title: Day28, Dysf+ISG15hi, Replicate2

city: Columbus

first: Yu-Lin

state: OH

address: 1645 Neil Avenue

department: Department of Microbial Infection and Immunity

institute: The Ohio State University

description:
Library name: Sample7
Dysf+ISG15hi_CB21

data_processing:
PolyA selected RNA was sequenced by Genewiz (Azenta) using Illumina HiSeq 2x150 bp sequencing, single index package.
Partek Flow was used for RNA-seq analysis. In brief, pre-alignment QC/QA analysis was performed on the FASTQ files, followed by read alignment on human whole genome hg38 using STAR.
The aligned reads were quantified to the hg38 RefSeq Transcripts 98 using the Quantify to annotation model on Partek and gene counts were filtered (exclude features where maximum < 50.0).
Assembly: hg38
Supplementary files format and content: normalized count using DESeq2 normalization

source_name: Cord blood

growth_protocol: Human cord blood mononuclear cells (CBMCs) were isolated and CD8+ T cells were expanded in vitro by TCR stimulation (soluble anti-CD3) from day 0-28 (chronic).

extract_protocol:
RNA was isolated from frozen cell pellets of human CD8⁺ T cells, which were FACS-purified from day 28, using the PicoPure RNA Isolation Kit (Applied Biosystems), following the manufacturer’s instructions.
Libraries were prepared for Illumina HiSeq by Genewiz (Azenta), with PolyA selection.

treatment_protocol: CD8+ T cells under chronic stimulation were treated with TGFB1 (5 ng/ml) and/or ISG15 (50nM) starting from day 7.

value: Cord blood

value: CD8+ T cell

value: Recombinant human ISG15 (5 nM) and TGF-β1 (5 ng/ml) were added starting on day 7 and continued through day 28


title: B3: ureter ligation last for 3 days

status: Public on Aug 10 2025

type: SRA

city: Qingdao

first: wei

last: jiao

state: Shandong

address: No. 16 Jiangsu Road, Qingdao 266000, Shandong Province, China

department: urology

country: China

institute: The Affiliated Hospital of Qingdao University

description:
Library name: Sample 8
Column name in gene_expression_anno.txt:B3_FPKM

library_source: transcriptomic

data_processing:
The raw data (raw reads) in fastq format underwent initial processing using the fastp software (version 0.19.7). In this step, clean data (clean reads) were obtained by removing reads containing adapters, reads containing ploy-N (N represents reads for which base information cannot be determined), and low-quality reads from the raw data. The Q20, Q30, and GC-content of the clean data were calculated. All subsequent analyses were conducted based on the high-quality clean data. The clean data were mapped to the reference genome using Hisat2 (v2.0.4). The quantification of gene expression levels was performed as follows: the gene expression levels were estimated using the fragments per kilobase of transcript per million mapped fragments (FPKM) method.
Assembly: not provided
Supplementary files format and content: FPKM, count values for each sample

molecule: total RNA

organism: Rattus norvegicus

source_name: renal parenchyma

extract_protocol:
kidney tissues were thawed at 4 ◦C and homogenized using a grinding pestle. A 100 μL aliquot of the homogenized sample was transferred into a 2 mL centrifuge tube containing 400 μL of methanol, followed by vortexing for 1 min. The sample was then centrifuged at 10,000 g for 10 min at 4 ◦C. The supernatant was transferred into a new 2 mL centrifuge tube and concentrated until dry. The residue was reconstituted in 150 μL of 2‑chloro-L-phenylalanine (4 ppm) solution prepared in 80 % methanol water (stored at 4 ◦C), filtered through a 0.22 μm membrane, and transferred to a vial for LC-MS analysis.
The total RNA was extracted from kidney tissues using RNAiso Plus (TaKaRa, Dalian, China). The RNA quality and integrity were evaluated and high-quality mRNA from samples were used to construct libraries. The mRNA in the sample was enriched using magnetic beads with oligo (dT) and fragmented into short segments. The synthesis of the first strand cDNA utilized a random hexamer primer and M-MuLV Reverse Transcriptase (RNase H-), followed by the subsequent performance of second strand cDNA synthesis using DNA Polymerase I and RNase H. After 3′ -end repair, ligation of a poly(A)-tail to the sequencing linker, and selection of the fragment sizes, the double-stranded cDNA was purified and PCR amplified. Then, the PCR products were purified using the AMPureXP system (Beckman Coulter, Beverly, MA, USA) for sequencing.

tag: tissue

value: renal parenchyma

tag: genotype

value: unilateral ureteral obstruction (UUO) animal model

tag: treatment

value: ureter ligation last for 3 day

tag: batch

value: Day  5


title: B1: ureter ligation last for 3 days

description:
Library name: Sample 6
Column name in gene_expression_anno.txt:B1_FPKM


title: RNA-seq of Pectobacterium carotovorum PCC27 WT at mid-log phase (replicate 1)

city: Seoul

first: EUNHYE

last: GOO

address: 1, Gwanak-ro, Gwanak-gu

department: Agricultural Biotechnology

country: South Korea

institute: Seoul National University

description:
Library name: PCC27_WT_midlog_rep1
WT strain PCC27 harvested at mid-log phase; total RNA was extracted and rRNA-depleted prior to sequencing.

data_processing:
Adapter sequences and low-quality bases were trimmed using Trimmomatic v0.39.
Quality of trimmed reads was assessed using FastQC v0.11.9.
Trimmed reads were aligned to the P. carotovorum PCC27 reference genome (GCF_045277235.1) using HISAT2 v2.2.1.
Read counts per gene were calculated using HTSeq-count v0.12.4.
Differential expression analysis was performed using DESeq2 in R v4.2.1.
Assembly: GCF_045277235.1 (ASM4527723v1)
Supplementary files format and content: Processed data files include raw gene count tables in plain text format (.count) generated by HTSeq-count. Each file contains gene IDs and corresponding raw read counts.

organism: Pectobacterium carotovorum

source_name: bacterial cells

growth_protocol: Cells were grown in LB medium at 28°C with shaking at 220 rpm. Samples were collected at mid-log phase or early stationary phase, depending on the experimental condition.

extract_protocol:
Total RNA was extracted using the RNeasy Mini Kit (Qiagen) according to the manufacturer’s protocol, followed by DNase I treatment to remove genomic DNA contamination.
Ribosomal RNA was depleted using the Ribo-Zero rRNA Removal Kit (Bacteria, Illumina). Strand-specific RNA-seq libraries were constructed using the TruSeq Stranded Total RNA Library Prep Kit (Illumina) according to the manufacturer’s protocol.

treatment_protocol: No treatment was applied prior to RNA extraction.

tag: cell type

value: bacterial cells

value: wild type

value: mid-log phase


title: B2: ureter ligation last for 3 days

description:
Library name: Sample 7
Column name in gene_expression_anno.txt:B2_FPKM


title: 15_D6-3F

type: RNA

city: Paris

first: Rachel

last: Legendre

address: 28, rue du docteur Roux

department: Research and Resource Center for Scientific Informatics

country: France

institute: Institut Pasteur

hyb_protocol: After control of fragmentation using Bioanalyzer 2100, cDNA is then hybridized to GeneChip® ClariomS Human (LifeTechnologies) at 45°C for 17 hours.

scan_protocol: After overnight hybridization, chips are washed on the fluidic station FS450 following specific protocols (Affymetrix) and scanned using the GCS3000 7G

data_processing: The scanned images are then analyzed with Expression Console software (Affymetrix) to obtain raw data (cel files) and metrics for Quality Controls. The observations of some of these metrics and the study of the distribution of raw data show no outlier experiment.  RMA normalization is performed using R with Version 23 of Entrezgene CDF brain array.

label: Biotin

organism: Homo sapiens

source_name: BEAS-2B cell line, 6 day with flagellin

label_protocol: 100 ng of total RNA is reverse transcribed following the GeneChip® WT Plus Reagent Kit (Affymetrix) .  Briefly, the resulting double strand cDNA is used for in vitro transcription with T7 RNA polymerase (all these steps are included in the WT cDNA synthesis and amplification kit of Affymetrix). After purification according to Affymetrix protocol, 5.5 ug of Sens Target DNA are fragmented and biotin labelled.

growth_protocol: Human BEC (BEAS-2B; American Type Culture Collection) were maintained at 37°C and 5% CO2 in F12 medium (Invitrogen) supplemented with 10% fetal calf serum, 1% penicillin-streptomycin, and 10 mM HEPES and stimulated in F12 without antibiotics.

extract_protocol: RNA was isolated using a NucleoSpin RNA/Protein kit (Macherey Nagel, Duren, Germany).

treatment_protocol: On day 0, BEC were stimulated or not with ultrapure flagellin (InvivoGen) from Pseudomonas aeruginosa (5 μg/mL) for 48 hours. On day 2, cells were washed twice and incubated with medium, for resting period of 4 days, until day 6. BEC were lysed at D0 just before pre-stimulation by flagellin, D2 at the time of washing, and at D6 after the resting period to extract RNA .

tag: cells type

value: Bronchial epithelial cells

tag: cell line

tag: time

tag: stimulation

value: Flagellin


title: 12_D6-3

source_name: BEAS-2B cell line, 6 day without flagellin


title: 10_D6-1


title: Day28,Dysf,Replicate2

last: Chen

country: USA

description:
Library name: Sample5
Dysf_CB21

molecule: polyA RNA

value: Recombinant human TGF-β1 (5 ng/ml) was added starting on day 7 and continued through day 28


title: ChIP-seq of HexA in P. carotovorum PCC27 at early stationary phase (replicate 3)

description:
Library name: PCC27_HexA_earlystat_rep3
HexA ChIP-seq replicate 3 at early stationary phase in PCC27

library_source: genomic

data_processing:
Adapters and low-quality bases were trimmed using Trimmomatic (v0.38). Reads shorter than 36 bp after trimming were discarded.
Filtered reads were aligned to the P. carotovorum PCC27 reference genome using Bowtie (v1.1.2).
PCR duplicates were removed using Picard MarkDuplicates (v0.118).
Peaks were called using MACS2 (v2.1.1.20160309) with default parameters.
Peak annotation and TSS binding profiles were generated using ChIPseeker (v1.16.1).
Assembly: GCF_045277235.1 (ASM4527723v1)
Supplementary files format and content: Processed data include a signal intensity file in bedGraph format (PCC27_HexA_midlog_rep1.bedGraph) and a binary visualization file in bigWig format (PCC27_HexA_midlog_rep1.bw). These files represent normalized tag density or ChIP signal over the genome.

molecule: genomic DNA

growth_protocol: P. carotovorum PCC27 was grown in LB medium at 28°C with shaking until mid-log or early stationary phase, as determined by OD₆₀₀ measurements.

extract_protocol:
Formaldehyde was added to a final concentration of 1% to cross-link protein-DNA complexes. Cells were harvested, lysed, and chromatin was sonicated to 200–500 bp fragments. HexA-bound DNA was immunoprecipitated using anti-FLAG antibody.
Libraries were prepared using the TruSeq ChIP Sample Preparation Kit (Illumina) following the manufacturer’s protocol (TruSeq ChIP Sample Preparation Guide 15023092 Rev. B). Paired-end sequencing was performed on the Illumina NovaSeq 6000 platform with a read length of 101 bp.

treatment_protocol: No additional treatment was applied before cross-linking.

value: hexA mutant expressing HexA-FLAG

value: early stationary phase


title: 09_D2-3F

source_name: BEAS-2B cell line, 2 day with flagellin


title: 11_D6-2


title: A5: ureter ligation last for 1 day

description:
Library name: Sample 5
Column name in gene_expression_anno.txt:A5_FPKM

value: ureter ligation last for 1 day


title: 05_D2-2

source_name: BEAS-2B cell line, 2 day without flagellin


title: Tet2/3 DKO CD4 1st transfer BR4

city: New York

first: Tarmo

last: Äijö

address: 162 5th Avenue

department: Center for Computational Biology

institute: Flatiron Institute

description:
1st transfer
splj8_1

data_processing:
Library strategy: TCR-seq
bcl2fastq ver 2.20.0
Adapter trimming and quality filtering of the sequencing libraries was done using fastp (0.21.0) with the default parameters.
MiXCR (v4.7.0) was used to assess TCR repertoire from TCR-seq data using the  takara-mouse-rna-tcr-smarter preset (mixcr analyze takara-mouse-rna-tcr-smarter)
Genome_build: mm10
Supplementary_files_format_and_content: MiXCR produced clone count information.

organism: Mus musculus

source_name: Spleen

growth_protocol: All mice were bred and maintained under specific pathogen-free conditions at University of North Carolina (UNC) Genetic Medicine building, in a facility managed by the Division of Comparative Medicine at UNC Chapel Hill. The experimental procedures were approved by the UNC Institutional Animal Care and Use Committee.

extract_protocol:
RNA was isolated using RNA was isolated using the micro RNeasy plus kit (Qiagen) or the Rneasy plus mini kit (Qiagen) depending on the available starting material.
In order to analyze TCR repertoire, we used the SMARTer mouse TCR a/b profiling kit (Takara, cat no 634403) following the manufacturer’s instructions. We used 20 ng of total RNA  in order to synthesize cDNA for each library. The kit employs a 5'-RACE-based approach to capture complete V(D)J variable regions of TCR transcripts. The libraries were sequenced using a Miseq sequencer (Illumina) at the UNC High Throughput Sequencing Facility.  The alignment and assembly of the RNAseq reads and the exporting of the clonotypes were conducted using MiXCR.

treatment_protocol: Mice were euthanized by CO2 asphyxiation and thymi were isolated and dissociated to acquire single-cell suspensions. Total thymocytes were depleted of CD24hi cells by staining with biotinylated mouse anti-CD24 (clone:M1/69) purchased from Biolegend. Unwanted cells were depleted by binding to mouse streptavidin magnetic beads (anti-mouse Rapidspheres, cat no 19860, Stemcell Technologies) according to the manufacturer’s instructions. Enriched cells were stained with viability dye (eBioscience) to distinguish live cells from. Cells were stained with aGalactosyl-Ceramide loaded tetramer (conjugated with PE, obtained from NIH tetramer core), TCRb (clone:H57-597) conjugated with PERCP/Cy5.5 (Biolegend), anti-mouse CD4 AF488 (clone:RM4-5), CD8 (clone:53-6.7) conjugated with Brilliant Violet 650 or APC.  Live, CD24 low enriched cells CD4+, TCRb+, aGalCerCd1d tetramer-, CD8- cells were sorted in a purity higher than 98% and were used for RNA isolation. Splenocytes were lysed with ACK buffer (Life Technologies) to remove erythrocytes and were depleted for B cells using biotinylated anti-mouse CD19 (clone: 6D5), any remaining blood cells using biotinylated anti-mouse TER119/ Erythroid cells (clone: TER119). Cells were stained with CD4AF488 (clone: RM4-5), TCR PERCPCy5.5 (clone: H57-597), CD25 APC (clone: PC61), aGalactocyl-Ceramide (aGal-Cer) loaded Tetramer PE. Live CD4+, CD25-, TCR+, tetramer- cells were sorted and used in downstream applications. Expanded cells in congenic recipients were sorted after staining with CD45.1BV421 (clone: A20) and CD45.2PE (clone: 104).   In all cases, dead cells were excluded using a fixable viability dye eFluor780 (eBioscience).   The cells were sorted using an Aria Sorter (Becton Dickinson).

tag: tag

tag: cells

value: ex CD4,TCRb+, cd45.2+ cells


title: ChIP-seq of HexA in P. carotovorum PCC27 at early stationary phase (replicate 2)

description:
Library name: PCC27_HexA_earlystat_rep2
HexA ChIP-seq replicate 2 at early stationary phase in PCC27


title: A4: ureter ligation last for 1 day

description:
Library name: Sample 4
Column name in gene_expression_anno.txt:A4_FPKM


title: ChIP-seq of HexA in P. carotovorum PCC27 at early stationary phase (replicate 1)

description:
Library name: PCC27_HexA_earlystat_rep1
HexA ChIP-seq replicate 1 at early stationary phase in PCC27


title: 06_D2-3


title: Day28, Dysf+ISG15hi, Replicate1

description:
Library name: Sample4
Dysf+ISG15hi_CB12

value: Cord blood


title: Day28, ISG15hi, Replicate1

description:
Library name: Sample3
ISG15hi_CB12

value: CD8+ T cell

value: Recombinant human ISG15 (5 nM) was added starting on day 7 and continued through day 28


title: ChIP-seq of HexA in P. carotovorum PCC27 at mid-log phase (replicate 3)

description:
Library name: PCC27_HexA_midlog_rep3
HexA ChIP-seq replicate 3 at mid-log phase in PCC27


title: Day28, ISG15hi, Replicate2

description:
Library name: Sample6
ISG15hi_CB21


title: A3: ureter ligation last for 1 day

description:
Library name: Sample 3
Column name in gene_expression_anno.txt:A3_FPKM

value: renal parenchyma


title: Day28,Dysf,Replicate1

description:
Library name: Sample2
Dysf_CB12


title: 04_D2-1


title: A2: ureter ligation last for 1 day

description:
Library name: Sample 2
Column name in gene_expression_anno.txt:A2_FPKM

value: unilateral ureteral obstruction (UUO) animal model


title: ChIP-seq of HexA in P. carotovorum PCC27 at mid-log phase (replicate 2)

description:
Library name: PCC27_HexA_midlog_rep2
HexA ChIP-seq replicate 2 at mid-log phase in PCC27


title: ChIP-seq of HexA in P. carotovorum PCC27 at mid-log phase (replicate 1)

description:
Library name: PCC27_HexA_midlog_rep1
HexA ChIP-seq replicate 1 at mid-log phase in PCC27


title: A1: ureter ligation last for 1 day

description:
Library name: Sample 1
Column name in gene_expression_anno.txt:A1_FPKM


title: 03_D0-3

source_name: BEAS-2B cell line, basal state


title: Day28,ChronicTCR

description:
Library name: Sample1
ChTCR_CB24

value: CBMC-derived naïve CD8 T cells were stimulated for 28 days with soluble anti-CD3 antibody (OKT3)


title: 02_D0-2


title: Homo sapiens, plasma, Healthy Controls 4

status: Public on Aug 09 2025

city: Jinan

first: Xiaoyu

last: Ma

address: 247 Beiyuan Street

institute: The Second Hospital of Shandong University

description:
Library name: N4
Column name in gene_count.xls: N4

data_processing:
Sequence reads were trimmed for adaptor sequence/low-quality sequence using fastp v0.22.0 (the default parameters) Trimmed sequence reads were mapped to hg38 using hisat2 v2.2.0 (the default parameters). Read count was calculated and normalized by featurecount v2.0.6 (-p)
Assembly: hg38
Supplementary files format and content: Excel read count of genes for each sample

source_name: plasma

extract_protocol:
RNA extraction was performed simultaneously for all samples using the NucleoSpin RNA Plus kit (Macherey-Nagel), following the manufacturer’s protocol.
RNA sequencing was conducted by Novogene using Illumina platforms based on sequencing-by-synthesis (SBS) technology.

value: plasma


title: Homo sapiens, plasma, Patients 5

description:
Library name: P5
Column name in gene_count.xls: P5


title: Homo sapiens, plasma, Healthy Controls 5

description:
Library name: N5
Column name in gene_count.xls: N5


title: Homo sapiens, plasma, Healthy Controls 2

description:
Library name: N2
Column name in gene_count.xls: N2


title: Homo sapiens, plasma, Healthy Controls 1

description:
Library name: N1
Column name in gene_count.xls: N1


title: Homo sapiens, plasma, Patients 3

description:
Library name: P3
Column name in gene_count.xls: P3


title: Homo sapiens, plasma, Patients 4

description:
Library name: P4
Column name in gene_count.xls: P4


title: 01_D0-1


title: Homo sapiens, plasma, Patients 1

description:
Library name: P1
Column name in gene_count.xls: P1


title: Homo sapiens, plasma, Patients 2

description:
Library name: P2
Column name in gene_count.xls: P2


title: RNA-seq of Pectobacterium carotovorum PCC27 WT at mid-log phase (replicate 3)

description:
Library name: PCC27_WT_midlog_rep3
WT strain PCC27 harvested at mid-log phase; total RNA was extracted and rRNA-depleted prior to sequencing.

value: bacterial cells


title: Homo sapiens, SHsy5y cell, control

city: shenyang

first: jingxuan

last: sun

address: puhe

institute: china medical university

description:
Library name: SHsy5y
Column name in gene_count.xls: SHsy5y


title: Homo sapiens, SHsy5y cell, Experiment SHsy5y

description:
Library name: Ind
Column name in gene_count.xls: Ind


title: Homo sapiens, SHsy5y cell, Experiment SHsy5y_infection

description:
Library name: Hind
Column name in gene_count.xls: Hind


title: Leukocytes from islets of human donor C

city: St. Louis

first: Pavel

last: Zakharov

state: MO

address: 660 S Euclid Ave Box 8118

department: Pathology and Immunology

institute: Wash.Univ in St. Louis

description: Library name: Islets_Donor_C

library_source: transcriptomic single cell

data_processing:
Downstream filtering, unsupervised clustering, differential expression and plots generation were completed using R and Seurat package. Sample matrices were imported into the Seurat and combined into a Seurat object. Cells were filtered for mitochondrial reads < 5% and for number of reads per cell the thresholds were set to < 300 and not to exceed 3 standard deviations from the mead value: 300 < nCount_RNA < mean(nCount_RNA) + sd(nCount_RNA)*3, where "sd" is standard deviation, and "mean" is mean value. Filtering based on the number of features was done similarly: 300 < nFeature_RNA< mean(nCount_RNA) + sd(nFeature_RNA)*3. Transformation and normalization were performed using SCTransform (v2) to fit a negative binomial distribution and regress out mitochondrial read percentage and number of reads per cell. Principle components (PCs) were calculated, and an elbow plot generated to select the cutoff for number of significant PCs to use for downstream analysis. UMAP dimensionality reduction was then computed using the top 30 PCs. Unsupervised clustering was performed using the "FindNeighbors" and "FindClusters" functions. The differential expression between clusters was performed with "FindAllMarkers" function using "SCT" assay and "data" slot. The DEGs were identified with default Wilcoxon rank-sum method with P. adj < 0.05, log2FC cutoff of 0.25, and minimum percentage of cells expressing an individual gene (min.pct) set to 0.25.
Assembly: GRCh38
Supplementary files format and content: CellRanger output files: filtered expression matrix, features, and barcodes

source_name: pancreatic islets

extract_protocol:
Human islets were obtained from deceased unidentified individuals (such as death after accidents, etc.). Islets were non-enzymatically dispersed, incubated with FcR-blocking antibodies for 10 min, and stained with fluorescent antibodies and viability dye. Then, CD45+ cells were FACS-purified and subjected to 10X library preparation procedure.
Cells were processed using the Chromium Single Cell 3ʹ Reagent kit from 10X Genomics. A total of 3,000-8,000 cells per sample were used for GEM generation. Reverse transcription, barcoding, cDNA amplification and library preparation were performed per the Chromium 3ʹ protocol at Genome Technology Access Center at the McDonnell Genome Institute (GTAC@MGI).

value: pancreatic islets

tag: Sex

value: male

value: CD45+ cells

tag: age

tag: bmi

tag: 10x kit

value: 10X Genomics Chromium Single Cell 3' v2


title: Leukocytes from islets of human donor B

description: Library name: Islets_Donor_B

value: 10X Genomics Chromium Single Cell 3' v3


title: WTC CRISPRi (Control), Day 6, No Xav differentiation, GEX library

city: Springfield lakes

first: Enakshi

last: Sinniah

state: Queensland

address: 59 Orchard Crescent

country: Australia

institute: University of Queensland

description:
Library name: Sample_1_GEX
10X Genomics

data_processing:
The sequencing data were further processed to generate FASTQ files and the raw count matrix using the CellRanger pipeline. The multiplexed samples were demultiplexed to their original identities by mapping of sample reads to GRCh38 human reference genome and internal barcode sequences. The gene expression count matrix was loaded into Seurat with the barcode matrix added separate from RNA.
Assembly: GRCh38
Supplementary files format and content: Tab-separated values files and matrix files

source_name: Heart

extract_protocol:
Cells were harvested using 0.5% EDTA + Trypsin (1:10) solution and neutralised using RPMI with FBS (1:1). Samples relevant to this project were multiplexed as part of a larger experimental run using a mixture of internal and external barcode labelling. Cells from the experimental conditions relevant to this project were stained individually with their assigned cell-hashing antibody (TotalSeq-A anti-human cell-hashing antibodies). All samples were pooled and sorted for viability on a Beckman Coulter MoFlo Astrios EQ Cell Sorter with a ZombieDye stain (BioLegend). For each condition, 5×105 live cells were collected and 2×104 were run on the 10X Chromium instrument for single-cell capture, in accordance with the manufacturer’s protocol. Following the cell-hashing protocol, the transcriptome and antibody barcode libraries were prepared separately. The transcriptome libraries were sequenced on the Illumina NovaSeq 6000 and the cell-hashing libraries on the Illumina NextSeq 550Dx.
Library was performed according to the manufacturer's instructions (Chromium Single Cell 3’ Library, Gel Bead and Multiplex Kit). The sorted single cell suspensions were loaded onto 10x Genomics Single Cell 3’ Chips to form single cell gel beads in emulsion (GEMs).

treatment_protocol: On day -1 of differentiation, hPSCs were dissociated using 0.5% EDTA, plated into vitronectin coated plates at a density of 1.8 x 105 cells/cm2 (24 well-plate) or 2 x 104 cells/cm2 (T25 flask), and cultured overnight in mTeSR media (Stem Cell Technologies). Differentiation was induced on day 0 by first washing with PBS, then changing the culture media to RPMI (ThermoFisher, Cat.#11875119) containing 3μM CHIR99021 (Stem Cell Technologies, Cat.#72054), 500μg/mL BSA (Sigma Aldrich, Cat.#A9418), and 213μg/mL ascorbic acid (Sigma Aldrich, Cat.#A8960). After 3 days of culture, the media was replaced with RPMI containing 500μg/mL BSA, and 213μg/mL ascorbic acid. On day 5, the media was exchanged for RPMI containing 500μg/mL BSA, and 213μg/mL ascorbic acid without supplemental cytokines. Cells were cultured in this media till harvest on Day 6. The differentiation cultures were maintained at 37°C in a 5% CO2 incubator.

value: Heart

value: WTC CRISPRi GCaMP

value: human iPSCs

value: Control


title: PANC-1 HuR KO EV_4

city: Portland

first: Yifei

last: Guo

address: 2730 S Moody Ave

institute: Oregon Health & Science University

description: Library name: Sample 8

data_processing:
All alignment and abundance quantification performed with Kallisto pseudo-aligner: https://pachterlab.github.io/kallisto/about
Data filtering and quality control performed with Fastqc
Assembly: Gencode Release 24, GRCh38.p5. Built into a Kallisto transcriptome index titled gencodev24_GRCh38.p5_kalindex
Supplementary files format and content: Processed data is in tsv format with columns target_id, length, eff_length, est_counts, and transcripts per million (tpm).

source_name: pancreas

extract_protocol:
EV RNA was harvested with Izon RNA Extraction Kit. Cell RNA was harvested with Quiagen Rneasy mini kit.
EV RNA: takara SMARTSeq V4 kit + Nextera XT. Cell RNA: ABclonal mRNA-seq Lib Prep Kit for Illumina.

value: pancreas

value: PDAC

value: HuR KO


title: RP11-175E9.1-g1 CRISPRi WTC (NKX2-6-AS1 K/D), Day 6, No Xav differentiation, GEX library

description:
Library name: Sample_2_GEX
10X Genomics

value: RP11-175E9.1-g1 WTC CRISPRi GCaMP

value: Doxycycline-induced K/D


title: PANC-1 HuR KO EV_2

description: Library name: Sample 6


title: Leukocytes from islets of human donor A

description: Library name: Islets_Donor_A


title: Homo sapiens, plasma, Healthy Controls 3

description:
Library name: N3
Column name in gene_count.xls: N3


title: PANC-1 WT EV_4

description: Library name: Sample 4

value: WT


title: PANC-1 WT EV_3

description: Library name: Sample 3


title: PANC-1 HuR KO EV_3

description: Library name: Sample 7


title: PANC-1 HuR KO EV_1

description: Library name: Sample 5


title: PANC-1 WT EV_2

description: Library name: Sample 2


title: PANC-1 WT EV_1

description: Library name: Sample 1


title: Sterile Iscove MDM medium; Bulk cells; Day 14; Rep 1

city: Amsterdam

first: Felipe

last: Rojas-Rodriguez

state: Noord Holland

address: Plesmanlaan 125

department: Molecular hematology

country: Netherlands

institute: Sanquin research

description: Library name: BULK_IMDM_D14_1

data_processing:
Reads were trimmed for adaptors using cutadapt.
Reads were trimmed for low quality regions using FASTP.
Reads were mapped to using two-pass mode in STAR
Mapped reads were deduplicated using UMI tools.
Read count matrix was obtained using featureCounts.
Assembly: GRCh38 human genome (release 111).
Supplementary files format and content: raw_counts.txt (Tab delimited text file include counts for each sample).
Supplementary files format and content: normalized_counts.txt (Tab delimited text file include normalized counts for each sample).

source_name: CD34+ Hematopoietic stem cell

growth_protocol: CD34+ HSCs (from mobilized peripheral blood) were differentiated and maintained in SL-II medium supplemented with 10^4 U/mL penicillin and 10 ng/ml streptomycin or IMDM supplemented with 10% fetal calf serum (FCS), 200 mM glutamine, 10^4 U/mL penicillin and 10 ng/ml streptomycin. Cells were maintained in humidified atmosphere with 5% CO2 at 37C.

extract_protocol:
RNA was harvested from purified CD34+ derived neutrophils (CD16+ MACS or bulk). Therefore, cells were washed thrice with PBS and resuspended in RLT+ buffer before storage at -80C. Cell pellets were lysed in 600ul RNA Plus buffer supplemented with 1% mercapto-ethanol. Resuspended cells were homogenized with QiaShredder Columns and isolated with the Qiagen RNAeasy plus kit. RNA was eluted with 40ul of nuclease free water to obtain RNA concentrations between 10 and 100 ng/uL.
NEBNext Ultra II Directional RNA Library Prep Kit for Illumina was used to process samples according to protocol. rRNA was depleted from total RNA using the Qiagen fast select kit. After fragmentation of the rRNA reduced RNA, a cDNA synthesis used for ligation with sequencing adapters. A final concentration of 0.8 nM of DNA was used for sequencing.

treatment_protocol: Both cultures were supplemented with 10ng/mL interleukin 3  (IL3),  250 ng/mL Flt-3L,  10ng/mL GM-CSF, 30 ng/mL G-CSF and 500ng/mL human stem cell factor (hSCF) during the first three days. From day 3 till 17, only 30 ng/mL GM-CSF supplementation was used for differentiation towards neutrophils.

value: CD34+ Hematopoietic stem cell

value: Wild type

value: Sterile Iscove MDM medium:Unsorted

value: Day 14


title: Blood neutrophils; Rep 6

description: Library name: PMN_6

source_name: Blood derived neutrophils

value: Blood derived neutrophils


title: Sterile Iscove MDM medium; Bulk cells; Day 14; Rep 3

description: Library name: BULK_IMDM_D14_3


title: Blood neutrophils; Rep 5

description: Library name: PMN_5


title: Sterile Iscove MDM medium; Bulk cells; Day 14; Rep 2

description: Library name: BULK_IMDM_D14_2


title: Blood neutrophils; Rep 4

description: Library name: PMN_4


title: Blood neutrophils; Rep 3

description: Library name: PMN_3


title: Blood neutrophils; Rep 2

description: Library name: PMN_2


title: Stem-Line II; Day 17; CD16 positive; Rep 3

description: Library name: CD16pos_D17_SLII_3

value: Stem-Line II Medium; CD16 positive MACS fraction

value: Day 17


title: Stem-Line II; Day 17; CD16 positive; Rep 6

description: Library name: CD16pos_D17_SLII_6


title: Stem-Line II; Day 17; CD16 positive; Rep 2

description: Library name: CD16pos_D17_SLII_2


title: Stem-Line II; Day 17; CD16 positive; Rep 5

description: Library name: CD16pos_D17_SLII_5


title: Stem-Line II; Day 17; CD16 positive; Rep 4

description: Library name: CD16pos_D17_SLII_4


title: Stem-Line II; Day 17; CD16 positive; Rep 1

description: Library name: CD16pos_D17_SLII_1


title: Stem-Line II; Day 17; CD16 negative; Rep 1

description: Library name: CD16neg_D17_SLII_1

value: Stem-Line II Medium; CD16 negative MACS fraction


title: Stem-Line II; Day 17; CD16 negative; Rep 2

description: Library name: CD16neg_D17_SLII_2


title: Stem-Line II; Day 17; Bulk cells; Rep 4

description: Library name: BULK_D17_SLII_4

value: Stem-Line II Medium; Unsorted


title: Stem-Line II; Day 17; Bulk cells; Rep 2

description: Library name: BULK_D17_SLII_2


title: Stem-Line II; Day 17; Bulk cells; Rep 1

description: Library name: BULK_D17_SLII_1


title: Primary hepatocyte,shMCM7,,IL6 sitimulated 24h 3

city: Wuhan

first: RUI

last: Zhou

address: Donghu Road 185

institute: Wuhan University

data_processing:
Base calling of sequencing data was performed using bcl2fastq software (the software is officially delivered by illumina) to obtain fastq software.
Use fastp (v 0.19.7) fastq data for filtering to get cleandata. The main parameters are (-W 5-M 20-3-l 20-w 10).
Use the HiSat2 software (2.1.0) to compare cleandata to the reference genome with default parameters.
Use StringTie software (v1.3.4d) and annotate the result to calculate the data expression FPKM (Gene.FPKM.xls); Calculate ReadsCount (i.e. Gene.Count.xls) according to the script given by the official StringTie.
Assembly: GRCm39
Supplementary files format and content: tab-delimited tex file includes raw counts for each Sample
Supplementary files format and content: tab-delimited tex files include RPKM values for each Sample

source_name: Primary hepatocyte

growth_protocol: Primary hepatocytes were isolated from mice two weeks after tail vein injection with AAV-px552 and AAV-shMcm7. The isolated hepatocytes were cultured in vitro in DMEM supplemented with 10% fetal bovine serum (FBS) under standard conditions, including a humidified atmosphere with 5% CO₂ at 37°C.

extract_protocol:
RNA was harvested using Trizol(Takara). 0.5ug of total RNA was used for the construction of sequencing libraries.
RNA libraries for RNA-seq were prepared using KAPA mRNA HyperPrep Kit following manufacturer's protocols

treatment_protocol: Recombinant Mouse IL-6(PeproTech) was added at a concentration of 10 ng/ml to stimulate the cultured hepatocytes.

value: Primary hepatocyte

value: Mcm9 knockdown

value: recombinate IL-6, 24h


title: Stem-Line II; Day 14; CD16 positive; Rep 5

description: Library name: CD16pos_D14_SLII_5


title: Primary hepatocyte,shMCM7,,IL6 sitimulated 24h 1

value: Mcm7 knockdown


title: Primary hepatocyte,shMCM7,,IL6 sitimulated 24h 2

value: Mcm8 knockdown


title: Stem-Line II; Day 17; Bulk cells; Rep 3

description: Library name: BULK_D17_SLII_3


title: Stem-Line II; Day 14; CD16 positive; Rep 4

description: Library name: CD16pos_D14_SLII_4


title: Primary hepatocyte,shControl,IL6 sitimulated 24h 2

value: Control


title: Primary hepatocyte,shControl,IL6 sitimulated 24h 3


title: Stem-Line II; Day 14; CD16 positive; Rep 3

description: Library name: CD16pos_D14_SLII_3


title: Stem-Line II; Day 14; CD16 positive; Rep 2

description: Library name: CD16pos_D14_SLII_2


title: Blood neutrophils; Rep 1

description: Library name: PMN_1


title: Primary hepatocyte,shMCM7 3

value: untreatment


title: Primary hepatocyte,shControl 3


title: Primary hepatocyte,shMCM7 1


title: Stem-Line II; Day 17; CD16 negative; Rep 4

description: Library name: CD16neg_D17_SLII_4


title: Stem-Line II; Day 17; CD16 negative; Rep 3

description: Library name: CD16neg_D17_SLII_3


title: Primary hepatocyte,shControl,IL6 sitimulated 24h 1


title: Stem-Line II; Day 14; CD16 negative; Rep 3

description: Library name: CD16neg_D14_SLII_3


title: Stem-Line II; Day 14; CD16 negative; Rep 2

description: Library name: CD16neg_D14_SLII_2


title: Primary hepatocyte,shControl 1


title: Primary hepatocyte,shControl 2


title: Stem-Line II; Day 14; CD16 negative; Rep 1

description: Library name: CD16neg_D14_SLII_1


title: Stem-Line II; Day 14; Bulk cells; Rep 3

description: Library name: BULK_D14_SLII_3


title: ExT29-Tag_001

city: Noda City Chiba

first: Mikiya

last: Tsunoda

address: 2669 Yamazaki

department: Research Institute for Biomedical Sciences

country: Japan

institute: Tokyo university of science

description:
Library name: Sample 90
scRNAseqwithbulkTCRseq_immunotherapies.h5ad

library_source: other

data_processing:
For bulk TCR analysis, adapter trimming and quality filtering of sequencing data were performed by using Cutadapt-3.2 and PRINSEQ-0.20.4. Sequencing data were processed by MiXCR-3.0.5. In MiXCR, Filtered reads were aligned to reference mouse TCR V/D/J sequences registered in the international ImMunoGeneTics information system: -starting-material=rna, -5-end=no-v-primers-, -3-end=c-primers, -adapters=no-adapters, vParameters.geneFeatureToAlign=VTranscript, -vjAlignmentOrder=JThenV, then identical sequences were assembled and grouped in clones with PCR and sequencing error correlation with the following parameters: -badQualityThreshold=15, –separateByV=true, –separateByJ=true, -only-productive=true, –region-of-interest=CDR3. The Variable (V) and Joining (J) segment of TCRs were represented in IMGT gene nomenclature. List of final clones were analyzed by VDJtools-1.2.1. Then, the sequencing reads of sample was normalized to the six times of cell count in each sample by “DownSample” command of VDJtools. T-cell clones were determined as TCR reads with the same TCR V segment, J segment and CDR3 nucleotide sequence.
For single cell analysis, reads were trimmed with Cutadapt-2.1051. Filtered reads were then annotated using the Python script provided by BD Biosciences and mapped to reference RNA using Bowtie2-2.4.254. Cell barcode information of each read was then added to the bowtie2-mapped BAM files and read counts of each gene in each cell barcode were counted using mawk. Resultant count data were converted to a single-cell gene-expression matrix file and filtered for valid cells based on read counts. Expression data of targeted panel genes were converted to a Seurat object55, log normalized and scaled (regressed by the number of unique molecular identifiers per cell) using the NormalizeData and ScaleData functions respectively. Principal component analysis (PCA) was performed using RunPCA function, and enrichment of each PC was calculated using the JackStraw and ScoreJackStraw function, and PCs that were significantly enriched (P ≤ 0.05) were selected for clustering and dimensional reduction analysis. Dimensional reduction was performed using RunUMAP function with default settings. Cell clustering was performed using FindNeighbors and FindClusters (resolution = 0.6) against the significant PCs.
Assembly: IMGT reference sequences of mouse TRB, GRCh38
Supplementary files format and content: text files in VDJtools format include read count and frequency, complementarity determining region 3 (CDR3) nucleotide and amino acid sequences, Variable(V)/ Diversity(D)/ Joining(J) segment names, and V, D and J segment boundaries within CDR3 nucleotide sequence of individual clones
Supplementary files format and content: tab-delimited text files include raw tag-count values for each sample

molecule: protein

source_name: Tumor

growth_protocol: Lewis lung carcinoma (LLC) cells (5 × 10^5 cells /mouse) were inoculated subcutaneously (s.c.) into the left and right flanks of C57BL/6 mice.

extract_protocol:
Tumors were cut into small fragments and digested for 45 minutes at 37°C with 0.2% collagenase (FUJIFILM Wako) and DNase I (Sigma-Aldrich), except for experiments for single-cell analysis, in which tumors were digested with 0.2% collagenase NB8 (Nordmark Pharma GmbH). The cells were then subjected to two-phase density separation with upper 35% Percoll PLUS (GE Healthcare) (1000g, 10 minutes at room temperature). Leukocytes were recovered, washed and resuspended in preparation medium (PM) [5%FBS, 10mM HEPES (Nacalai), Penicillin-Streptomycin (Nacalai) in RPMI-1640 (Nacalai)]. After RBC lysis with ACK buffer [155mM of NH4Cl (Nacalai), 10mM of KHCO3 (Nacalai), and 0.5mM EDTA pH8.0 (NIPPON GENE)] for 1 minute at room temperature, cells were washed and resuspended in PM, and the cell concentrations were measured using Flow-Count fluorospheres (Beckman Coulter) and a CytoFLEX S flow cytometer (Beckman Coulter).Cells were stained with a mix of Fc Block (anti-mouse CD16/CD32 mAb; clone 2.4G2, BioLegend) and fluorophore-conjugated anti-mouse monoclonal antibodies (Supplementary Table 5). Antibody mixtures were prepared using BD Horizon Brilliant Stain Buffer Plus (BD) diluted 5x with FACS buffer (D-PBS(-) supplemented with 2% FBS, and 0.05% Sodium Azide). Staining was performed for 20 minutes at 4°C. In the single-cell RNA-seq/TCR-seq experiment, cells were stained with antibodies and anti-CD45 Sample Tag oligonucleotide-conjugated antibodies from the Single-Cell Multiplexing Kit (BD Biosciences). Before sorting, T cells were enriched from tumor suspensions by further staining with BD IMag APC Magnetic Particles-DM (BD Biosciences), then placed on magnetic plate. After washing by MACS buffer (D-PBS(-) supplemented with 10% BSA, and 2 mM EDTA pH8.0), APC and APC-Cy7(CD8, mouse TCRb)-positive cells were collected as positive fraction. Cell sorting was performed using a FACS Aria II or Aria III (BD Biosciences). Cells were stained with 5µg/mL of propidium iodide (PI, Sigma-Aldrich) immediately before sorting. Nonviable cells were excluded from the analysis based on forward and side scatter profiles and PI staining. For bulk TCR repertoire analysis and bulk RNA-seq analysis, cells were sorted into Lysis buffer [1% w/v LiDS, 100 mM Tris-HCl pH7.5, 500 mM LiCl, 5 mM dithiothreitol, and 10 mM EDTA pH8.0] and stored at −80°C. For single-cell RNA/TCR-seq, cells were sorted into D-PBS(-) with 10% FBS. For the bulk TCR-seq experiments, cells were sorted from the CD8+, CD8+PD-1+Ly108+TIM-3-, CD8+PD-1+Ly108+TIM-3+, CD8+PD-1+Ly108-TIM-3+ and CD8+PD-1- fractions to produce five datasets per mouse. For the single-cell RNA/TCR-seq and bulk TCR-seq experiment, cells were sorted from the CD8+PD-1+Ly108+TIM-3-, CD8+PD-1+Ly108+TIM-3+, CD8+PD-1+Ly108-TIM-3+ and CD8+PD-1- fractions.
Bulk TCR libraries were prepared on purified T cells lysed in lysis buffer [1% LiDS, 100 mM Tris-HCl (pH 7.5), 500 mM LiCl, and 10 mM EDTA]. PolyA RNAs were isolated according to a previous report with some modifications (GSE110711).To perform reverse transcription and template-swiching, beads were then suspended in 10 µL of RT mix [1× First Strand buffer (Thermo Fisher Scientific), 1 mM dNTP, 2.5 mM DTT (Thermo Fisher Scientific), 1 M betaine (Sigma-Aldrich), 9 mM MgCl2, 1 U/µL RNaseIn Plus Rnase Inhibitor (Promega), 10 U/µL Superscript II (Thermo Fisher Scientific), and 1µM of i5-TSO], and incubated for 60 min at 42ºC and immediately cooled on ice. Beads were washed once with with B&W-T buffer [5 mM Tris-HCl (pH 7.5), 1 M NaCl, 0.5 mM EDTA, and 0.1% Tween-20], and once with Tris-HCl (pH 8.0). To amplify the TCR cDNA containing complementarity determining region 3 (CDR3), nested PCR of the TCR locus was performed as follows. Beads with containing cDNA was resuspended with the 25 µL of first PCR mixture [0.4 µM of primers (i5, Trac_ex, and Trbc_ex), and 1x KAPA Hifi Hotstart ReadyMix (KAPA Biosystems, Wilmington, MA, #KK2602)], and the thermal cycling was performed as following condition: denaturation at 95°C for 3 min, 10 cycles of denaturation for 20 sec at 98°C, annealing for 15 sec at 65°C and extension for 30 sec at 72°C, followed by a final extension at 72°C for 2 min. Then, 2.5 µL of first PCR product was mixed with the 22.5 µl of second PCR mixture [0.35 µM of primers (i5_2nd, i7-BC_mTrac, and i7-BC_mTrbc), and 1x KAPA Hifi Hotstart ReadyMix], , and the thermal cycling was performed as following condition: denaturation at 95°C for 3 min, 18 cycles of denaturation for 20 sec at 98°C, annealing for 15 sec at 65°C and extension for 30 sec at 72°C, followed by a final extension at 72°C for 2 min.. The second-PCR products were purified by an Agencort AM Pure XP kit (Bexkman-Coulter, CA, #A63881) at a 0.7:1 ratio of beads to sample, and eluted with 20 µL of Tris-HCl (pH 8.0).To amplify TCR library and add adaptor sequence for next generation sequencer, the third PCR was performed as follows. 5µL of purified second PCR product was mixed with the 20 µl of third PCR mixture [0.4 µM of primers (i5-BC and i7-BC), and 1x KAPA Hifi Hotstart ReadyMix (KAPA Biosystems, Wilmington, MA, USA, #KK2602)], and the thermal cycling was performed under the same condition as first PCR, excepted for the number of cycles (23 cycles). The third-PCR products were purified as second PCR. The products were pooled and then purified and subjected to dual size selection using ProNex size-selective purification system (Promega) and eluted with 25 μL of Tris-HCl (pH 8.5). Final TCR libraries, whose lengths were about 600 base pairs were sequenced using an Illumina Novaseq 6000 S4 flowcell (67 bp read 1 and 140 bp read 2) (Illumina, USA). Only read2 contained the sequence regarding the definition of T cell clones. For the sinfle cell library, live cells were stained with Calcein AM (Nacalai) and loaded on an BD Rhapsody cartridge (BD) and processed using BD Rhapsody Cartridge Reagent Kit (BD) following the manufacturer’s instructions. Single-cell cDNA synthesis was performed using BD Rhapsody cDNA Kit (BD) following the manufacturer’s protocol. Whole transcriptome analysis (WTA), TCRseq, and Sample tag Index libraries were prepared using Immune Response Panel Mm (BD) according to the manufacturer’s instructions. Amplified libraries were quantified using a KAPA SYBR Fast qPCR Kit (KAPA Biosystems) and its size distribution was analyzed by a MultiNA. WTA, TCRseq, and Sample tag libraries were sequenced on an Illumina Novaseq 6000 S4 flowcell (67 bp read 1 and 140 bp read 2) using NovaSeq 6000 S4 Reagent Kit v1.5 to a depth of approximately 20,000 reads, 7,000 reads, and 10,000 reads per cell, respectively.

treatment_protocol: For treatment, anti-LAG-3 (clone C9B7W, BioLegend or Selleck) and combined anti-PD-L1 (clone 10F.9G2, BioLegend) and anti-CTLA-4 (clone 9D9, Bioxcell) were injected intraperitoneally at a dose of 200μg per mouse (100μg per antibody per mouse for combined anti-PD-L1 and anti-CTLA-4 treatment) on days 14 and 16 after tumor inoculation.

value: Tumor

value: Lewis lung carcinoma

value: lineage-negative (CD11b, B220, NK1.1 and Ter119), TCRb+,  CD4-, CD8+ T cells

value: WT


title: ExT29-WTA_001

description:
Library name: Sample 91
scRNAseqwithbulkTCRseq_immunotherapies.h5ad


title: Stem-Line II; Day 14; Bulk cells; Rep 1

description: Library name: BULK_D14_SLII_1

value: CD34+ Hematopoietic stem cell

value: Wild type

value: Day 14


title: ExT29scTCR_001

description:
Library name: Sample 89
scRNAseqwithbulkTCRseq_immunotherapies.h5ad

data_processing:
For bulk TCR analysis, adapter trimming and quality filtering of sequencing data were performed by using Cutadapt-3.2 and PRINSEQ-0.20.4. Sequencing data were processed by MiXCR-3.0.5. In MiXCR, Filtered reads were aligned to reference mouse TCR V/D/J sequences registered in the international ImMunoGeneTics information system: -starting-material=rna, -5-end=no-v-primers-, -3-end=c-primers, -adapters=no-adapters, vParameters.geneFeatureToAlign=VTranscript, -vjAlignmentOrder=JThenV, then identical sequences were assembled and grouped in clones with PCR and sequencing error correlation with the following parameters: -badQualityThreshold=15, –separateByV=true, –separateByJ=true, -only-productive=true, –region-of-interest=CDR3. The Variable (V) and Joining (J) segment of TCRs were represented in IMGT gene nomenclature. List of final clones were analyzed by VDJtools-1.2.1. Then, the sequencing reads of sample was normalized to the six times of cell count in each sample by “DownSample” command of VDJtools. T-cell clones were determined as TCR reads with the same TCR V segment, J segment and CDR3 nucleotide sequence.
For single cell analysis, reads were trimmed with Cutadapt-2.1051. Filtered reads were then annotated using the Python script provided by BD Biosciences and mapped to reference RNA using Bowtie2-2.4.254. Cell barcode information of each read was then added to the bowtie2-mapped BAM files and read counts of each gene in each cell barcode were counted using mawk. Resultant count data were converted to a single-cell gene-expression matrix file and filtered for valid cells based on read counts. Expression data of targeted panel genes were converted to a Seurat object55, log normalized and scaled (regressed by the number of unique molecular identifiers per cell) using the NormalizeData and ScaleData functions respectively. Principal component analysis (PCA) was performed using RunPCA function, and enrichment of each PC was calculated using the JackStraw and ScoreJackStraw function, and PCs that were significantly enriched (P ≤ 0.05) were selected for clustering and dimensional reduction analysis. Dimensional reduction was performed using RunUMAP function with default settings. Cell clustering was performed using FindNeighbors and FindClusters (resolution = 0.6) against the significant PCs.
Assembly: IMGT reference sequences of mouse TRB, GRCh38
Supplementary files format and content: text files in VDJtools format include read count and frequency, complementarity determining region 3 (CDR3) nucleotide and amino acid sequences, Variable(V)/ Diversity(D)/ Joining(J) segment names, and V, D and J segment boundaries within CDR3 nucleotide sequence of individual clones
Supplementary files format and content: tab-delimited text files include raw tag-count values for each sample
Library strategy: scTCR-seq


title: Stem-Line II; Day 10; Bulk cells; Rep 4

description: Library name: BULK_D10_SLII_4

value: Day 10


title: Stem-Line II; Day 14; CD16 positive; Rep 1

description: Library name: CD16pos_D14_SLII_1

value: Stem-Line II Medium; CD16 positive MACS fraction


title: Stem-Line II; Day 10; Bulk cells; Rep 3

description: Library name: BULK_D10_SLII_3

value: Stem-Line II Medium; Unsorted


title: Primary hepatocyte,shMCM7 2

value: Primary hepatocyte


title: Stem-Line II; Day 14; Bulk cells; Rep 2

description: Library name: BULK_D14_SLII_2


title: ExT29_day21_Tumor_Tim3_PDL1CTLA4_9 (bulk TCR)

description: Library name: Sample 86

data_processing:
For bulk TCR analysis, adapter trimming and quality filtering of sequencing data were performed by using Cutadapt-3.2 and PRINSEQ-0.20.4. Sequencing data were processed by MiXCR-3.0.5. In MiXCR, Filtered reads were aligned to reference mouse TCR V/D/J sequences registered in the international ImMunoGeneTics information system: -starting-material=rna, -5-end=no-v-primers-, -3-end=c-primers, -adapters=no-adapters, vParameters.geneFeatureToAlign=VTranscript, -vjAlignmentOrder=JThenV, then identical sequences were assembled and grouped in clones with PCR and sequencing error correlation with the following parameters: -badQualityThreshold=15, –separateByV=true, –separateByJ=true, -only-productive=true, –region-of-interest=CDR3. The Variable (V) and Joining (J) segment of TCRs were represented in IMGT gene nomenclature. List of final clones were analyzed by VDJtools-1.2.1. Then, the sequencing reads of sample was normalized to the six times of cell count in each sample by “DownSample” command of VDJtools. T-cell clones were determined as TCR reads with the same TCR V segment, J segment and CDR3 nucleotide sequence.
For single cell analysis, reads were trimmed with Cutadapt-2.1051. Filtered reads were then annotated using the Python script provided by BD Biosciences and mapped to reference RNA using Bowtie2-2.4.254. Cell barcode information of each read was then added to the bowtie2-mapped BAM files and read counts of each gene in each cell barcode were counted using mawk. Resultant count data were converted to a single-cell gene-expression matrix file and filtered for valid cells based on read counts. Expression data of targeted panel genes were converted to a Seurat object55, log normalized and scaled (regressed by the number of unique molecular identifiers per cell) using the NormalizeData and ScaleData functions respectively. Principal component analysis (PCA) was performed using RunPCA function, and enrichment of each PC was calculated using the JackStraw and ScoreJackStraw function, and PCs that were significantly enriched (P ≤ 0.05) were selected for clustering and dimensional reduction analysis. Dimensional reduction was performed using RunUMAP function with default settings. Cell clustering was performed using FindNeighbors and FindClusters (resolution = 0.6) against the significant PCs.
Assembly: IMGT reference sequences of mouse TRB, GRCh38
Supplementary files format and content: text files in VDJtools format include read count and frequency, complementarity determining region 3 (CDR3) nucleotide and amino acid sequences, Variable(V)/ Diversity(D)/ Joining(J) segment names, and V, D and J segment boundaries within CDR3 nucleotide sequence of individual clones
Supplementary files format and content: tab-delimited text files include raw tag-count values for each sample
Library strategy: bulk TCR-seq

value: lineage-negative (CD11b, B220, NK1.1 and Ter119), TCRb+,  CD4-, CD8+, PD-1+, Ly108- and Tim3+ T cells


title: Stem-Line II; Day 14; CD16 negative; Rep 4

description: Library name: CD16neg_D14_SLII_4

value: Stem-Line II Medium; CD16 negative MACS fraction


title: ExT29_day21_Tumor_Tim3_PDL1CTLA4_11 (bulk TCR)

description: Library name: Sample 88


title: Stem-Line II; Day 10; Bulk cells; Rep 2

description: Library name: BULK_D10_SLII_2


title: ExT29_day21_Tumor_Tim3_PDL1CTLA4_10 (bulk TCR)

description: Library name: Sample 87


title: ExT29_day21_Tumor_Tim3_PDL1CTLA4_8 (bulk TCR)

description: Library name: Sample 85


title: ExT29_day21_Tumor_Tim3_PDL1CTLA4_5 (bulk TCR)

description: Library name: Sample 84


title: Stem-Line II; Day 10; Bulk cells; Rep 1

description: Library name: BULK_D10_SLII_1


title: ExT29_day21_Tumor_Tim3_PDL1CTLA4_2 (bulk TCR)

description: Library name: Sample 83


title: Hematopoietic stem cell; Day 0; Rep 4

description: Library name: D0_SLII_4

value: Day 0


title: ExT29_day21_Tumor_Tim3_LAG3_7 (bulk TCR)

description: Library name: Sample 82


title: Hematopoietic stem cell; Day 0; Rep 6

description: Library name: D0_SLII_6


title: Hematopoietic stem cell; Day 0; Rep 5

description: Library name: D0_SLII_5


title: ExT29_day21_Tumor_Tim3_LAG3_6 (bulk TCR)

description: Library name: Sample 81


title: Stem-Line II; Day 14; Bulk cells; Rep 4

description: Library name: BULK_D14_SLII_4


title: ExT29_day21_Tumor_Tim3_LAG3_4 (bulk TCR)

description: Library name: Sample 80


title: Hematopoietic stem cell; Day 0; Rep 3

description: Library name: D0_SLII_3


title: 08_D2-2F

value: Bronchial epithelial cells

value: Flagellin


title: ExT29_day21_Tumor_Tim3_LAG3_1 (bulk TCR)

description: Library name: Sample 78


title: Hematopoietic stem cell; Day 0; Rep 1

description: Library name: D0_SLII_1


title: ExT29_day21_Tumor_PD1nega_PDL1CTLA4_11 (bulk TCR)

description: Library name: Sample 77

value: lineage-negative (CD11b, B220, NK1.1 and Ter119), TCRb+,  CD4-, CD8+, PD-1- T cells


title: ExT29_day21_Tumor_Tim3_LAG3_3 (bulk TCR)

description: Library name: Sample 79

value: Tumor


title: circFCHO2_KD_RIP_PTBP1

city: Hefei

first: Yucong

last: Wang

state: Anhui

address: 443 Huangshan Road

institute: USTC

description: Library name: Smaple16

data_processing:
To analyze RNA-seq, the high-throughput sequencing tools, Hisat2, and feature_counts, were used to map clean reads to Homo sapiens reference genome (hg19) and calculate the gene expression level. The differentially expressed gene (DEG) was determined by the “DEseq2” package in R software using the corresponding cutoff (q < 0.05, |log2(fold change)|> 1 . These DEGs were pre-ranked and the GSEA analysis was implemented in GSEA software (version 4.1.0, https:// www. gsea- msigdb. org).
Reads from RIP-seq cleandata were aligned to the hg19 genome with Bowtie2 version 2.4.1 using default settings, with all duplicates, unmapped reads, reads with more than three mismatches and non-uniquely mapped reads were removed by samtools version 1.1. The bamCoverage pipeline in deeptools (v 3.5.1) was used to generate PTBP1 binding reads coverage files (.bw). The multiBigwigSummary pipeline in deeptools was used to calculate the PTBP1 binding level on each genes. The PTBP1 binding reads coverage files was visualized by IGV (v 2.11.4) software.
Assembly: hg19
Supplementary files format and content: RNA expression level and PTBP1binding level on genes (.txt format)

molecule: other

extract_protocol:
RNA from RIP sample or whole-cell samplewas extracted with TRIzol reagent (Invitrogen) according to the manufacturer’s protocol.
The RNA from RIP sample or whole-cell sample were utilized to construct whole transcriptome library by the TruSeq Ribo Profile Library Prep Kit (Illumina, United States), following manufacturer’s instructions. In brief, rRNA was removed by Illumina Ribo-Zero Gold kit and surplus RNA was purified for end repair and 50-adaptor ligation. Then, reverse transcription was performed with random primers containing 3’ adaptor sequences. Finally, the cDNAs were purified and amplified with PCR reaction. The products with 300-500 length were purified and quantified. These libraries were utilized to 150 nt paired-end sequencing with an Illumina Nova seq 6000 system (Novogene, China). Each library was generated a depth of ~8G basepairs (for mRNA-seq sample) or ~6G basepairs (for RIP sample), and then adapters were removed with cutadapt software to obtain cleandata.

value: lung epithelial cell


title: Hematopoietic stem cell; Day 0; Rep 2

description: Library name: D0_SLII_2


title: circFCHO2_OE_RIP_PTBP1

description: Library name: Smaple15


title: ExT29_day21_Tumor_PD1nega_PDL1CTLA4_9 (bulk TCR)

description: Library name: Sample 75

value: Lewis lung carcinoma


title: ExT29_day21_Tumor_PD1nega_PDL1CTLA4_10 (bulk TCR)

description: Library name: Sample 76

value: WT


title: ExT29_day21_Tumor_PD1nega_PDL1CTLA4_5 (bulk TCR)

description: Library name: Sample 73


title: ExT29_day21_Tumor_PD1nega_PDL1CTLA4_8 (bulk TCR)

description: Library name: Sample 74


title: circFCHO2_OE_RIP_input

description: Library name: Smaple13


title: Control_OE_RIP_PTBP1

description: Library name: Smaple11


title: circFCHO2_KD_RIP_input

description: Library name: Smaple14


title: Control_KD_RIP_input

description: Library name: Smaple10


title: ExT29_day21_Tumor_PD1nega_PDL1CTLA4_2 (bulk TCR)

description: Library name: Sample 72


title: Control_OE_RIP_input

description: Library name: Smaple9


title: circFCHO2_KD_RNA_rep2

description: Library name: Smaple8


title: ExT29_day21_Tumor_PD1nega_LAG3_4 (bulk TCR)

description: Library name: Sample 69


title: ExT29_day21_Tumor_PD1nega_LAG3_6 (bulk TCR)

description: Library name: Sample 70


title: circFCHO2_KD_RNA_rep1

description: Library name: Smaple7


title: circFCHO2_OE_RNA_rep2

description: Library name: Smaple6


title: ExT29_day21_Tumor_PD1nega_LAG3_1 (bulk TCR)

description: Library name: Sample 67


title: circFCHO2_OE_RNA_rep1

description: Library name: Smaple5


title: ExT29_day21_Tumor_Ly108_PDL1CTLA4_11 (bulk TCR)

description: Library name: Sample 66

value: lineage-negative (CD11b, B220, NK1.1 and Ter119), TCRb+,  CD4-, CD8+, PD-1+, Ly108+ and Tim3- T cells


title: Control_KD_RNA_rep2

description: Library name: Smaple4

value: Wild type

value: Control


title: ExT29_day21_Tumor_Ly108_PDL1CTLA4_10 (bulk TCR)

description: Library name: Sample 65


title: Control_KD_RNA_rep1

description: Library name: Smaple3


title: ExT29_day21_Tumor_Ly108_PDL1CTLA4_9 (bulk TCR)

description: Library name: Sample 64


title: ExT29_day21_Tumor_Ly108_PDL1CTLA4_5 (bulk TCR)

description: Library name: Sample 62


title: Control_OE_RNA_rep2

description: Library name: Smaple2


title: ExT29_day21_Tumor_Ly108_PDL1CTLA4_2 (bulk TCR)

description: Library name: Sample 61


title: ExT29_day21_Tumor_PD1nega_LAG3_7 (bulk TCR)

description: Library name: Sample 71


title: ExT29_day21_Tumor_Ly108_PDL1CTLA4_8 (bulk TCR)

description: Library name: Sample 63


title: Control_KD_RIP_PTBP1

description: Library name: Smaple12


title: iPSC-derived endothelial cells, condition 1, replicate 1

city: Basel

first: Martina

last: Pigoni

address: Grenzacherstrasse 124

department: NRD

country: Switzerland

institute: Hoffmann-La Roche

data_processing:
Raw reads were processed with Cell Ranger software (version 7.1.0) and aligned to hg38 reference transcriptome (2020-A reference package provided by Cell Ranger).
Count matrices generated by Cell Ranger were corrected for ambient RNA using CellBender 0.2.0 with the expected cells parsed from the Cell Ranger web summary. The number of total droplets to include was set to 25000, the false positive rate set to 0.01 and the algorithm was trained for 150 epochs.
CellBender corrected count matrices were then concatenated into an AnnData object.
Assembly: hg38
Supplementary files format and content: CellBender corrected count matrices concatenated into one AnnData object.

source_name: Endothelial cell iPSC derived

extract_protocol:
300,000 cells differentiated to iECs (cond 1), iEC-rep (cond 2), iCE-BECs (cond 3) were plated in 6 well plates at day 11 and grown at confluence to day 14. At day 14, cells were detached and resuspended into a single cell suspension concentrated 1x106 cells per mL.
Cells were then processed for library generation following the Chromium Next GEM Single Cell 3’ v3.1 protocol for GEM generation and barcoding followed by gene expression library construction according to the manufacturer’s instructions. Targeted cell recovery of 10’000 per sample was performed.

value: Endothelial cell iPSC derived


title: iPSC-derived endothelial cells, condition 3, replicate 3


title: Control_OE_RNA_rep1

description: Library name: Smaple1


title: iPSC-derived endothelial cells, condition 2, replicate 3


title: ExT29_day21_Tumor_Ly108_LAG3_6 (bulk TCR)

description: Library name: Sample 59


title: iPSC-derived endothelial cells, condition 3, replicate 2


title: ExT29_day21_Tumor_Ly108_LAG3_3 (bulk TCR)

description: Library name: Sample 57


title: ExT29_day21_Tumor_PD1nega_LAG3_3 (bulk TCR)

description: Library name: Sample 68


title: iPSC-derived endothelial cells, condition 1, replicate 3


title: ExT29_day21_Tumor_Ly108_LAG3_7 (bulk TCR)

description: Library name: Sample 60


title: ExT29_day21_Tumor_Ly108_LAG3_4 (bulk TCR)

description: Library name: Sample 58


title: iPSC-derived endothelial cells, condition 1, replicate 2


title: iPSC-derived endothelial cells, condition 2, replicate 1


title: ExT29_day21_Tumor_DP_PDL1CTLA4_11 (bulk TCR)

description: Library name: Sample 55

value: lineage-negative (CD11b, B220, NK1.1 and Ter119), TCRb+,  CD4-, CD8+, PD-1+, Ly108+ and Tim3+ T cells


title: ExT29_day21_Tumor_DP_PDL1CTLA4_9 (bulk TCR)

description: Library name: Sample 53


title: ExT29_day21_Tumor_Ly108_LAG3_1 (bulk TCR)

description: Library name: Sample 56


title: ExT29_day21_Tumor_DP_PDL1CTLA4_8 (bulk TCR)

description: Library name: Sample 52


title: iPSC-derived endothelial cells, condition 2, replicate 2


title: ExT29_day21_Tumor_DP_PDL1CTLA4_10 (bulk TCR)

description: Library name: Sample 54


title: Whole blood, KY-0116-07, PETCT2, Normal lung, Hot LN_No, Hot Lung_No

city: Melbourne

first: Dylan

last: Sheerin

state: VIC

address: 1G Royal Parade

institute: WEHI

data_processing:
Forward and reverse reads were then aligned to the reference genome index using the Rsubread align function.
Gene counts were quantified using the BAM files generated from this step, featureCounts (v2.0.3).
Assembly: GRCh38 DNA primary assembly
Supplementary files format and content: .txt formatted gene count matrix

source_name: Whole blood

extract_protocol:
RNA was extracted from thawed Tempus tubes using the PerfectPure™ Blood RNA Kit, including DNAse treatment.
1000 ng of the isolated total RNA was globin reduced and TruSeq Stranded kit used for library preparation with 100bp pair-end (PE100) read chemistry.

value: Whole blood


title: iPSC-derived endothelial cells, condition 3, replicate 1


title: Whole blood, KY-0061-02, TB_end, Subclinical TB-inactive, Hot LN_Yes, Hot Lung_Yes

tag: initial age


title: Whole blood, KY-0116-07, PETCT1, Subclinical TB, Hot LN_Yes, Hot Lung_Yes


title: ExT29_day21_Tumor_DP_PDL1CTLA4_5 (bulk TCR)

description: Library name: Sample 51


title: ExT29_day21_Tumor_DP_LAG3_7 (bulk TCR)

description: Library name: Sample 49


title: Whole blood, KY-0106-02, PETCT1, Normal lung, Hot LN_No, Hot Lung_No


title: ExT29_day21_Tumor_DP_PDL1CTLA4_2 (bulk TCR)

description: Library name: Sample 50


title: ExT29_day21_Tumor_DP_LAG3_4 (bulk TCR)

description: Library name: Sample 47


title: Whole blood, KY-0106-02, Post Tx, No PETCT, Hot LN_NA, Hot Lung_NA

tag: petct definition

value: No PETCT

tag: other hot lung

tag: combined par + ln vs change summary


title: Whole blood, KY-0090-03, PETCT2, Normal lung, Hot LN_No, Hot Lung_No

tag: tissue


title: Whole blood, KY-0090-02, PETCT2, Subclinical TB-inactive, Hot LN_No, Hot Lung_Yes


title: ExT29_day21_Tumor_DP_LAG3_3 (bulk TCR)

description: Library name: Sample 46

tag: genotype


title: ExT29_day21_Tumor_DP_LAG3_1 (bulk TCR)

description: Library name: Sample 45

tag: cell type


title: Whole blood, KY-0097-06, PETCT2, Other cold lung lesion, Hot LN_No, Hot Lung_No


title: ExT29_day14_Tumor_Tim3_PDL1CTLA4_11 (bulk TCR)

description: Library name: Sample 44

value: lineage-negative (CD11b, B220, NK1.1 and Ter119), TCRb+,  CD4-, CD8+, PD-1+, Ly108- and Tim3+ T cells


title: Whole blood, KY-0076-02, PETCT2, Subclinical TB, Hot LN_Yes, Hot Lung_Yes


title: ExT29_day14_Tumor_Tim3_PDL1CTLA4_10 (bulk TCR)

description: Library name: Sample 43


title: ExT29_day14_Tumor_Tim3_PDL1CTLA4_9 (bulk TCR)

description: Library name: Sample 42


title: Whole blood, KY-0076-02, TB_end, Subclinical TB, Hot LN_Yes, Hot Lung_Yes


title: Whole blood, KY-0016-04, TB_end, Subclinical TB, Hot LN_No, Hot Lung_Yes


title: Whole blood, KY-0081-07, PETCT2, Subclinical TB-inactive, Hot LN_No, Hot Lung_Yes

value: Minimal worsening


title: Whole blood, KY-0016-04, PETCT2, Normal lung, Hot LN_No, Hot Lung_No


title: ExT29_day14_Tumor_Tim3_PDL1CTLA4_5 (bulk TCR)

description: Library name: Sample 40


title: ExT29_day14_Tumor_Tim3_PDL1CTLA4_2 (bulk TCR)

description: Library name: Sample 39


title: Whole blood, KY-0015-02, TB_end, Subclinical TB, Hot LN_Yes, Hot Lung_Yes


title: Whole blood, KY-0121-04, PETCT1, Subclinical TB, Hot LN_Yes, Hot Lung_Yes

tag: participant id


title: Whole blood, KY-0015-02, PETCT2, Subclinical TB, Hot LN_No, Hot Lung_Yes


title: ExT29_day14_Tumor_Tim3_LAG3_6 (bulk TCR)

description: Library name: Sample 37

tag: cell line


title: Whole blood, KY-0098-02, PETCT1, Other cold lung lesion, Hot LN_No, Hot Lung_No


title: ExT29_day14_Tumor_Tim3_LAG3_7 (bulk TCR)

description: Library name: Sample 38


title: ExT29_day21_Tumor_DP_LAG3_6 (bulk TCR)

description: Library name: Sample 48


title: ExT29_day14_Tumor_Tim3_LAG3_3 (bulk TCR)

description: Library name: Sample 35


title: Whole blood, KY-0097-06, PETCT1, Other cold lung lesion, Hot LN_No, Hot Lung_No


title: ExT29_day14_Tumor_Tim3_LAG3_4 (bulk TCR)

description: Library name: Sample 36


title: ExT29_day14_Tumor_PD1nega_PDL1CTLA4_11 (bulk TCR)

description: Library name: Sample 33


title: ExT29_day14_Tumor_Tim3_LAG3_1 (bulk TCR)

description: Library name: Sample 34


title: Whole blood, KY-0087-04, PETCT1, Normal lung, Hot LN_No, Hot Lung_No


title: ExT29_day14_Tumor_PD1nega_PDL1CTLA4_10 (bulk TCR)

description: Library name: Sample 32


title: Whole blood, KY-0084-05, PETCT1, Normal lung, Hot LN_No, Hot Lung_No


title: ExT29_day14_Tumor_PD1nega_PDL1CTLA4_9 (bulk TCR)

description: Library name: Sample 31


title: ExT29_day14_Tumor_PD1nega_PDL1CTLA4_8 (bulk TCR)

description: Library name: Sample 30


title: ExT29_day14_Tumor_PD1nega_PDL1CTLA4_5 (bulk TCR)

description: Library name: Sample 29


title: ExT29_day14_Tumor_Tim3_PDL1CTLA4_8 (bulk TCR)

description: Library name: Sample 41


title: ExT29_day14_Tumor_PD1nega_LAG3_7 (bulk TCR)

description: Library name: Sample 27


title: Whole blood, KY-0097-07, PETCT2, Subclinical TB-inactive, Hot LN_No, Hot Lung_Yes


title: Whole blood, KY-0074-05, PETCT1, Subclinical TB, Hot LN_No, Hot Lung_Yes


title: Whole blood, KY-0031-03, Post Tx, Subclinical TB, Hot LN_No, Hot Lung_Yes


title: ExT29_day14_Tumor_PD1nega_PDL1CTLA4_2 (bulk TCR)

description: Library name: Sample 28


title: Whole blood, KY-0031-03, PETCT1, Subclinical TB, Hot LN_No, Hot Lung_Yes


title: Whole blood, KY-0097-07, PETCT1, Subclinical TB, Hot LN_No, Hot Lung_Yes


title: Whole blood, KY-0090-03, PETCT1, Normal lung, Hot LN_No, Hot Lung_No


title: ExT29_day14_Tumor_PD1nega_LAG3_6 (bulk TCR)

description: Library name: Sample 26


title: ExT29_day14_Tumor_PD1nega_LAG3_1 (bulk TCR)

description: Library name: Sample 23


title: Whole blood, KY-0093-03, PETCT1, Subclinical TB, Hot LN_No, Hot Lung_Yes


title: Whole blood, KY-0097-02, PETCT1, Subclinical TB, Hot LN_No, Hot Lung_Yes


title: Whole blood, KY-0110-05, PETCT1, Subclinical TB, Hot LN_Yes, Hot Lung_Yes


title: ExT29_day14_Tumor_PD1nega_LAG3_4 (bulk TCR)

description: Library name: Sample 25


title: ExT29_day14_Tumor_PD1nega_LAG3_3 (bulk TCR)

description: Library name: Sample 24


title: Whole blood, KY-0089-09, PETCT1, Normal lung, Hot LN_No, Hot Lung_No


title: Whole blood, KY-0089-11, PETCT1, Normal lung, Hot LN_No, Hot Lung_No


title: ExT29_day14_Tumor_Ly108_PDL1CTLA4_10 (bulk TCR)

description: Library name: Sample 21


title: ExT29_day14_Tumor_Ly108_PDL1CTLA4_11 (bulk TCR)

description: Library name: Sample 22


title: ExT29_day14_Tumor_Ly108_PDL1CTLA4_9 (bulk TCR)

description: Library name: Sample 20


title: Whole blood, KY-0090-01, PETCT1, Other cold lung lesion, Hot LN_No, Hot Lung_No


title: ExT29_day14_Tumor_Ly108_PDL1CTLA4_8 (bulk TCR)

description: Library name: Sample 19


title: Whole blood, KY-0088-01, PETCT1, Normal lung, Hot LN_No, Hot Lung_No


title: ExT29_day14_Tumor_Ly108_PDL1CTLA4_5 (bulk TCR)

description: Library name: Sample 18


title: Whole blood, KY-0089-04, PETCT1, Normal lung, Hot LN_No, Hot Lung_No


title: Whole blood, KY-0085-05, PETCT2, Normal lung, Hot LN_No, Hot Lung_No

value: F


title: ExT29_day14_Tumor_Ly108_LAG3_6 (bulk TCR)

description: Library name: Sample 15


title: ExT29_day14_Tumor_Ly108_LAG3_7 (bulk TCR)

description: Library name: Sample 16


title: ExT29_day14_Tumor_Ly108_LAG3_4 (bulk TCR)

description: Library name: Sample 14


title: Whole blood, KY-0084-06, PETCT1, Other hot lung lesion, Hot LN_No, Hot Lung_Yes


title: Whole blood, KY-0084-01, PETCT1, Subclinical TB, Hot LN_No, Hot Lung_No


title: Whole blood, KY-0082-03, PETCT1, Normal lung, Hot LN_No, Hot Lung_No


title: ExT29_day14_Tumor_Ly108_LAG3_1 (bulk TCR)

description: Library name: Sample 12


title: Whole blood, KY-0081-10, PETCT1, Subclinical TB-inactive, Hot LN_No, Hot Lung_No


title: ExT29_day14_Tumor_DP_PDL1CTLA4_10 (bulk TCR)

description: Library name: Sample 10


title: Whole blood, KY-0090-02, PETCT1, Subclinical TB-inactive, Hot LN_No, Hot Lung_Yes


title: Whole blood, KY-0081-05, PETCT1, Subclinical TB, Hot LN_Yes, Hot Lung_Yes


title: Whole blood, KY-0081-07, PETCT1, Subclinical TB-inactive, Hot LN_No, Hot Lung_Yes


title: Whole blood, KY-0081-05, Post Tx, Subclinical TB, Hot LN_Yes, Hot Lung_Yes


title: ExT29_day14_Tumor_DP_PDL1CTLA4_11 (bulk TCR)

description: Library name: Sample 11


title: ExT29_day14_Tumor_DP_PDL1CTLA4_8 (bulk TCR)

description: Library name: Sample 8


title: Whole blood, KY-0081-03, PETCT2, Subclinical TB-inactive, Hot LN_No, Hot Lung_No


title: ExT29_day14_Tumor_Ly108_PDL1CTLA4_2 (bulk TCR)

description: Library name: Sample 17


title: Whole blood, KY-0081-03, PETCT1, Subclinical TB, Hot LN_No, Hot Lung_Yes


title: Whole blood, KY-0081-02, PETCT2, Subclinical TB, Hot LN_Yes, Hot Lung_Yes


title: Whole blood, KY-0085-05, PETCT1, Normal lung, Hot LN_No, Hot Lung_No


title: ExT29_day14_Tumor_DP_LAG3_6 (bulk TCR)

description: Library name: Sample 4


title: ExT29_day14_Tumor_DP_PDL1CTLA4_5 (bulk TCR)

description: Library name: Sample 7


title: ExT29_day14_Tumor_DP_LAG3_7 (bulk TCR)

description: Library name: Sample 5


title: ExT29_day14_Tumor_DP_LAG3_4 (bulk TCR)

description: Library name: Sample 3


title: Whole blood, KY-0084-06, PETCT2, Other hot lung lesion, Hot LN_No, Hot Lung_Yes


title: Whole blood, KY-0081-01, PETCT1, Other hot lung lesion, Hot LN_No, Hot Lung_Yes


title: ExT29_day14_Tumor_Ly108_LAG3_3 (bulk TCR)

description: Library name: Sample 13


title: ExT29_day14_Tumor_DP_LAG3_3 (bulk TCR)

description: Library name: Sample 2


title: ExT29_day14_Tumor_DP_LAG3_1 (bulk TCR)

description: Library name: Sample 1


title: Whole blood, KY-0077-02, PETCT1, Other cold lung lesion, Hot LN_No, Hot Lung_No


title: Whole blood, KY-0078-01, PETCT1, Other hot lung lesion, Hot LN_Yes, Hot Lung_Yes


title: BilateralWTA1_L004

description:
Library name: Sample 52
scRNAseqwithbulkTCRseq_untreated.h5ad

data_processing:
For bulk TCR analysis, adapter trimming and quality filtering of sequencing data were performed by using Cutadapt-3.2 and PRINSEQ-0.20.4. Sequencing data were processed by MiXCR-3.0.5. In MiXCR, Filtered reads were aligned to reference mouse TCR V/D/J sequences registered in the international ImMunoGeneTics information system: -starting-material=rna, -5-end=no-v-primers-, -3-end=c-primers, -adapters=no-adapters, vParameters.geneFeatureToAlign=VTranscript, -vjAlignmentOrder=JThenV, then identical sequences were assembled and grouped in clones with PCR and sequencing error correlation with the following parameters: -badQualityThreshold=15, –separateByV=true, –separateByJ=true, -only-productive=true, –region-of-interest=CDR3. The Variable (V) and Joining (J) segment of TCRs were represented in IMGT gene nomenclature. List of final clones were analyzed by VDJtools-1.2.1. Then, the sequencing reads of sample was normalized to the six times of cell count in each sample by “DownSample” command of VDJtools. T-cell clones were determined as TCR reads with the same TCR V segment, J segment and CDR3 nucleotide sequence.
For single cell analysis, reads were trimmed with Cutadapt-2.1051. Filtered reads were then annotated using the Python script provided by BD Biosciences and mapped to reference RNA using Bowtie2-2.4.254. Cell barcode information of each read was then added to the bowtie2-mapped BAM files and read counts of each gene in each cell barcode were counted using mawk. Resultant count data were converted to a single-cell gene-expression matrix file and filtered for valid cells based on read counts. Expression data of targeted panel genes were converted to a Seurat object55, log normalized and scaled (regressed by the number of unique molecular identifiers per cell) using the NormalizeData and ScaleData functions respectively. Principal component analysis (PCA) was performed using RunPCA function, and enrichment of each PC was calculated using the JackStraw and ScoreJackStraw function, and PCs that were significantly enriched (P ≤ 0.05) were selected for clustering and dimensional reduction analysis. Dimensional reduction was performed using RunUMAP function with default settings. Cell clustering was performed using FindNeighbors and FindClusters (resolution = 0.6) against the significant PCs.
Assembly: IMGT reference sequences of mouse TRB, GRCh38
Supplementary files format and content: text files in VDJtools format include read count and frequency, complementarity determining region 3 (CDR3) nucleotide and amino acid sequences, Variable(V)/ Diversity(D)/ Joining(J) segment names, and V, D and J segment boundaries within CDR3 nucleotide sequence of individual clones
Supplementary files format and content: tab-delimited text files include raw tag-count values for each sample
Supplementary files format and content: h5ad for scRNA-seq samples

extract_protocol:
Tumors were cut into small fragments and digested for 45 minutes at 37°C with 0.2% collagenase (FUJIFILM Wako) and DNase I (Sigma-Aldrich), except for experiments for single-cell analysis, in which tumors were digested with 0.2% collagenase NB8 (Nordmark Pharma GmbH). The cells were then subjected to two-phase density separation with upper 35% Percoll PLUS (GE Healthcare) (1000g, 10 minutes at room temperature). Leukocytes were recovered, washed and resuspended in preparation medium (PM) [5%FBS, 10mM HEPES (Nacalai), Penicillin-Streptomycin (Nacalai) in RPMI-1640 (Nacalai)]. After RBC lysis with ACK buffer [155mM of NH4Cl (Nacalai), 10mM of KHCO3 (Nacalai), and 0.5mM EDTA pH8.0 (NIPPON GENE)] for 1 minute at room temperature, cells were washed and resuspended in PM, and the cell concentrations were measured using Flow-Count fluorospheres (Beckman Coulter) and a CytoFLEX S flow cytometer (Beckman Coulter).Cells were stained with a mix of Fc Block (anti-mouse CD16/CD32 mAb; clone 2.4G2, BioLegend) and fluorophore-conjugated anti-mouse monoclonal antibodies (Supplementary Table 5). Antibody mixtures were prepared using BD Horizon Brilliant Stain Buffer Plus (BD) diluted 5x with FACS buffer (D-PBS(-) supplemented with 2% FBS, and 0.05% Sodium Azide). Staining was performed for 20 minutes at 4°C. In the single-cell RNA-seq/TCR-seq experiment, cells were stained with antibodies and anti-CD45 Sample Tag oligonucleotide-conjugated antibodies from the Single-Cell Multiplexing Kit (BD Biosciences). Before sorting, T cells were enriched from tumor suspensions by further staining with BD IMag APC Magnetic Particles-DM (BD Biosciences), then placed on magnetic plate. After washing by MACS buffer (D-PBS(-) supplemented with 10% BSA, and 2 mM EDTA pH8.0), APC and APC-Cy7(CD8, mouse TCRb)-positive cells were collected as positive fraction. Cell sorting was performed using a FACS Aria II or Aria III (BD Biosciences). Cells were stained with 5µg/mL of propidium iodide (PI, Sigma-Aldrich) immediately before sorting. Nonviable cells were excluded from the analysis based on forward and side scatter profiles and PI staining. For bulk TCR repertoire analysis and bulk RNA-seq analysis, cells were sorted into Lysis buffer [1% w/v LiDS, 100 mM Tris-HCl pH7.5, 500 mM LiCl, 5 mM dithiothreitol, and 10 mM EDTA pH8.0] and stored at −80°C. For single-cell RNA/TCR-seq, cells were sorted into D-PBS(-) with 10% FBS. For the bulk TCR-seq experiments, cells were sorted from the CD8+, CD8+PD-1+Ly108+TIM-3-, CD8+PD-1+Ly108+TIM-3+, CD8+PD-1+Ly108-TIM-3+ and CD8+PD-1- fractions to produce five datasets per mouse. For the single-cell RNA/TCR-seq and bulk TCR-seq experiment, cells were sorted from the CD8+PD-1+Ly108+TIM-3-, CD8+PD-1+Ly108+TIM-3+, CD8+PD-1+Ly108-TIM-3+ and CD8+PD-1- fractions.
Bulk TCR libraries were prepared on purified T cells lysed in lysis buffer [1% LiDS, 100 mM Tris-HCl (pH 7.5), 500 mM LiCl, and 10 mM EDTA]. PolyA RNAs were isolated according to a previous report with some modifications (GSE110711).To perform reverse transcription and template-swiching, beads were then suspended in 10 µL of RT mix [1× First Strand buffer (Thermo Fisher Scientific), 1 mM dNTP, 2.5 mM DTT (Thermo Fisher Scientific), 1 M betaine (Sigma-Aldrich), 9 mM MgCl2, 1 U/µL RNaseIn Plus Rnase Inhibitor (Promega), 10 U/µL Superscript II (Thermo Fisher Scientific), and 1µM of i5-TSO], and incubated for 60 min at 42ºC and immediately cooled on ice. Beads were washed once with with B&W-T buffer [5 mM Tris-HCl (pH 7.5), 1 M NaCl, 0.5 mM EDTA, and 0.1% Tween-20], and once with Tris-HCl (pH 8.0). To amplify the TCR cDNA containing complementarity determining region 3 (CDR3), nested PCR of the TCR locus was performed as follows. Beads with containing cDNA was resuspended with the 25 µL of first PCR mixture [0.4 µM of primers (i5, Trac_ex, and Trbc_ex), and 1x KAPA Hifi Hotstart ReadyMix (KAPA Biosystems, Wilmington, MA, #KK2602)], and the thermal cycling was performed as following condition: denaturation at 95°C for 3 min, 10 cycles of denaturation for 20 sec at 98°C, annealing for 15 sec at 65°C and extension for 30 sec at 72°C, followed by a final extension at 72°C for 2 min. Then, 2.5 µL of first PCR product was mixed with the 22.5 µl of second PCR mixture [0.35 µM of primers (i5_2nd, i7-BC_mTrac, and i7-BC_mTrbc), and 1x KAPA Hifi Hotstart ReadyMix], , and the thermal cycling was performed as following condition: denaturation at 95°C for 3 min, 18 cycles of denaturation for 20 sec at 98°C, annealing for 15 sec at 65°C and extension for 30 sec at 72°C, followed by a final extension at 72°C for 2 min.. The second-PCR products were purified by an Agencort AM Pure XP kit (Bexkman-Coulter, CA, #A63881) at a 0.7:1 ratio of beads to sample, and eluted with 20 µL of Tris-HCl (pH 8.0).To amplify TCR library and add adaptor sequence for next generation sequencer, the third PCR was performed as follows. 5µL of purified second PCR product was mixed with the 20 µl of third PCR mixture [0.4 µM of primers (i5-BC and i7-BC), and 1x KAPA Hifi Hotstart ReadyMix (KAPA Biosystems, Wilmington, MA, USA, #KK2602)], and the thermal cycling was performed under the same condition as first PCR, excepted for the number of cycles (23 cycles). The third-PCR products were purified as second PCR. The products were pooled and then purified and subjected to dual size selection using ProNex size-selective purification system (Promega) and eluted with 25 μL of Tris-HCl (pH 8.5). Final TCR libraries, whose lengths were about 600 base pairs were sequenced using an Illumina Novaseq 6000 S4 flowcell (67 bp read 1 and 140 bp read 2) (Illumina, USA). Only read2 contained the sequence regarding the definition of T cell clones. For the single cell library, live cells were stained with Calcein AM (Nacalai) and loaded on an BD Rhapsody cartridge (BD) and processed using BD Rhapsody Cartridge Reagent Kit (BD) following the manufacturer’s instructions. Single-cell cDNA synthesis was performed using BD Rhapsody cDNA Kit (BD) following the manufacturer’s protocol. Whole transcriptome analysis (WTA), TCRseq, and Sample tag Index libraries were prepared using Immune Response Panel Mm (BD) according to the manufacturer’s instructions. Amplified libraries were quantified using a KAPA SYBR Fast qPCR Kit (KAPA Biosystems) and its size distribution was analyzed by a MultiNA. WTA, TCRseq, and Sample tag libraries were sequenced on an Illumina Novaseq 6000 S4 flowcell (67 bp read 1 and 140 bp read 2) using NovaSeq 6000 S4 Reagent Kit v1.5 to a depth of approximately 20,000 reads, 7,000 reads, and 10,000 reads per cell, respectively.

value: lineage-negative (CD11b, B220, NK1.1 and Ter119), TCRb+,  CD4-, CD8+ T cells


title: BilateralWTA1_L001

description:
Library name: Sample 49
scRNAseqwithbulkTCRseq_untreated.h5ad


title: BilateralWTA1_L003

description:
Library name: Sample 51
scRNAseqwithbulkTCRseq_untreated.h5ad


title: BilateralWTA1_L002

description:
Library name: Sample 50
scRNAseqwithbulkTCRseq_untreated.h5ad


title: Whole blood, KY-0076-03, PETCT1, Normal lung, Hot LN_No, Hot Lung_No


title: Whole blood, KY-0076-02, PETCT1, Subclinical TB, Hot LN_Yes, Hot Lung_Yes


title: ExT29_day14_Tumor_DP_PDL1CTLA4_9 (bulk TCR)

description: Library name: Sample 9


title: Whole blood, KY-0074-04, PETCT1, Subclinical TB-inactive, Hot LN_Yes, Hot Lung_No


title: Whole blood, KY-0081-02, PETCT1, Subclinical TB-inactive, Hot LN_Yes, Hot Lung_Yes


title: Whole blood, KY-0081-01, PETCT2, Other hot lung lesion, Hot LN_Yes, Hot Lung_Yes


title: Whole blood, KY-0073-04, PETCT1, Normal lung, Hot LN_No, Hot Lung_No


title: Whole blood, KY-0074-03, PETCT1, Other hot lung lesion, Hot LN_Yes, Hot Lung_Yes


title: Whole blood, KY-0072-01, PETCT2, Subclinical TB-inactive, Hot LN_Yes, Hot Lung_No


title: BilateralTag1_L003

description:
Library name: Sample 43
scRNAseqwithbulkTCRseq_untreated.h5ad


title: ExT29_day14_Tumor_DP_PDL1CTLA4_2 (bulk TCR)

description: Library name: Sample 6


title: Whole blood, KY-0070-05, PETCT1, Normal lung, Hot LN_No, Hot Lung_No


title: BilateralTCR1_L003

description:
Library name: Sample 47
scRNAseqwithbulkTCRseq_untreated.h5ad


title: BilateralTag1_L004

description:
Library name: Sample 44
scRNAseqwithbulkTCRseq_untreated.h5ad


title: Whole blood, KY-0072-01, PETCT1, Subclinical TB-inactive, Hot LN_Yes, Hot Lung_No


title: BilateralTCR1_L001

description:
Library name: Sample 45
scRNAseqwithbulkTCRseq_untreated.h5ad


title: BilateralTCR1_L002

description:
Library name: Sample 46
scRNAseqwithbulkTCRseq_untreated.h5ad


title: BilateralTag1_L002

description:
Library name: Sample 42
scRNAseqwithbulkTCRseq_untreated.h5ad


title: BilateralTag1_L001

description:
Library name: Sample 41
scRNAseqwithbulkTCRseq_untreated.h5ad


title: Whole blood, KY-0069-02, PETCT1, Normal lung, Hot LN_No, Hot Lung_No


title: Whole blood, KY-0067-01, PETCT2, Subclinical TB, Hot LN_No, Hot Lung_Yes


title: Whole blood, KY-0070-01, PETCT1, Normal lung, Hot LN_Yes, Hot Lung_No


title: Whole blood, KY-0073-01, PETCT1, Other cold lung lesion, Hot LN_Yes, Hot Lung_No


title: ExT6_day21_Tumor_Tim3_5

description: Library name: Sample 40

tag: time


title: Whole blood, KY-0075-02, PETCT1, Other hot lung lesion, Hot LN_Yes, Hot Lung_Yes


title: BilateralTCR1_L004

description:
Library name: Sample 48
scRNAseqwithbulkTCRseq_untreated.h5ad


title: ExT6_day21_Tumor_Tim3_2

description: Library name: Sample 37


title: Whole blood, KY-0065-07, PETCT1, Normal lung, Hot LN_No, Hot Lung_No


title: Whole blood, KY-0067-01, PETCT1, Subclinical TB-inactive, Hot LN_No, Hot Lung_Yes


title: ExT6_day21_Tumor_Tim3_1

description: Library name: Sample 36


title: ExT6_day21_Tumor_Tim3_3

description: Library name: Sample 38


title: Whole blood, KY-0065-07, PETCT2, Normal lung, Hot LN_No, Hot Lung_No


title: ExT6_day21_Tumor_PD1nega_5

description: Library name: Sample 35


title: ExT6_day21_Tumor_PD1nega_4

description: Library name: Sample 34


title: Whole blood, KY-0065-06, PETCT1, Subclinical TB-inactive, Hot LN_No, Hot Lung_No


title: Whole blood, KY-0065-06, PETCT2, Subclinical TB, Hot LN_No, Hot Lung_Yes


title: ExT6_day21_Tumor_PD1nega_3

description: Library name: Sample 33


title: Whole blood, KY-0064-01, PETCT1, Other hot lung lesion, Hot LN_Yes, Hot Lung_Yes


title: Whole blood, KY-0064-01, PETCT2, Other hot lung lesion, Hot LN_Yes, Hot Lung_Yes


title: Whole blood, KY-0063-01, PETCT1, Normal lung, Hot LN_No, Hot Lung_No


title: ExT6_day21_Tumor_PD1nega_2

description: Library name: Sample 32


title: RP11-175E9.1-g1 CRISPRi WTC (NKX2-6-AS1 K/D), Day 6, No Xav differentiation, HTO library

description:
Library name: Sample_2_HTO
10X Genomics

value: Heart

value: RP11-175E9.1-g1 WTC CRISPRi GCaMP

value: human iPSCs

tag: treatment

value: Doxycycline-induced K/D


title: WTC CRISPRi (Control), Day 6, No Xav differentiation, HTO library

description:
Library name: Sample_1_HTO
10X Genomics

value: WTC CRISPRi GCaMP


title: Whole blood, KY-0061-02, PETCT1, Subclinical TB, Hot LN_Yes, Hot Lung_Yes


title: HPaMEC cells, HuR KO EV, 24h_1

description: Library name: Sample 12

value: pancreas

value: HPaMEC

value: pancreatic endothelial cell

value: WT

value: HuR KO EV


title: HPaMEC cells, HuR KO EV, 24h_2

description: Library name: Sample 13


title: ExT6_day21_Tumor_Tim3_4

description: Library name: Sample 39

value: Tumor

value: Lewis lung carcinoma


title: ExT6_day21_Tumor_PD1nega_1

description: Library name: Sample 31

value: lineage-negative (CD11b, B220, NK1.1 and Ter119), TCRb+,  CD4-, CD8+, PD-1- T cells


title: ExT6_day21_Tumor_Ly108_5

description: Library name: Sample 30

value: lineage-negative (CD11b, B220, NK1.1 and Ter119), TCRb+,  CD4-, CD8+, PD-1+, Ly108+ and Tim3- T cells


title: HPaMEC cells, WT EV, 24h_3

description: Library name: Sample 11

value: WT EV


title: Whole blood, KY-0060-03, PETCT1, Normal lung, Hot LN_No, Hot Lung_No

value: Whole blood


title: HPaMEC cells, WT EV, 24h_2

description: Library name: Sample 10


title: HPaMEC cells, WT EV, 24h_1

description: Library name: Sample 9


title: ExT6_day21_Tumor_Ly108_4

description: Library name: Sample 29


title: Whole blood, KY-0060-02, PETCT2, Normal lung, Hot LN_No, Hot Lung_No


title: shSHCBP1-IL-1beta-3

description:
HL7702,shSHCBP1，8h
shSHCBP1-IL-1β-3

data_processing:
Base calling of sequencing data was performed using bcl2fastq software (the software is officially delivered by illumina) to obtain fastq software.
Use fastp (v 0.19.7) fastq data for filtering to get cleandata. The main parameters are (-W 5-M 20-3-l 20-w 10).
Use the HiSat2 software (2.1.0) to compare cleandata to the reference genome with default parameters.
Use StringTie software (v1.3.4d) and annotate the result to calculate the data expression FPKM (Gene.FPKM.xls); Calculate ReadsCount (i.e. Gene.Count.xls) according to the script given by the official StringTie.
Assembly: GRCh38
Supplementary files format and content: tab-delimited tex file includes raw counts for each Sample
Supplementary files format and content: tab-delimited tex files include RPKM values for each Sample

growth_protocol: HL7702 cells were mantained in DMEM supplemented with 10% fetal bovine serum(FBS) and antibiotics in humidified atmosphere with 5% CO2 at 37℃

treatment_protocol: 10ng/ml Recombinant human IL-1β (PeproTech) was added to stimulate the HL7702

value: hepatocyte

value: SHCBP1 knockdown

value: recombinate IL-1beta, 8h


title: Whole blood, KY-0060-02, PETCT1, Normal lung, Hot LN_No, Hot Lung_No


title: shSHCBP1-IL-1beta-1

description:
HL7702,shSHCBP1，8h
shSHCBP1-IL-1β-1


title: shSHCBP1-IL-1beta-2

description:
HL7702,shSHCBP1，8h
shSHCBP1-IL-1β-2


title: ExT6_day21_Tumor_Ly108_2

description: Library name: Sample 27


title: shMCM7-IL-1beta-3

description:
HL7702,shMCM7，8h
shMCM7-IL-1β-3

value: MCM7 knockdown


title: Whole blood, KY-0056-02, PETCT2, Other cold lung lesion, Hot LN_No, Hot Lung_No


title: ExT6_day21_Tumor_Ly108_3

description: Library name: Sample 28


title: ExT6_day21_Tumor_Ly108_1

description: Library name: Sample 26


title: Whole blood, KY-0056-02, PETCT1, Other cold lung lesion, Hot LN_No, Hot Lung_No


title: shMCM7-IL-1beta-1

description:
HL7702,shMCM7，8h
shMCM7-IL-1β-1


title: HPaMEC cells, HuR KO EV, 24h_3

description: Library name: Sample 14


title: shMCM7-IL-1beta-2

description:
HL7702,shMCM7，8h
shMCM7-IL-1β-2


title: PLKO.1-IL-1beta-3

description:
HL7702,shControl，8h
PLKO.1-IL-1β-3


title: Whole blood, KY-0053-04, PETCT1, Other cold lung lesion, Hot LN_No, Hot Lung_No


title: ExT6_day21_Tumor_DP_5

description: Library name: Sample 25

value: lineage-negative (CD11b, B220, NK1.1 and Ter119), TCRb+,  CD4-, CD8+, PD-1+, Ly108+ and Tim3+ T cells


title: PLKO.1-IL-1beta-2

description:
HL7702,shControl，8h
PLKO.1-IL-1β-2


title: ExT6_day21_Tumor_DP_4

description: Library name: Sample 24


title: Whole blood, KY-0053-03, PETCT2, Normal lung, Hot LN_No, Hot Lung_No


title: ExT6_day21_Tumor_DP_3

description: Library name: Sample 23


title: Whole blood, KY-0053-03, PETCT1, Normal lung, Hot LN_No, Hot Lung_No


title: shSHCBP1-1

description:
HL7702,shSHCBP1
shSHCBP1-1

value: untreatment


title: shMCM7-3

description:
HL7702,shMCM7
shMCM7-3


title: PLKO.1-IL-1beta-1

description:
HL7702,shControl，8h
PLKO.1-IL-1β-1


title: shSHCBP1-3

description:
HL7702,shSHCBP1
shSHCBP1-3


title: shSHCBP1-2

description:
HL7702,shSHCBP1
shSHCBP1-2


title: Whole blood, KY-0051-10, PETCT2, Subclinical TB, Hot LN_Yes, Hot Lung_Yes


title: shMCM7-2

description:
HL7702,shMCM7
shMCM7-2


title: shMCM7-1

description:
HL7702,shMCM7
shMCM7-1


title: ExT6_day21_Tumor_DP_1

description: Library name: Sample 21


title: PLKO.1-3

description:
HL7704,shControl
PLKO.1-3


title: PLKO.1-2

description:
HL7703,shControl
PLKO.1-2


title: Whole blood, KY-0051-10, PETCT1, Subclinical TB, Hot LN_Yes, Hot Lung_Yes


title: PLKO.1-1

description:
HL7702,shControl
PLKO.1-1


title: ExT6_day14_Tumor_Tim3_5

description: Library name: Sample 20

value: lineage-negative (CD11b, B220, NK1.1 and Ter119), TCRb+,  CD4-, CD8+, PD-1+, Ly108- and Tim3+ T cells


title: Whole blood, KY-0051-08, PETCT1, Other cold lung lesion, Hot LN_No, Hot Lung_No


title: ExT6_day14_Tumor_Tim3_4

description: Library name: Sample 19


title: ExT6_day14_Tumor_Tim3_3

description: Library name: Sample 18


title: Whole blood, KY-0051-01, PETCT2, Other hot lung lesion, Hot LN_No, Hot Lung_Yes


title: testes of blue catfsh sampled at age 4.2, biological rep3

city: Auburn

first: Ian

last: Butts

state: Alabama

address: 203, Swingle Hall

department: School of Fisheries, Aquaculture & Aquatic Sciences

institute: Auburn University

description: Library name: Seasonal_Sample_13

data_processing:
Raw, demultiplexed sequence files were provided in FASTQ format. The quality of raw RNA sequencing reads was assessed by FastQC (V.0.11.9). Clean reads were obtained by removing low-quality reads and adapter sequences from the raw data using Trimmomatic (V.0.39).
The genome of blue catfish (I. furcatus) was obtained from NCBI and remaining high-quality reads were aligned to the genome using STAR aligner (V.2.7.5). Post mapping quality control was performed using MultiQC (V.1.10.1).
BAM files were used to generate a gene-level count data matrix for all samples via HTSeq-count (V. 0.11.1).
Pairwise differential gene expression analyses were performed using edgeR (V.4.4.2)
Assembly: GCF_023375685.1
Supplementary files format and content: CSV files include raw counts for each sample

organism: Ictalurus furcatus

source_name: Testis

extract_protocol:
Total RNA was extracted using Quick-RNA Miniprep Plus Kit (Zymo Research) following the manufacturer’s instructions. Total RNA was treated with DNase I and resuspended in 50 µL of nuclease-free water.
Sequencing libraries were prepared using a NEBNext Ultra II RNA Library Preparation Kit (NEB, Ipswich, MA, U.S.), following manufacturer’s instructions

value: Testis

value: male

value: Age 4.2


title: testes of blue catfsh sampled at age 4.2, biological rep4

description: Library name: Seasonal_Sample_14


title: testes of blue catfsh sampled at age 4.2, biological rep1

description: Library name: Seasonal_Sample_11


title: Whole blood, KY-0051-01, PETCT1, Other hot lung lesion, Hot LN_No, Hot Lung_Yes


title: ExT6_day14_Tumor_Tim3_2

description: Library name: Sample 17

value: WT


title: testes of blue catfsh sampled at age 4.2, biological rep2

description: Library name: Seasonal_Sample_12


title: Whole blood, KY-0048-03, PETCT2, Subclinical TB, Hot LN_No, Hot Lung_Yes


title: testes of blue catfsh sampled at age 3.9, biological rep2

description: Library name: Seasonal_Sample_10

value: Age 3.9


title: ExT6_day21_Tumor_DP_2

description: Library name: Sample 22

value: Tumor

value: Lewis lung carcinoma


title: Whole blood, KY-0048-03, PETCT1, Subclinical TB, Hot LN_No, Hot Lung_Yes


title: testes of blue catfsh sampled at age 3.1, biological rep3

description: Library name: Seasonal_Sample_7

value: Age 3.1


title: ExT6_day14_Tumor_PD1nega_5

description: Library name: Sample 15

value: lineage-negative (CD11b, B220, NK1.1 and Ter119), TCRb+,  CD4-, CD8+, PD-1- T cells


title: ExT6_day14_Tumor_Tim3_1

description: Library name: Sample 16


title: testes of blue catfsh sampled at age 3.1, biological rep1

description: Library name: Seasonal_Sample_5


title: testes of blue catfsh sampled at age 3.1, biological rep2

description: Library name: Seasonal_Sample_6


title: testes of blue catfsh sampled at age 3.1, biological rep4

description: Library name: Seasonal_Sample_8


title: Whole blood, KY-0043-02, PETCT2, Other cold lung lesion, Hot LN_Yes, Hot Lung_No

value: Whole blood


title: ExT6_day14_Tumor_PD1nega_4

description: Library name: Sample 14


title: Whole blood, KY-0043-02, PETCT1, Normal lung, Hot LN_Yes, Hot Lung_No


title: testes of blue catfsh sampled at age 2.9, biological rep4

description: Library name: Seasonal_Sample_4

value: Age 2.9


title: ExT6_day14_Tumor_PD1nega_3

description: Library name: Sample 13


title: Whole blood, KY-0040-01, PETCT2, Normal lung, Hot LN_Yes, Hot Lung_No


title: rep3_dox

city: Cold Spring Harbor

first: Adrian

middle: R

last: Krainer

state: NY

address: 1 Bungtown Rd

institute: Cold Spring Harbor Laboratory

data_processing:
Bulk RNA-seq analyses were implemented and integrated using the “CodeSpringLab” platform developed by the CSHL Bioinformatics core, downloaded from https://github.com/RadUtama/CodeSpringLab.git. Quality control of raw sequencing files (fastq) was implemented using FastQC v0.11.8 and FastQ Screen v0.15.2 with default parameters.
Sequence alignment was performed using STAR v2.7.10a with parameters set to ‘--outFilterMismatchNmax-2’, ‘--outFilterMultimapNmax-2’, ‘--outSAMtype BAM SortedByCoordinate’, ‘--outSAMunmapped None’, ‘--outSAMstrandField None’. The human genome reference (FASTA assembly and GTF annotation) was extracted from GENCODE GRCh38.p13 v42. Gene quantification was performed using featureCounts from Subread v2.0.2 with parameters set to ‘-p --countReadPairs’, ‘-t exon’, ‘-s 0’, ‘-Q 12’, ‘-C –minOverlap 1’.
Assembly: GENCODE GRCh38.p13 v42

growth_protocol: Cells were used within 10 passages for all experiments, and were confirmed to be Mycoplasma-free routinely during the course of the study, using a Lonza MycoAlert kit. Cells were cultured at 37C and 5% CO2. SNU449 (human HCC) cells were obtained from ATCC and cultured in RPMI 1640 (Corning) supplemented with 10% FBS and 1% penicillin/streptomycin (P/S).

extract_protocol:
RNA isolation was performed using a Direct-zol RNA Miniprep kit (Zymo Research).
RNA-seq libraries were prepared with Kapa mRNA Hyper Prep kit (Roche).

treatment_protocol: For transfection with either siRNA or PKM-ASO in a 6-well plate, 7.5 µL Lipofectamine 3000 (Thermo Fisher Scientific) was mixed with 250 µL opti-MEM (Thermo Fisher Scientific), then mixed with 250 µL opti-MEM containing 10x of the final ASO or siRNA concentration, and incubated for 20 min. 500 µL of the mix was then added dropwise to 2 mL of culture medium without 1% P/S. For Doxycycline treatment, 0.25 ug/ml Dox was added to the media.

value: heptaocellular carcinoma cells

value: dox


title: testes of blue catfsh sampled at age 2.9, biological rep1

description: Library name: Seasonal_Sample_1


title: ExT6_day14_Tumor_PD1nega_2

description: Library name: Sample 12


title: Whole blood, KY-0040-01, PETCT1, Other hot lung lesion, Hot LN_Yes, Hot Lung_Yes


title: rep3_si156


title: ExT6_day14_Tumor_Ly108_5

description: Library name: Sample 10

value: lineage-negative (CD11b, B220, NK1.1 and Ter119), TCRb+,  CD4-, CD8+, PD-1+, Ly108+ and Tim3- T cells


title: rep3_tmo2

value: PKM-ASO


title: rep3_lipo

value: lipofectamine


title: testes of blue catfsh sampled at age 3.9, biological rep1

description: Library name: Seasonal_Sample_9


title: Whole blood, KY-0037-02, PETCT1, Other cold lung lesion, Hot LN_No, Hot Lung_No


title: rep2_dox


title: Whole blood, KY-0034-15, PETCT1, Other hot lung lesion, Hot LN_No, Hot Lung_Yes


title: ExT6_day14_Tumor_Ly108_4

description: Library name: Sample 9


title: rep2_si156


title: Whole blood, KY-0034-15, PETCT2, Other cold lung lesion, Hot LN_No, Hot Lung_No