tool ‘MarkDuplicates’ (version 2.26.2) (http://broadinstitute.github.io/picard/).
Assembly: mm10
Supplementary files format and content: bigWig

growth_protocol: For hematopoietic stem and progenitor Lin-c-Kit+ (LK) cell selection, magnetic-activated cell sorting was applied with autoMACS® Pro Separator (Miltenyi Biotec). Briefly, the lineage-positive cells (Lin+) were firstly depleted from total BM cells of 6–8 weeks old mice using the Direct Lineage Cell Depletion Kit (Miltenyi Biotec, 130-110-470), and then the lineage-negative cells (Lin-) were sorted with c-Kit (CD117) MicroBeads (Miltenyi Biotec, 130-091-224). The purity of selected cells was analyzed by flow cytometry. The LK cells were also incubated in suspension culture containing 30% FBS and 2% BSA in complete RPMI-1640 medium supplemented with 100 ng/mL mSCF, 10 ng/mL mIL-3, 50 ng/mL mTPO and 10 ng/ml mGM-CSF. All mESCs were kept in DMEM (Gibco, 11995065) supplemented with 15% Stem Cell Qualified Fetal Bovine Serum, Heat Inactivated (Gemini Bio Products, 100-525), 1 × L-glutamine (Gibco, 25030081), NEAA (Gibco, 25030081), LIF (MilliporeSigma, ESG1107), 1 × β-mercaptoethanol (Gibco, 21985023), 3 μM CHIR99021 (STEMCELL Technologies, 72052) and 1 μM PD0325901 (STEMCELL Technologies, 72182) at 37 °C and 5% CO2. The medium was replaced every day. ES cells were passaged on gelatin-coated plates twice to clear feeder cells before experiments.

extract_protocol: NGS libraries were obtained directly from cultured cells. Library constructions were conducted by following the manufacturer's instructions for ATAC-Seq Kit (Active Motif, 53150).


title: NU_MCC_030, lymph node - gd VDJ


title: mLK, Tet2 WT, ATAC, rep2


title: NU_MCC_030, lymph node - ab VDJ


title: NU_MCC_030, lymph node - RNA


title: mLK, Tet2 KO, ATAC, rep1


title: mESC, Pspc1 KO, ATAC, rep1

value: Pspc1 knockout


title: NU_MCC_021, adrenal - ab VDJ

value: adrenal


title: NU_MCC_021, adrenal - RNA

value: RNA


title: mLK, Tet2 WT, ATAC, rep1

value: bone marrow


title: mESC, Tet2 WT, ATAC, rep2


title: NU_MCC_021, adrenal - ADT

value: ADT


title: mESC, Pspc1 KO, ATAC, rep2


title: NU_MCC_043, lymph node - ADT


title: mESC, Tet2 WT, ATAC, rep1


title: NU_MCC_005, skin  - ab VDJ

value: skin


title: NU_MCC_005, skin  - gd VDJ


title: NU_MCC_044Post2_RNA,skin


title: Small intestinal epithelial cells Phf16-/y Sample2 IR (7 dpi)

city: Pohang

first: Somi

last: Kim

address: 47, Jigok-ro

department: Life Sciences

country: South Korea

institute: POSTECH

description:
10x Genomics
Phf16KO_7dpi_2

data_processing:
Raw FASTQ files for scRNA-seq were processed using the Cell Ranger software suite (v3.0.2; 10x Genomics). Reads were mapped to the mouse reference genome (GRCm38) with the Ensembl GRCm38.98 GTF file. For each replicate, a gene-by-cell count matrix was generated with default arguments except for expect-cells=8,000 for normal cells at basal condition, and 4,000 and 6,000 for irradiated cells at 3 dpi and 7 dpi, respectively.
Empty droplets were identified and removed using the EmptyDrops function of the Dropletutils (v1.6.1) R package with lower=100 for cells at baseline and 3 dpi, and 500 for cells at 7 dpi considering the different basal amounts of ambient RNA in samples. Poor-quality cells with < 2.5 log10-scaled unique molecular identifiers (UMIs) and < 15% of UMIs assigned to mitochondrial genes were identified and excluded using the calculateQCMetric function of the scater (v1.14.0) R package.
To remove cell-specific biases, cells were clustered using the quickCluster function of the scran (v1.12.0) R package with default arguments and cell-specific size factors were calculated using the computeSumFactors function of the same package. Raw counts of each cell were divided by cell-specific size factor and then log2-transformed with a pseudocount of 1.
Assembly: GRCm38.98
Supplementary files format and content: Tab-separated values files and matrix files

source_name: Small intestine crypt

growth_protocol: All animal studies and procedures were approved by the Institutional Animal Care and Use Committee (IACUC) of Seoul National University (SNU-211117-1). Eight-to-ten-week-old male mice were used for the experiments. Mice were housed under controlled temperature (22-23 °C) and light (12 hours light:12 hours dark) conditions. Food and water were available ad libitum.

extract_protocol:
Mice were euthanized with CO2, and the small intestine was collected and washed with DPBS (cat. no. SH30028.02, HyClone) using a 10-mL syringe. The intestinal section was cut longitudinally, with the lumen of the intestine facing up, then washed three times with DPBS. The intestine was then cut into 2-mm pieces. To dissociate the villi and crypts, the tissue pieces were incubated at 4 C in DPBS with EDTA (5 mM) for 30 min. After incubation, pieces were gently pipetted up and down five times with a pre-wetted 10-mL serological pipette. The supernatant containing villi and crypts was filtered through a 70-m filter to separate the villi (filtered) and crypts (filtrate). To isolate single cells from small intestine crypts, the pellet was further incubated in 1× TrypLE™ Express (cat. no. 12605010, Gibco/Thermo Fisher Scientific) supplemented with 0.8 kU ml−1 DNase I (cat. no. 04536282001, Roche). Crypt cells isolated from each mouse genotype were stained with propidium iodide (PI) to remove dead cells and PI negative cells were sorted using a FACSAria™ III cell sorter (BD Biosciences). Sorted cells were washed with 0.04% BSA in PBS and the concentration was adjusted to 2 x 105 ~ 1 x 106 cells/mL.
Cells were added to the RT reaction mix and then loaded onto a Chromium Single Cell B Chip (PN-1000074; 10x Genomics) to capture 6,000 cells per sample. Gel bead-in-emulsions (GEMs) were generated using a Chromium Controller (10x Genomics). Libraries for scRNA-seq were prepared using a Chromium Single Cell 3ʹ GEM Library & Gel Bead Kit v3 (PN-1000092; 10x Genomics). Subsequent DNA purification and library generation were performed according to the manufacturer’s instructions. Libraries were sequenced (paired-end 100-bp reads) on an Illumina NovaSeq 6000 (Illumina), generating an average of 80,000 read pairs per cell.

treatment_protocol: The WT and PHF16 knockout mice were irradiated with 12Gy using GC3000 Elan (MDS Nordion, Canada).

value: Small intestine crypt

value: Epithelial cells

tag: time

value: 7 dpi

value: whole body irradiation (10-16 Gy)


title: mESC, Pspc1 WT, ATAC, rep1

value: embryos


title: NU_MCC_021, adrenal - gd VDJ

value: gd VDJ


title: NU_MCC_005, skin  - ADT


title: NU_MCC_005, skin  - RNA


title: NU_MCC_030, lymph node - ADT


title: mESC, Tet2 KO, ATAC, rep2

value: mESC

value: mouse embronic stem cell


title: MO_MCC_103, skin - gd VDJ


title: Small intestinal epithelial cells WT Sample2 IR (7 dpi)

description:
10x Genomics
WT_7dpi_2

value: wild-type


title: mESC, Pspc1 WT, ATAC, rep2

value: wildtype


title: MO_MCC_103, skin  - ab VDJ

value: ab VDJ

value: Merkel Cell Carcinoma (MCC) cell


title: MO_MCC_102, skin  - gd VDJ


title: Small intestinal epithelial cells WT Sample1 IR (7 dpi)

description:
10x Genomics
WT_7dpi_1


title: MO_MCC_102, skin  - ab VDJ


title: Small intestinal epithelial cells WT Sample2 IR (3 dpi)

description:
10x Genomics
WT_3dpi_2

value: 3 dpi


title: MO_MCC_102, skin  - ADT


title: MO_MCC_103, skin  - ADT


title: Small intestinal epithelial cells Phf16-/y Sample2 IR (3 dpi)

description:
10x Genomics
Phf16KO_3dpi_2


title: MO_MCC_103, skin  - RNA

value: RNA


title: Small intestinal epithelial cells Phf16-/y Sample2 normal

description:
10x Genomics
Phf16KO_0dpi_2

value: 0 dpi


title: Small intestinal epithelial cells Phf16-/y Sample1 IR (7 dpi)

description:
10x Genomics
Phf16KO_7dpi_1


title: mESC, Tet2 KO, ATAC, rep1

value: Tet2 knockout


title: MO_MCC_102, skin  - RNA

value: skin


title: MO_MCC_101, skin  - ab VDJ


title: Small intestinal epithelial cells Phf16-/y Sample1 IR (3 dpi)

description:
10x Genomics
Phf16KO_3dpi_1

value: Small intestine crypt


title: MO_MCC_101, skin  - ADT

value: ADT


title: Small intestinal epithelial cells WT Sample1 IR (3 dpi)

description:
10x Genomics
WT_3dpi_1

value: Epithelial cells


title: Small intestinal epithelial cells Phf16-/y Sample1 normal

description:
10x Genomics
Phf16KO_0dpi_1


title: MO_MCC_101, skin  - gd VDJ


title: Periosteum and hematoma, tibia 7 days post-fracture

city: Créteil

first: Céline

last: Colnot

address: 8 rue du Général Sarrail

country: France

institute: INSERM U955

data_processing:
Fastq files were processed with the CellRanger pipeline with the default’s parameters
Reads were align against the mm10 reference genome
Filtered features barcodes matrices were used to perform data analysis
Assembly: mm10

source_name: Periosteum

extract_protocol:
Injured tibias from 4 mice were collected and the surrounded tissues were removed. The activated periosteum was scraped and collected with the hematoma/callus. Collected tissues were minced and placed 5 min in ice-cold Nuclei Buffer (NUC101, Merck) before mechanical nuclei extraction using a glass douncer. Extraction was performed by 20 strokes of pestle A followed by 10-20 of pestle B. Nuclei suspension was filtered, centrifuged and resuspended in RNAse-free PBS (AM9624, ThermoFischer Scientific) with 2% Bovine Serum Albumin (A2153, Merck) and 0.2 U/µL RNAse inhibitor (3335399001, Roche). A second step of centrifugation was performed to reduce contamination by cytoplasmic RNAs. Sytox™ AADvanced™ (S10349, ThermoFischer Scientific) was added (1/200) to label nuclei and Sytox-AAD+ nuclei were sorted using Sony SH800.
The snRNA-seq libraries were generated using Chromium Single Cell Next GEM 3′ Library & Gel Bead Kit v.3.1 (10x Genomics) according to the manufacturer’s protocol. Briefly, 10 000 to 20 000 nuclei were loaded in the 10x Chromium Controller to generate single-nuclei gel- beads in emulsion. After reverse transcription, gel-beads in emulsion were disrupted. Barcoded complementary DNA was isolated and amplified by PCR. Following fragmentation, end repair and A-tailing, sample indexes were added during index PCR. The purified libraries were sequenced on a Novaseq (Illumina) with 28 cycles of read 1, 8 cycles of i7 index and 91 cycles of read 2. Sequencing data were processed using the Cell Ranger Count pipeline and reads were mapped on the HG38 2020-A reference human genome, with intronic and exonic sequences.

value: Periosteum

value: Prss56Cre; R26tdTom; Nf1fl/fl


title: Small intestinal epithelial cells WT Sample2 normal

description:
10x Genomics
WT_0dpi_2


title: MO_MCC_101, skin  - RNA


title: MO_MCC_100, skin  - ab VDJ


title: Lesioned WM_Old_rep3

city: Munich

first: Vini

last: Tiwari

address: Feodor Lynen Straße 17

institute: Technical University Munich, (TUM), Deutsches Zentrum für Neurodegenerative Erkrankungen

description: 10x Genomics

data_processing:
The demultiplexing, barcoded processing, gene counting and aggregation were made using the Cell Ranger software v6.1.0 (https://support.10xgenomics.com/single-cell-gene-expression/software/pipelines/latest/what-is-cell-ranger)
Assembly: mm10
Supplementary files format and content: Tab-separated values files and matrix files

source_name: LesionWM, Brain

extract_protocol:
The frozen white matter tissues were homogenized in the cold Nuclei EZ Prep lysis buffer (Millipore Sigma) in a 2ml Dounce grinder, followed by clean-up using OptiPrep Density Gradient Medium (Millipore Sigma) (Corces et al., 2017). The nuclei recovered from the interface of OptiPrep medium were used for snRNA-seq preparations.
Single-nucleus RNA-seq libraries were prepared using Chromium Next Gem Single Cell 3’ v3.1 Reagents following the manufacturer’s instruction (10X Genomics).

value: LesionWM, Brain

tag: Sex

value: male

tag: age

value: 15 months old


title: MO_MCC_100, skin  - gd VDJ

value: gd VDJ


title: MO_MCC_100, skin  - RNA


title: MO_MCC_100, skin  - ADT


title: MO_MCC_099, skin  - gd VDJ


title: Lesioned WM_Young_rep3

description: 10x Genomics

value: 3 months old


title: Lesioned WM_Old_rep2

description: 10x Genomics


title: Lesioned WM_Old_rep1

description: 10x Genomics


title: Lesioned WM_Young_rep1

description: 10x Genomics


title: MO_MCC_099, skin  - ab VDJ

value: ab VDJ


title: Microglia_Input_Control_rep2

data_processing:
FastQC was used to check the quality of raw fastq files
The raw reads with a phred score > 30 were further used to align to the mm10 genome using bowtie2 using default parameters . Duplicate reads were removed using the Picard tools.
Peak calling was performed using MACS2 (create_model - - mfold (5-50) - -bw 300’) in Galaxy normalised to input . Detection of differential binding sites was done using csaw .
Bigwig files were generated from the bedgraph treatment files generated by MACS2 using the command wigtoBigWig in Galaxy. The replicates in each condition were merged using the Bigwig merge to generate a combined bigwig file for each condition.. NarrowPeak files were generated using MACS 2 .Merged and sorted narrow peak file were used as regions to plot and merged bigwig files were used as score file to compute matrix for coverage heatmap.
Assembly: mm10
Supplementary files format and content: bigWig, narrowPeak (except for Input sample)

extract_protocol:
Cells were fixed with 1% formaldehyde for 10 minutes. After quenching and cell lysis, chromatin was sheared using Bioruptor® Pico sonication device (Diagenode Cat# B01060001)
ChIP-seq libraries were prepared using IP-Star® Compact Automated System (Diagenode Cat# B03000002) from input and ChIP’d DNA using MicroPlex Library Preparation Kit v3 /96 rxns (Diagenode Cat# C05010002) with 24 UDI for MicroPlex v3 - Set I (Diagenode Cat# C05010008). 94 ng of DNA was used for ip samples. Chromatin corresponsing to 1% was set as an input. Optimal library amplification was assessed by qPCR using KAPA SYBR® FAST (Sigma-Aldrich) on Light Cycler® 96 System (Roche) and by using High Sensitivity NGS Fragment Analysis Kit (DNF-474) on a Fragment Analyzer™ (Agilent). Libraries were then purified using Agencourt® AMPure® XP (Beckman Coulter) and quantified using Qubit™ dsDNA HS Assay Kit (Thermo Fisher Scientific, Q32854). Finally, their fragment size was analyzed by High Sensitivity NGS Fragment Analysis Kit (DNF-474) on a Fragment Analyzer™ (Agilent).

treatment_protocol: 200ul of reconstituted BCG containing 8x105-12x106 CFU was injected i.v in aged mice whereas the control mice received Saline

value: Brain

value: Cd11b+ microglia

value: 12 months

value: Saline

value: Male

tag: chip antibody


title: Small intestinal epithelial cells WT Sample1 norma

description:
10x Genomics
WT_0dpi_1

value: wild-type

value: 0 dpi


title: MO_MCC_099, skin  - RNA

value: RNA

value: skin

value: Merkel Cell Carcinoma (MCC) cell


title: Lesioned WM_Young_rep2

description: 10x Genomics


title: MO_MCC_096, skin  - gd VDJ


title: MO_MCC_099, skin  - ADT

value: ADT


title: Microglia_Input_BCG_rep1

value: BCG


title: Microglia_Input_BCG_rep2


title: Microglia_H3K27ac_Control_rep2


title: MO_MCC_096, skin  - ab VDJ


title: Microglia_H3K27ac_Control_rep1


title: MO_MCC_096, skin  - RNA


title: MO_MCC_095, skin  - gd VDJ


title: Microglia_H3K27ac_BCG_rep2


title: MO_MCC_095, skin  - ab VDJ


title: Microglia_H3K27ac_BCG_rep1


title: MO_MCC_095, skin  - ADT


title: Microglia_H3K4me3_Control_rep1


title: MO_MCC_094, skin  - ab VDJ


title: Microglia_Input_Control_rep1


title: Microglia_H3K4me3_BCG_rep2


title: MO_MCC_094, skin  - gd VDJ


title: MO_MCC_094, skin  - ADT


title: MO_MCC_093, skin  - ab VDJ


title: MO_MCC_093, skin  - gd VDJ


title: Microglia_H3K4me3_BCG_rep1


title: MO_MCC_096, skin  - ADT


title: MO_MCC_093, skin  - ADT


title: MO_MCC_093, skin  - RNA


title: MO_MCC_092, skin  - gd VDJ


title: MO_MCC_092, skin  - ab VDJ


title: MO_MCC_092, skin  - ADT


title: MO_MCC_095, skin  - RNA


title: MO_MCC_092, skin  - RNA


title: Microglia_H3K4me3_Control_rep2


title: MO_MCC_094, skin  - RNA


title: MO_MCC_089, skin  - gd VDJ


title: MO_MCC_089, skin  - ADT


title: MO_MCC_089, skin  - RNA


title: MO_MCC_089, skin  - ab VDJ


title: MO_MCC_088, skin  - ab VDJ


title: MO_MCC_088, skin  - gd VDJ


title: MO_MCC_088, skin  - ADT


title: MO_MCC_088, skin  - RNA


title: MO_MCC_086, skin  - gd VDJ


title: MO_MCC_086, skin  - ab VDJ


title: MO_MCC_086, skin  - ADT


title: MO_MCC_086, skin  - RNA


title: MO_MCC_085Pre, skin  - ab VDJ


title: Old microglia_lesion WM_rep2

data_processing:
FastQC was used to check the quality of raw fastq files
ATAC-seq reads were trimmed for Nextera transposase adapters and reads <20 bp with quality cutoff 20 was done using Cutadapt.
The trimmed fastq files were aligned to the mm10 genome using bowtie2 using default parameters. The mapped reads were further filtered for mitochondrial reads, reads that are not properly paired and with low mapping quality (phred scale >=30) using BAM tools.
Peak calling was performed using MACS2 (‘- -nonmodel - - shift-100 - -extsize 200’) in Galaxy. Detection of differential binding sites was done using csaw .
Bigwig files were generated from the bedgraph treatment files generated by MACS2 using the command wigtoBigWig in Galaxy. The replicates in each condition were merged using the Bigwig merge to generate a combined bigwig file for each condition.
Assembly: mm10
Supplementary files format and content: bigWig

extract_protocol:
Cd11b+ cells were isolated using Magnetic activated cell sorting (MACS) and cryopreserved with 5% DMSO to allow slow cooling rate inorder to minimise cell lysis. Further, viable cells were prepared for library using the original procedure (Buenrostro et al., 2013) with some added modifications for frozen cells (Fujiwara et al., 2019).Briefly, cell pellets were re-suspended in ice-cold lysis buffer (10 mM Tris-Cl, pH 7.4, 10 mM NaCl, 3 mM MgCl2, and 0.1 % (v/v) Igepal CA-630) to extract the cell nuclei
The library of transposed DNA fragments was prepared according to  (Buenrostro et al., 2013).DNA was purified with the ZymoResearch DNA Clear & Concentrator TM-5 kit and eluted in 10 µl of elution buffer. qPCR was then performed to determine the additional number of PCR cycles required for the final library. The profiles of libraries were assessed with the Bioanalyzer2100.

value: LesionWM, Brain

value: male

value: 20 months old


title: MO_MCC_085Pre, skin  - gd VDJ

value: gd VDJ


title: MO_MCC_085Pre, skin  - ADT


title: Primary uveal melanoma tumor 24

type: genomic

city: Philadelphia

first: Arupa

last: Ganguly

state: PA

address: 560 Clinical Research Building 415 Curie Boulevard

institute: University of Pennsylvania

hyb_protocol: Standard protocol

scan_protocol: HiScan

data_processing: minfi

label: Cy3 and Cy5

source_name: Uveal melanoma tumor

label_protocol: Standard protocol

extract_protocol: See manuscript

value: Uveal melanoma tumor


title: MO_MCC_085Pre, skin  - RNA


title: Primary uveal melanoma tumor 19


title: Primary uveal melanoma tumor 17


title: MO_MCC_085Post, skin  - gd VDJ


title: Primary uveal melanoma tumor 18


title: Young microglia_lesion WM_rep1

value: 3 months old


title: Primary uveal melanoma tumor 14


title: Young microglia_lesion WM_rep2

value: Cd11b+ microglia


title: Batch 4; samples 12,13; sorted; non-C1 VLM cell nuclei

city: Charlottesville

first: John

middle: N

last: Campbell

state: VA

address: 485 MCCORMICK RD, GILMER HALL ROOM 229 PO BOX 400328

department: Biology

institute: University of Virginia

data_processing:
we used CellRanger v5.0 (10X Genomics) to demultiplex the sequencing reads, align them to the mouse genome mm10 including intronic reads with the command --use-introns, and quantify gene-level (feature) expression values for each cell-specific barcode as a feature-barcode matrix
Assembly: mm10
Supplementary files format and content: CellRanger output .h5 files including all cell barcodes (raw_feature_bc_matrix)

source_name: ventrolateral medulla

growth_protocol: not applicable

extract_protocol:
We then processed brain tissue into cell nuclei suspensions using one of two protocols, as described below. Importantly, tissue for all sample batches was dissected using the same approach, though the postdissection processing of tissue samples differed between batches 1-2 and batches 3-6, as described below (details provided in table 1). For sample batches 1 and 2, we transferred the dissected tissue to an RNase-free microcentrifuge tube, froze it in a slurry of dry ice and methanol, and then stored at -80 ℃ for less than a month. After thawing a batch of tissue samples on ice, we isolated cell nuclei from the thawed tissue using a previously described hypotonic lysis protocol (Matson et al., 2018), modified as follows: trituration with 5 Pasteur pipettes, starting with an unpolished pipette, followed by a series of four pipettes with tips fire polished to progressively smaller diameters; 5 to 10 strokes for each size pipette, or until the tissue passed smoothly through the pipette; and filtered resuspended nuclei with 20 µm strainer (pre-separation filter; catalog # 130-101-812; Miltenyi Biotec; Bergisch Gladbach, Germany). We fluorescently labeled nuclei with propidium iodide and then quantified them with a DeNovix CellDrop automated cell counter. a 20 µm cell strainer (Miltenyi Biotec pre-separation filter), fluorescently labeled DNA with NucBlue Live ReadyProbes Reagent (catalog # R37605, Thermo Fisher Scientific), and sorted the nuclei on a Becton Dickenson Influx cell sorter equipped with an 85 µm nozzle and set to purity mode. All sorting was performed with a Specialist in Cytometry (SCYM; ASCP) certified technical assistance at the University of Virginia Flow Cytometry Core. To separate nuclei from non-nucleated debris, we gated for events with high relative intensity of NucBlue fluorescence. We then gated the nuclei by forward scatter area vs. side scatter area to eliminate large aggregates, followed by forward scatter area vs. forward scatter height to select for singlets. Finally, from the single nuclei, we selected H2B-mCherry+ and -negative nuclei based on their high and low relative fluorescence, respectively, with 561 nm excitation and 610/20 nm collection filter. We separated H2BmCherry+ and -negative nuclei singlets into low-bind microcentrifuge tubes, each containing 18.8 uL of RT Reagent B (from Chromium Next GEM Single Cell 3ʹ v3.1 kit). For sample batches 3 through 6, which involved fluorescence-activated cell sorting (FACS), we isolated cell nuclei from preserved RVLM tissue samples based on a published protocol (Habib et al., 2016; Todd et al., 2020). RVLM tissue was microdissected as described above, but then transferred to RNase-free microcentrifuge tubes containing ice-cold RNAprotect reagent (catalog # 76104; Qiagen; Venlo, The Netherlands). We stored the tissue samples in RNAprotect at 4℃ overnight, then removed the liquid and transferred the preserved tissue samples to -80 ℃ storage for no more than 3 months. When ready to proceed, we thawed a batch of tissue samples on ice, homogenized them in 1 mL of homogenization buffer 25 times with pestle A and 25 times with pestle B (Kimble Kontes Dounce Tissue Grinder, DWK Life Sciences, catalog #885300-0001), then separated the nuclei from debris by density gradient centrifugation according to the protocol (Habib et al., 2016). We filtered the resuspended nuclei through NucBlue Live ReadyProbes Reagent (catalog # R37605, Thermo Fisher Scientific), and sorted the nuclei on a Becton Dickenson Influx cell sorter equipped with an 85 µm nozzle and set to purity mode. All sorting was performed with a Specialist in Cytometry (SCYM; ASCP) certified technical assistance at the University of Virginia Flow Cytometry Core. To separate nuclei from non-nucleated debris, we gated for events with high relative intensity of NucBlue fluorescence. We then gated the nuclei by forward scatter area vs. side scatter area to eliminate large aggregates, followed by forward scatter area vs. forward scatter height to select for singlets. Finally, from the single nuclei, we selected H2B-mCherry+ and -negative nuclei based on their high and low relative fluorescence, respectively, with 561 nm excitation and 610/20 nm collection filter. We separated H2BmCherry+ and -negative nuclei singlets into low-bind microcentrifuge tubes, each containing 18.8 uL of RT Reagent B (from Chromium Next GEM Single Cell 3ʹ v3.1 kit
We added the remainder of the Step 1 mastermix components and then proceeded with the Chromium Next GEM Single Cell 3ʹ v3.1 kit (catalog # PN-1000128 or PN-1000121; 10X Genomics; Pleasanton, CA; U.S.A.) according to the manufacturer’s instructions (CG000204 Rev D; 10X Genomics), tagging each sequencing library with a different 10X Sample Specific Index (catalog # PN-1000213; 10X Genomics) for sequencing libraries in multiplex.

treatment_protocol: To avoid stress- and anesthesia-related changes in gene expression, we rapidly decapitated unanesthetized mice upon scruffing and extracted their brains. Each sample batch consisted of tissue from 5-8 mice of each sex (8-20 weeks of age). Each brain was placed in ice-cold Earl’s Balanced Salt Solution (catalog # E2888; Sigma-Aldrich; St. Louis, MO, U.S.A.) or Hibernate-A (catalog # A1247501; Thermo Fisher Scientific; Waltham, MA, U.S.A.) for 2-5 min, transferred dorsal surface up to a chilled brain matrix (catalog # SA-2175; Roboz; Gaithersburg, MD, U.S.A.), and sectioned in the coronal plane at 1 mm intervals. Keeping the tissue chilled throughout, we microdissected RVLM tissue with a scalpel blade on an ice-filled petri dish.

value: ventrolateral medulla

value: not applicable

value: neural


title: Primary uveal melanoma tumor 23


title: Primary uveal melanoma tumor 15


title: Primary uveal melanoma tumor 22


title: Primary uveal melanoma tumor 12


title: Primary uveal melanoma tumor 20


title: Batch 4; sample 10; sorted; A5 adrenergic neuron nuclei


title: Unlesioned WM, Saline_5_5_38_rep5

data_processing:
Low-quality reads and adapters were trimmed using Trim Galore using the following parameters (reads <20 bp with quality cutoff 20).
The trimmed FASTQ files were mapped to the mm10 reference genome using STAR. The mapped reads (of lesion tissue sections) belonging to the same slide were merged using the merge BAM files tool in Galaxy.(https://usegalaxy.eu/)
To quantify the number of reads mapping to the exons of each gene, Feature counts was used. Differential analysis was performed using Deseq2.
Assembly: mm10
Supplementary files format and content: text files of count values (feature counts) for each sample analyzed

source_name: WM, Brain

extract_protocol:
RNA was isolated from PFA-fixed OCT-embedded frozen brain sections using the method that digest proteins to free cross-linked RNA (Ji et al., 2023).Five WM tissue sections were scratched from each slide and condition.
1 ng mRNA from each sample was taken and continued with the Smart-seq2 protocol as described in (Safaiyan et al., 2021) (Kaya et al., 2022).

value: WM, Brain

value: Saline

value: 12 months


title: Primary uveal melanoma tumor 5


title: Batch 3; sample 7; sorted; C1 catecholaminargic VLM neuron nuclei


title: Primary uveal melanoma tumor 7


title: MO_MCC_085Post, skin  - ADT

value: skin


title: Primary uveal melanoma tumor 8


title: Primary uveal melanoma tumor 13


title: Primary uveal melanoma tumor 16


title: Primary uveal melanoma tumor 21


title: Unlesioned WM, Saline_5_4_95_rep5


title: Primary uveal melanoma tumor 10


title: Batch 3; sample 6; sorted; A5 adrenergic neuron nuclei


title: Primary uveal melanoma tumor 2


title: MO_MCC_085Post, skin  - RNA

value: RNA

value: Merkel Cell Carcinoma (MCC) cell


title: Old microglia_lesion WM_rep1


title: Unlesioned WM, Saline_5_3_6_rep5


title: Batch 3; samples 8,9; sorted; non-C1 VLM cell nuclei


title: Batch 1; samples 1,2,3; unsorted; all VLM cell nuclei

description: Contains three technical replicates


title: Primary uveal melanoma tumor 9


title: MO_MCC_084, skin  - gd VDJ


title: Unlesioned WM, Saline_5_2_92_rep5


title: Thy1.1 5

city: Regensburg

first: LIT

last: NGS-Core

address: Franz-Josef-Strauß-Allee 11

department: NGS-Core

institute: Leibniz Institute for Immunotherapy

data_processing:
Sequenced reads were aligned to the mm10 genome assembly using Bowtie 2.2.6 and Tophat 2.1.1
R package edgeR was used to identify differentially expressed genes
Assembly: mm10
Supplementary files format and content: tab-delimited text file includes raw counts for each sample

source_name: Spleen

extract_protocol:
Total cellular RNA was isolated from CD8+ T-cells using the RNeasy Mini Kit (Qiagen) according to the manufacturer’s instructions. The concentration and quality of the purified RNA was analyzed using the RNA ScreenTape Kit (Agilent).
Generation of dsDNA libraries for Illumina sequencing from total cellular RNA was carried out using the TruSeq Stranded Total RNA Kit (Illumina) according to the manufacturer’s instructions.
The quality of dsDNA libraries was analyzed using the High Sensitivity D1000 ScreenTape Kit (Agilent) and concentrations were assessed with the Qubit dsDNA HS Kit (Thermo Fisher Scientific).

value: Spleen

value: CD8 T cells


title: Primary uveal melanoma tumor 4


title: Primary uveal melanoma tumor 11


title: Thy1.1 4


title: Primary uveal melanoma tumor 3


title: MO_MCC_084, skin  - ab VDJ

value: ab VDJ


title: Thy1.1 3


title: Primary uveal melanoma tumor 6


title: Thy1.1 2


title: Unlesioned WM, Saline_4_5_8_rep4

value: male


title: Thy1.1 1


title: MO_MCC_084, skin  - ADT

value: ADT


title: Primary uveal melanoma tumor 1

value: Uveal melanoma tumor


title: Lmo4 4


title: Unlesioned WM, Saline_4_4_36_rep4


title: MO_MCC_084, skin  - RNA


title: Lmo4 3


title: Unlesioned WM, Saline_5_1_90_rep5


title: MO_MCC_082skin, skin  - gd VDJ

value: gd VDJ


title: MO_MCC_082skin, skin  - ab VDJ


title: Lmo4 2


title: Unlesioned WM, Saline_4_3_42_rep4


title: MO_MCC_085Post, skin  - ab VDJ


title: FC;EA;rep4;scRNA-seq

city: Hangzhou

first: Congcong

last: Ma

state: Zhejiang

address: No. 548, binwen Road

institute: Zhejiang Chinese Medical University

description: 10x Genomics

data_processing:
Sequencing results were demultiplexed and converted to FASTQ format using Illumina bcl2fastq software (version 2.20). Sample demultiplexing, barcode processing and single-cell 3’gene counting were performed using the Cell Ranger pipeline (https://support.10xgenomics.com/single-cell-geneexpression/software/pipelines/latest/what-is-cell-ranger, version 6.1.1). A total of 98,338 single cells captured from 5 EA group and 5 CON group were processed using 10X Genomics Chromium Single Cell 3’Solution. The Cell Ranger output loaded into Seurat (version 3.1.1) was used for Dimensional reduction, clustering, and analysis of scRNA-seq data. Overall, 80,713 cells passed the quality control threshold: all genes expressed in less than 1 cell were removed; number of genes expressed per cell >500 as low cut-off; UMI counts <500; the percent of mitochondrial-DNA derived gene-expression <25%. To visualize the data, we further reduced the dimensionality of all cells using Seurat and used t-SNE to project the cells into 2D space. The steps include: 1. Using the LogNormalize method of the "Normalization" function of the Seurat software to calculate the expression value of genes; 2. Principal component analysis (PCA) was performed using the normalized expression value within all the PCs, the top 10 PCs were used to process clustering and t-SNE analysis; 3. Selecting weighted Shared Nearest Neighbor (SNN) graph-based clustering method to identify clusters. Marker genes for each cluster were identified with the Wilcoxon rank-sum test (default parameters are "bimod": Likelihood-ratio test) with default parameters via the FindAllMarkers function in Seurat. This selects marker genes which are expressed in more than 10% of the cells in a cluster with average log value (Fold Change) greater than 0.25 (default parameters: 0.26).
Assembly: CellRanger
Supplementary files format and content: barcodes.tsv      The barcode information of each cell was recorded in the file.
Supplementary files format and content: features.tsv        Gene id and symbol information contained in the gene annotation file
Supplementary files format and content: matrix.mtx         The number of reads of genes.

organism: Rattus norvegicus

source_name: Rat brain frontal cortex tissue

extract_protocol:
At the end of the intervention, the rats were anesthetized with pentobarbital (50 mg/kg) intraperitoneally, and the normal saline was perfused through the left heart. The brain was removed by decapitation, and the frontal cortex tissue was quickly stripped. A sterile RNase-free culture dish containing an appropriate amount of calcium-free and magnesium-free 1×PBS was placed on ice and the tissue was transferred to the culture dish and cut into 0.5 mm3 pieces. The tissues were then washed with 1× PBS, and the non-purpose tissues, such as blood stains and brain white matter, were removed as many as possible.
Tissues were dissociated into single cells in dissociation solution in a 37 ℃ water bath with shaking for 20 min at 100 rpm. Digestion was terminated by 1×PBS containing 10% fetal bovine serum (FBS, V/V), then pipetting of 5-10 times with a Pasteur pipette was applied. The resulting cell suspension was filtered through 70-30 μm stacked cell strainer and centrifuged at 300×g for 5 min at 4 °C. The cell pellet was resuspended in 100 μl 1×PBS (0.04% BSA) and added with 1 ml 1×red blood cell lysis buffer (MACS 130-094-183), and was incubated at room temperature or on ice for 2-10 min to lyse remaining red blood cells. After incubation, the suspension was centrifuged at 300×g  for 5 min at room temperature. The pellet was resuspended in 100 μl Dead Cell Removal MicroBeads (MACS 130-090-101) and dead cells were removed using Miltenyi® Dead Cell Removal Kit (MACS 130-090-101). Then, the cells were resuspended in 1×PBS (0.04% BSA) and centrifuged at 300×g for 3 min at 4 °C (repeat twice). The cell pellet was finally resuspended in 50 μl of 1×PBS (0.04% BSA). The overall cell viability was confirmed by trypan blue exclusion test, with viability above 85% as acceptable standard. The single cell suspensions were quantified using a haemocytometer/Countess II Automated Cell Counter and the concentration was adjusted to 700-1,200 cells/μl.
Single-cell suspensions were loaded to 10X Chromium to capture single cell according to the manufacturer’s instructions of 10X Genomics Chromium Single-Cell 3’kit (V3). The following cDNA amplification and library construction steps were performed according to the standard protocol. Libraries were sequenced on an Illumina NovaSeq 6000 sequencing system (paired-end multiplexing run, 150 bp) by LC-Bio Technology Co., Ltd., (HangZhou, China) at a minimum depth of 20,000 reads per cell.

value: Rat brain frontal cortex tissue

value: brain cell

value: WT

value: EA


title: FC;EA;rep5;scRNA-seq

description: 10x Genomics


title: FC;EA;rep3;scRNA-seq

description: 10x Genomics


title: MO_MCC_082skin, skin  - ADT

data_processing:
scRNA-seq Pre-processing: The GRCh38.p13 primary assembly from Ensembl Release 98 and the GENCODE v32 primary assembly were modified as follows. The sequence headers in the Ensembl FASTA were modified to match the primary assembly from GENCODE, by replacing metadata with original contig names, appending "chr" to names of autosomes and sex chromosomes, and handle the mitochondrial chromosome. The GTF file was modified to remove the version suffixes from transcript, gene, and exon IDs to match Previous CellRanger references. The GTF file was further filtered to include genes passing one or more of allowable gene_type, allowable transcript_type, no "PAR" tag, or no "readthrough_transcript" tag. Cellranger 6.1.0 was used to create a reference for scRNA-seq alignment. The MCPyV sequence was added to the primary assembly and to the final GTF file.
FASTQ files containing gene exPression and feature barcodes were aligned to the human hg38 genome and MCPyV genome using CellRanger 6.1.0. Ambient RNA was removed using the CellBender pipeline. Processed read counts for each sample were imported as a Seruat object after which TCR-seq data was added. We then filtered the data based on mitochondrial reads (25% cutoff) and minimum RNA genes (200 genes). Doublets were removed using DoubletFinder, and cells were annotated with ProjectTILS and SingleR, using Monaco and HPCA reference datasets. The data were merged into a single Seurat object. We performed standard Seurat (4.0) clustering, Harmony batch correction, and manual cell cluster annotation based on marker genes, including transcription factors, effector molecules, and Previously annotated cell types. The data were subset by major cell types and subclustered. Contaminating cells were filtered out based on marker gene exPression. Differential gene exPression, gene enrichment, and GSVA were performed, followed by comparison of cell type proportions, analysis of TCR clonotype exPression, and TCR linkage analysis.


title: Unlesioned WM, Saline_4_1_53_rep4

value: WM, Brain


title: Unlesioned WM, Saline_4_2_93_rep4

value: Saline


title: Unlesioned WM, Saline_3_5_3_rep3

value: 12 months


title: MO_MCC_082skin, skin  - RNA

value: skin


title: Lmo4 5


title: WM35_siAsah1_1_B_3J_n2

city: Toulouse

first: Marie

last: Tosolini

address: 2 avenue Hubert Curien

department: CRCT

institute: INSERM

data_processing:
FASTQ files were aligned using bowtie2 and Homo sapiens transcriptome reference GRCh38 (v97)
the raw count matrix was generated using  RSEM (v1.2.25)
Assembly: GRCh38 (v97)
Supplementary files format and content: Tab-delimited text file including raw count for each sample

growth_protocol: The WM35 cells were cultured in medium containing 80% (v/v) filtered MCDB153 medium (Sigma-Aldrich) complemented with 17.86 mM NaHCO3 and adjusted to pH 7.4, 20% (v/v) Leibovitz’s L-15 medium (Gibco, Thermo Fisher Scientific) supplemented with 1.68 mM CaCl2, 5 μg/mL human insulin solution (Sigma-Aldrich) and 2% Fetal Bovine Serum (FBS).

extract_protocol:
RNAs were extracted using a Qiagen RNA extraction kit (RNeasy Plus mini kit)
RNA Libraries for RNA-seq were prepared using Illumina Stranded mRNA Prep Ligation kit following manufacturer's instructions (Illumina).

treatment_protocol: Cells were seeded at Day 0, and then treated at Day 1 with or without 50 ng/mL human recombinant TNF (Peprotech). For transfection experiments with siRNAs, a solution of Opti-MEM containing RNAimax lipofectamine (Invitrogen, dilution 1/100) and 10 nM of siRNA was prepared and left for 20 min at room temperature. The cells were then seeded with this solution. siCTRL (5’GUACCGCACGUCAUUCGUAUCdTdT3’), siASAH1-1 (5'AAUCAACCUAUCCUCCUUCAGdTdT3') and siASAH1-2 (5'CACGAUUAACUGUGAAAUGUAdTdT3') were from Eurofins. siASAH1-3 was from ThermoFisher (Catalog number: AM16708 ID:14389).

value: Day 3


title: WM35_siAsah1_1_A_3J_n2


title: WM35_siAsah1_1_B_3J_n1


title: MO_MCC_082LN, lymph node - gd VDJ

value: lymph node

value: Merkel Cell Carcinoma (MCC) cell


title: WM35_siAsah1_1_A_3J_n1


title: MO_MCC_082LN, lymph node - ADT


title: FC;CON;rep4;scRNA-seq

description: 10x Genomics

value: Control


title: Batch 2; samples 4,5; unsorted; all VLM cell nuclei

description: Contains two technical replicates

value: ventrolateral medulla

value: not applicable

value: neural


title: FC;CON;rep5;scRNA-seq

description: 10x Genomics


title: FC;CON;rep3;scRNA-seq

description: 10x Genomics


title: WM35_siCtrl_3J_n1

value: siCtl


title: Unlesioned WM, Saline_3_3_35_rep3

value: male


title: Unlesioned WM, Saline_3_4_12_rep3


title: FC;EA;rep2;scRNA-seq

description: 10x Genomics

value: Rat brain frontal cortex tissue


title: Unlesioned WM, Saline_2_5_50_rep2


title: FC;EA;rep1;scRNA-seq

description: 10x Genomics

value: brain cell


title: MO_MCC_081Pre, skin  - RNA

value: RNA


title: MO_MCC_081Pre, skin  - ADT

value: ADT


title: WM35_TNF_72h_n3

value: WT

value: TNF


title: Unlesioned WM, Saline_2_4_40_rep2

value: WM, Brain

value: Saline

value: 12 months


title: Lmo4 1

value: Spleen

value: CD8 T cells


title: rDRG Glia maintained in base glia media Replicate 4

city: Boston

first: Chelsey

last: LeBlang

state: Massachusetts

address: 360 Longwood Ave

department: Cancer Biology

institute: Dana Farber Cancer Institute

data_processing:
Fastq files were generated using bcl2fastq v2.20
Reads were aligned with STAR (v2.7.3a)
Assembly: UCSC Hg38 and UCSC Rn6
Supplementary files format and content: CSV file contains raw counts aligned to human genome
Supplementary files format and content: CSV file contains raw counts aligned to rat genome

source_name: mixed primary rat dorsal root ganglion cells

growth_protocol: Primary e14.5 rat DRG glia were cultured for one week in base media. After one week, rDRG glia were either maintained in base media, treated with neuronal maturation media, or cocultured with terminally differentiating human induced neural crest precursors for one week. Human induced neural crest precursors were also plated alone and terminally differentiated one week.

extract_protocol:
RNA was collected using the RNeasy Plus Mini Kit from Qiagen acording to manufacturer's protocol.
Roche Kapa mRNA HyperPrep strand specific sample preparation kits  according to the manufacturer’s protocol on a Beckman Coulter Biomek i7.

value: Glia Base Media


title: MO_MCC_080, skin  - gd VDJ

value: gd VDJ

value: skin

value: Merkel Cell Carcinoma (MCC) cell


title: MO_MCC_082LN, lymph node - RNA

value: lymph node


title: WM35_siCtrl_3J_n2

value: siCtl

value: Day 3


title: Unlesioned WM, Saline_3_2_5_rep3

value: male


title: MO_MCC_080, skin  - ab VDJ

value: ab VDJ


title: Unlesioned WM, Saline_3_1_94_rep3


title: Unlesioned WM, Saline_2_2_56_rep2


title: rDRG Glia maintained in base glia media Replicate 3

value: mixed primary rat dorsal root ganglion cells


title: WM35_TNF_72h_n4

value: WT


title: FC;CON;rep2;scRNA-seq

description: 10x Genomics

value: Rat brain frontal cortex tissue

value: brain cell

value: Control


title: MO_MCC_082LN, lymph node - ab VDJ


title: rDRG Glia maintained in base glia media Replicate 2


title: rDRG Glia maintained in base glia media Replicate 1


title: FC;CON;rep1;scRNA-seq

description: 10x Genomics


title: Unlesioned WM, Saline_2_3_16_rep2

value: WM, Brain

value: Saline

value: 12 months


title: Unlesioned WM, Saline_1_5_55_rep1


title: MO_MCC_080, skin  - ADT

value: ADT


title: WM35_TNF_72h_n1

value: TNF


title: WM35_TNF_48h_n4


title: Unlesioned WM, Saline_2_1_1_rep2


title: WM35_TNF_48h_n3


title: WM35_TNF_72h_n2


title: WM35_TNF_48h_n2


title: MO_MCC_079Pre, skin  - gd VDJ

value: gd VDJ

value: skin


title: MO_MCC_079Pre, skin  - ab VDJ

value: Merkel Cell Carcinoma (MCC) cell


title: Unlesioned WM, Saline_1_3_89_rep1


title: rDRG Glia treated with NCATS Maturation Media Replicate 2

value: Maturation Media


title: WM35_TNF_48h_n1


title: rDRG Glia treated with NCATS Maturation Media Replicate 3


title: Unlesioned WM, Saline_1_2_45_rep1


title: Unlesioned WM, Saline_1_4_49_rep1


title: MO_MCC_079Pre, skin  - RNA

value: RNA


title: MO_MCC_079Pre, skin  - ADT


title: WM35_TNF_24h_n4


title: Unlesioned WM, Saline_1_1_73_rep1

value: male


title: SUVH6-YFParid1_IP_Rep2

first: Lei

last: Li

address: Xinjiangwancheng

institute: Fudan University

data_processing:
Following a FastQC (version 0.12.0) quality control, adapter trimming was performed using TrimGalore (version 0.4.1).
Reads were mapped to the Arabidopsis genome (TAIR10) using Bowtie2 (version 2.1.0), and subsequently PCR duplicates were removed using sambamba (version 0.6.6).
Deeptools (version 3.1.2) was used to examine correlations between the ChIP samples and generate Genome browser tracks.
Narrow peaks were called with MACS2 (version 2.1.0) using an FDR cutoff of 0.05 or P value cutoff of 0.00001.
Assembly: TAIR10
Supplementary files format and content: bigWig

organism: Arabidopsis thaliana

source_name: mature pollen

extract_protocol:
~150 µL of mature pollens were collected and resuspended in ice-cold pollen nuclei buffer (1.3 mM H3BO3, 3.6 mM CaCl2·2H2O, 0.74 mM KH2PO4, 438 mM Sucrose, 7 mM MOPS, 0.83 mM MgSO4·7H2O, adjust pH to 6.0), and then transferred to a 2 ml tube containing 2 mm BashingBead, vortex at the 49 Hz for 3 min with homogenizer. Debris was removed by filtering the suspension through a 10 µm nylon mesh. Nuclei were precipitated by centrifugation and resuspended in M1 buffer (10 mM sodium phosphate pH 7.0, 100 mM NaCl, 1 M hexylene glycol, 10 mM 2-mercaptoethanol, and protease inhibitor) containing 1% formaldehyde and fixed with 20 min, the cross-linking reaction was stopped by the addition of 125 mM glycine. Nuclei were precipitated by centrifugation and washed three times with M2 buffer (10 mM sodium phosphate pH 7.0, 100 mM NaCl, 1 M hexylene glycol, 10 mM MgCl2, 0.5% Triton X-100, 10 mM 2-mercaptoethanol, and protease inhibitor), and then further washed once with M3 buffer (10 mM sodium phosphate pH 7.0, 100 mM NaCl, 10 mM 2-mercaptoethanol, and protease inhibitor). Nuclei pellets were rinsed and resuspended in sonication buffer (10 mM Tris-HCl pH 8.0, 1 mM EDTA, 0.1% SDS, and protease inhibitor), and then sonicated with a sonicator (QSonica, Q800R3) for 30 min (20 s on/40 s off). After chromatin shearing, the debris was removed by centrifugation and the solutions containing chromatin fragments were diluted with three times the volume of ChIP dilution buffer (16.7 mM Tris-HCl pH 8.0, 167 mM NaCl, 1.2 mM EDTA, 1.1% Triton X-100, 0.01% SDS, and protease inhibitor). For H3K9me2 ChIP, after dilution, protein G magnetic beads (Thermo Fisher Scientific) were added to sheared chromatin and incubated at 4˚C for 1 hour. Pre-cleared samples were collected and incubated with anti-H3K9me2 antibodies at 4˚C overnight with rotation. After incubation, samples were mixed with 30 µL of protein G magnetic beads, incubated at 4˚C for 3 hours with rotation. For SUVH6 ChIP, after dilution, the pre-cleared samples were collected and incubated with GFP-trap beads at 4˚C overnight with rotation. The beads then washed twice with low salt buffer (20 mM Tris-HCl pH 8.0, 150 mM NaCl, 2 mM EDTA, 1% Triton X-100, and 0.1% SDS), twice with high salt buffer (20 mM Tris-HCl pH 8.0, 500 mM NaCl, 2 mM EDTA, 1% Triton X-100, and 0.1% SDS), once with LiCl buffer (10 mM Tris-HCl pH 8.0, 1 mM EDTA, 0.25 M LiCl, 1% NP-40, and 0.1% deoxycholic acid), and twice with TE buffer (10 mM Tris-HCl pH 8.0 and 1 mM EDTA). Immunoprecipitated DNA was incubated at 65 °C using 150 μL of elution buffer (1% SDS, 10 mM EDTA, 0.1 M NaHCO3) twice with shaking rigorously. After elution, the DNA-protein complex was reverse-crosslinked using 12 μL 5 M NaCl at 65 °C for 6 hours. 1 μL of Proteinase K, 10 μL of 0.5 M EDTA, 20 μL of 1 M Tris pH 6.5, 10 µg of RNaseA were added to digest the proteins and RNA. The DNA was then purified using phenol: chloroform and precipitated with sodium acetate, Glycogen and ethanol overnight at -20 °C. The precipitated DNA was collected and processed for the library construction.
ChIP-Seq libraries were prepared using the VAHTS Universal DNA Library Prep Kit for Illumina V4 (Vazyme, ND610) following the manufacturer’s instructions. CUT&Tag libraries were prepared using the Hyperactive Universal CUT&Tag Assay Kit for Illumina (Vazyme, TD903) following the manufacturer’s instructions. The libraries were sequenced with an Illumina NovaSeq 6000 or Illumina Hiseq 3000 to generate pair-end 150 bp reads. Samples were prepared from three or two independent biological replicates.

value: mature pollen


title: WM35_TNF_24h_n3


title: MO_MCC_079Post, skin  - gd VDJ


title: SUVH6-YFParid1_IP_Rep1


title: MO_MCC_078, lymph node - gd VDJ

value: lymph node


title: WM35_TNF_24h_n2

value: WT


title: MO_MCC_079Post, skin  - RNA


title: Unlesioned WM, BCG_5_4_46_rep5

value: BCG


title: WM35_TNF_24h_n1


title: SUVH6-YFParid1_input_Rep2


title: SUVH6-YFParid1_input_Rep1


title: WM35_TNF_6h_n4


title: MO_MCC_078, lymph node - ab VDJ

value: ab VDJ


title: SUVH6-YFP_IP_Rep2


title: rDRG Glia treated with NCATS Maturation Media Replicate 1

value: mixed primary rat dorsal root ganglion cells


title: WM35_TNF_6h_n3


title: MO_MCC_078, lymph node - ADT


title: Unlesioned WM, BCG_5_2_51_rep5

value: WM, Brain


title: MO_MCC_080, skin  - RNA


title: WM35_TNF_6h_n2


title: SUVH6-YFP_IP_Rep1


title: MO_MCC_078, lymph node - RNA


title: Unlesioned WM, BCG_5_1_41_rep5


title: SUVH6-YFP_input_Rep2


title: MO_MCC_077, skin  - gd VDJ


title: Unlesioned WM, BCG_4_5_7_rep4


title: WM35_TNF_6h_n1


title: Unlesioned WM, BCG_5_5_15_rep5

value: Saline


title: SUVH6-YFP_input_Rep1


title: Unlesioned WM, BCG_4_4_47_rep4

value: 12 months


title: Unlesioned WM, BCG_5_3_11_rep5


title: MO_MCC_077, skin  - ab VDJ


title: ARID1-GFP_input_Rep2


title: WM35_Ctrl_n3


title: Unlesioned WM, BCG_4_2_13_rep4


title: WM35_Ctrl_n4


title: Unlesioned WM, BCG_4_3_2_rep4


title: MO_MCC_077, skin  - ADT

value: ADT


title: WM35_Ctrl_n2


title: ARID1-GFP_input_Rep1


title: MO_MCC_076, skin  - ADT


title: Unlesioned WM, BCG_4_1_9_rep4


title: BEAS-2B cells, 5 ng/mL of TGF-β1

city: Ilsandong-gu, Goyang-si

first: Dong-Hee

last: Han

state: Gyeonggi-do

address: 32, Dongguk-ro

department: Life science

institute: Dongguk University

data_processing:
Illumina Casava1.8 software used for basecalling.
Sequenced reads were trimmed for adaptor sequence, and masked for low-complexity or low-quality sequence(using fastx_trimmer), then mapped to hsa (mirbase) whole genome using Bowtie2
Read count extraction using bedtools.
Assembly: hsa (mirbase) whole genom
Supplementary files format and content: tab-delimited text files include read count values for each Sample

source_name: Bronchus

growth_protocol: A549, BEAS-2B cells were maintained in RPMI1640 medium supplemented with 10% fetal bovine serum, 1% antibiotics, and 1% sodium pyruvate in a humidified atmosphere at 5% CO2 and 37°C.

extract_protocol:
RNA was harvested using Trizol reagent (Invitrogen). Total RNA was used for the construction of sequencing libraries.
RNA libraries were prepared for sequencing using NEBNext Multiplex Small RNA Library Prep kit (New England BioLabs, Inc., USA) according to the manufacturer’ instructions.

treatment_protocol: 24 hours after cell seeding, cells were treated with 1 and 5 ng/mL of TGF-β1 along with RPMI1640 medium containing 1% FBS, fetal bovine serum, 1% antibiotics, and 1% sodium pyruvate. TGF-β1 treatment was carried out for 48 hours.

value: Bronchus

value: Bronchial epithelial cells

value: 5 ng/mL of TGF-β1


title: arid1_H3K9me2_Rep2


title: Unlesioned WM, BCG_3_5_96_rep3


title: MO_MCC_076, skin  - RNA

first: Jaehyuk

last: Choi

address: 303 E. Superior St. 5-220

institute: Northwestern University

extract_protocol: CryoPreserved digested MCC tumor samples were thawed and processed following a modified version of the 10x Demonstrated Protocol "Thawing Dissociated Tumor Cells for Single Cell RNA Sequencing" and BioLegend TotalSeq C protocol. The protocol aimed to maintain high cell viability, decrease background, and reduce costs. Samples were thawed, labeled with TotalSeq C antibodies, and stained with 7-AAD viability staining solution. Cells were then subjected to FACS to sort CD45+ and CD45- cell populations for target enrichment of 2/3 CD45+ cells. After sorting, the final cell Preparation was resuspended in RPMI + 10% FBS + RNase Inhibitor, filtered, and counted to ensure the optimal concentration of 1e6 cells/mL. Finally, the cell suspension was processed for encapsulation and reverse transcription using the 10x 5’ v2 Chromium platform according to manufacture instructions.

value: skin

value: Merkel Cell Carcinoma (MCC) cell


title: MO_MCC_077, skin  - RNA

value: RNA


title: Unlesioned WM, BCG_3_4_4_rep3

value: male


title: A549 cells, 5 ng/mL of TGF-β1

source_name: Lung

value: Lung

value: Lung carcinoma epithelial cells


title: Unlesioned WM, BCG_3_3_34_rep3


title: WM35_Ctrl_n1

value: WT


title: MO_MCC_075, skin  - gd VDJ

value: gd VDJ


title: MO_MCC_075, skin  - ab VDJ

value: ab VDJ


title: Col-0_H3K9me2_Rep2

value: mature pollen


title: Unlesioned WM, BCG_3_2_74_rep3

value: WM, Brain

value: BCG


title: MO_MCC_075, skin  - RNA


title: MO_MCC_075, skin  - ADT


title: Unlesioned WM, BCG_3_1_48_rep3

value: 12 months


title: A549 cells, 1 ng/mL of TGF-β1

value: 1 ng/mL of TGF-β1


title: arid1_H3K9me2_Rep3


title: Unlesioned WM, BCG_2_5_10_rep2


title: arid1_input_Rep3


title: A549 cells, control


title: Unlesioned WM, BCG_2_4_14_rep2


title: arid1_H3K9me2_Rep1


title: MO_MCC_074, skin  - ab VDJ


title: Col-0_H3K9me2_Rep3


title: arid1_input_Rep1


title: Batch 6; sample 18; SpinalTRAP-positive VLM cells

value: ventrolateral medulla

value: not applicable

value: neural


title: BEAS-2B cells, 1 ng/mL of TGF-β1

value: Bronchus

value: Bronchial epithelial cells


title: Batch 6; sample 21; SpinalTRAP-negative VLM cells


title: arid1_input_Rep2


title: Unlesioned WM, BCG_2_3_33_rep2


title: BEAS-2B cells, control


title: Batch 6; sample 20; SpinalTRAP-negative VLM cells


title: Col-0_input_Rep2


title: Batch 6; sample 19; SpinalTRAP-negative VLM cells


title: MO_MCC_074, skin  - ADT

value: ADT

value: skin

value: Merkel Cell Carcinoma (MCC) cell


title: Col-0_input_Rep1


title: MO_MCC_073, skin  - ab VDJ


title: Batch 5; sample 14; SpinalTRAP-positive VLM cells


title: Unlesioned WM, BCG_1_5_91_rep1

value: male


title: Col-0_input_Rep3


title: Unlesioned WM, BCG_2_2_39_rep2


title: Bec_LPCs rep 2

first: hexin

last: Yan

address: Pujian Road

institute: Shanghai Reji Hospital

data_processing:
Raw data (raw reads) were processed using Trimmomatic. The reads containing poly-N and the low quality reads were removed to obtain the clean reads.
Then the clean reads were mapped to reference genome using hisat2.
FPKM value of each gene was calculated using cufflinks
and the read counts of each gene were obtained by htseq-count.
DEGs were identified using the DESeq (2012) R package functions estimate Size Factors and nbinomTest.
Assembly: mm10
Supplementary files format and content: *.txt: Tab-delimited text files include raw counts of each gene.

source_name: intrahepatic bile duct

growth_protocol: The intrahepatic biliary epithelial cells (BECs) were isolated from 6- to 8-week-old mice as described. After liver perfusion, hepatocytes, portal vein branches and connective tissue were removed using the dissecting microscope and immersed the pieces of bile ducts in a solution of digestive enzymes for 20 min (Celliver Biotechnology Inc.). Then small pieces of bile ducts were plated into matrigel and cultured in Transition and Expansion Medium (TEM). After 1-2 passages, BECs grew out of the pieces and formed a monolayer. Single-cell suspensions were sorted by Beckman MoFlo XDP equipped with 405 nm, 488 nm, 561 nm, and 640 nm excitation lasers.

extract_protocol:
Total RNA was isolated using RNeasy mini kit (Qiagen, Germany). Total RNA was quantified using a NanoDrop ND-2000 spectrophotometer (Thermo Fisher Scientific, Waltham, MA, USA).
The paired-end libraries were constructed using TruSeq Stranded mRNA LTSample Prep Kit (Illumina, San Diego, CA, USA) according to the manufacturer’s instructions.

treatment_protocol: Cultured in Transition and Expansion Medium (TEM).

value: intrahepatic bile duct

value: liver progenitor cells

value: WT

value: 6-8 weeks

tag: condition


title: MO_MCC_073, skin  - gd VDJ

value: gd VDJ


title: Unlesioned WM, BCG_2_1_44_rep2

value: WM, Brain

value: BCG


title: MO_MCC_074, skin  - gd VDJ


title: Col-0_H3K9me2_Rep1

value: mature pollen


title: Batch 5; sample 17; SpinalTRAP-negative VLM cells

value: ventrolateral medulla


title: MO_MCC_074, skin  - RNA

value: RNA


title: Bec_LPCs rep 1

value: cultured in Transition and Expansion Medium (TEM)


title: Unlesioned WM, BCG_1_3_37_rep1

value: 12 months


title: Batch 5; sample 15; SpinalTRAP-negative VLM cells

value: not applicable

value: neural


title: MO_MCC_073, skin  - RNA

value: skin


title: Unlesioned WM, BCG_1_1_52_rep1


title: Batch 4; sample 11; sorted; C1 catecholaminargic VLM neuron nuclei


title: MO_MCC_073, skin  - ADT

value: ADT

value: Merkel Cell Carcinoma (MCC) cell


title: Unlesioned WM, BCG_1_2_43_rep1

value: male


title: Batch 5; sample 16; SpinalTRAP-negative VLM cells