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Series GSE188391 Query DataSets for GSE188391
Status Public on Aug 29, 2023
Title A genetic-metabolic axis for metastatic liver organotropism
Organisms Homo sapiens; Mus musculus; blank sample
Experiment type Expression profiling by high throughput sequencing
Other
Summary To generate CRISPR/Cas9 knockout screen, we used mouse Kinome CRISPR Knockout Library (Brie) (Addgene # 75316) purchased from addgene. The mouse kinase sgRNA pool includes 2,852 sgRNAs targeting 713 kinase genes and 100 control non-targeting sgRNAs. Cas9 expressing HCmel12 melanoma cells were edited for 14 days, and tumor cells were injected for metastasis screening studies. RNA-seq data: HCmel12 melanoma cells and CRISPR knockout HCmel12 melanoma cells - Pip4k2c KO were treated for 4 hours with insulin. Then cells were lysed in RLT buffer and used to perform mRNA profiling.
We performed single-cell RNA-sequencing (scRNA-seq) of eight mice with concurrent liver and lung metastasis. A total of 95,606 cells expressing 19,624 genes were included for downstream analyses.
Bulk RNA-sequencing analysis of 15 human melanoma liver metastasis samples from FFPE sample. Tumor areas were macro-dissected and RNA was extracted using Qiagen kit All Prep DNA/RNA FFPE Kit (Catalog Number 80234), concentration was quantified using Qu-bit and stored at-80 °C until use.
 
Overall design In vitro and in vivo CRISPR knockout screen of HCmel12 melanoma cells, and identification of drivers of site-specific metastasis. mRNA profiles following of Parental and Pip4k2c KO HCmel12 melanoma cells with and without insulin treatment for 4hours.
C57/BL6 mice were injected 5x10^5 HCmel12 melanoma cells of indicated genotypes via lateral tail vein and after 21 days mice were euthanized and organs harvested. Around 10,000 cells per channel were loaded on a Chromium controller (10x genomics, Pasadena, CA). Sequencing libraries were prepared using standard parameters, and libraries were sequenced on a NovaSeq with S4 flow cell (Illumina, San Diego, CA) and 2x100 bp paired end reads. For downstream analyses, we include single-cell RNA sequencing of 95,606 cells from 16 samples (MR1-MR16), with matched liver and lung metastasis from HCmel12 mouse melanoma model of indicated genotypes. MR1-MR4 are from HCmel12 melanoma cells with Pip4k2c knockout; MR5-MR10 are from HCmel12 melanoma cells with Pten knockout; MR11-MR16 are from HCmel12 melanoma cells with Pip4k2c and Pten double knockouts.
Bulk RNA-sequencing of 15 independent human melanoma liver metastasis samples were taken from FFPE samples. 5 patients were primary type cutaneous melanoma, 10 were primary type uveal melanoma. Raw counts for each transcript were quantified with kallisto using human reference GRCh38. Raw count data was normalized to transcript per million (TPM).

>>>Submitter states raw data can not be provided for Human samples due to patient privacy concerns<<<
 
Contributor(s) Rogava M, Hug C, Izar B
Citation missing Has this study been published? Please login to update or notify GEO.
Submission date Nov 08, 2021
Last update date Aug 29, 2023
Contact name Meri Rogava
E-mail(s) mr4020@cumc.columbia.edu
Organization name Columbia University
Department Hematology Oncology
Lab Izar Lab
Street address 650 W 168th St, New York, NY 10032
City New York
State/province New York
ZIP/Postal code 10032
Country USA
 
Platforms (4)
GPL17021 Illumina HiSeq 2500 (Mus musculus)
GPL24247 Illumina NovaSeq 6000 (Mus musculus)
GPL24676 Illumina NovaSeq 6000 (Homo sapiens)
Samples (139)
GSM5678509 cells_R1
GSM5678510 cells_R2
GSM5678511 cells_R3
Relations
BioProject PRJNA778715
SRA SRP345023

Download family Format
SOFT formatted family file(s) SOFTHelp
MINiML formatted family file(s) MINiMLHelp
Series Matrix File(s) TXTHelp

Supplementary file Size Download File type/resource
GSE188391_RNAseq_normalized_counts.tsv.gz 1.1 Mb (ftp)(http) TSV
GSE188391_RNAseq_normalized_counts_human.tsv.gz 1.2 Mb (ftp)(http) TSV
GSE188391_RNAseq_raw_counts.csv.gz 455.7 Kb (ftp)(http) CSV
GSE188391_RNAseq_raw_counts_human.csv.gz 469.3 Kb (ftp)(http) CSV
GSE188391_guideRNA_counts.tsv.gz 290.5 Kb (ftp)(http) TSV
GSE188391_guideRNA_lognorm.tsv.gz 814.1 Kb (ftp)(http) TSV
GSE188391_sc_barcodes.tsv.gz 435.7 Kb (ftp)(http) TSV
GSE188391_sc_counts.csv.gz 449.5 Mb (ftp)(http) CSV
GSE188391_sc_features.tsv.gz 150.6 Kb (ftp)(http) TSV
GSE188391_sc_matrix.mtx.gz 397.4 Mb (ftp)(http) MTX
GSE188391_sc_raw_feature_bc_matrix.h5 842.1 Mb (ftp)(http) H5
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Raw data are available in SRA
Processed data are available on Series record

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