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Series GSE60530 Query DataSets for GSE60530
Status Public on Apr 09, 2015
Title Profile of gene expression in U87-MG xenografts expressing control vector (V0), the ubiquitin ligase KPC1 or the p50 subunit of the NF-kB transcription factor, using RNASeq analysis of transcripts mapped independently to the human and murine genomes
Organism Homo sapiens
Experiment type Expression profiling by high throughput sequencing
Summary Purpose: We identified KPC1 as the ubiquitin ligase that binds to the p105 precursor of NF-kB, ubiquitinates it and mediates its proteasomal processing to generate the p50 active subunit of the transcription factor. Using U87-MG human glioblastoma xenografts, we observed that overexpression of KPC1 results in strong inhibition of tumor growth mediated via excessive generation of p50.The goal of this RNASeq study was to analyze the profile of gene expression in xenografts overexpressing control (V0), KPC1 or p50 vectors, and to further understand how the altered gene expression patterns can explain the tumor suppressive effect we observed.
Results:Transcript analysis of U87-MG xenografts overexpressing control (V0), KPC1 or p50 vector mapped to the human genome revealed:
• A strong similarity between overexpression of p50 and KPC1 (correlation of 0.51, p-value<10-300 )
• A specific signature of NF-kB targets [21 of the consistently changed genes are known to be regulated by NF-kB (p-value<3.4×10-9 )]
• A significant (p-value<1.4×10-18) increase in the expression of 40 tumor suppressor genes, with no significant change in other classes.
• A significant down regulation of a cluster of genes including LIN28B, IL-6, HMAGA2 and VEGFA. This finding links well to an established regulatory axis involving LIN28B, Let-7 microRNA, and IL-6 in inflammation and cell transformation that is regulated by NF-kB.

Overall design Exponentially growing U87-MG cells were stably transfected with an empty vector (V0) or vectors coding for Myc-KPC1 or Flag-p50. Cells were dissociated with trypsin, washed with PBS, and brought to a concentration of 50×10^6 cells/ml. Cell suspension (5×10^6/0.1 ml) was inoculated subcutaneously at the right flank of 7-weeks old male Balb/C nude mice (n=7). Following 21 days, mRNA from U87-MG xenografts was isolated using an RNA purification kit, and analyzed using the Illumina HiSeq 2500 sequencer. The RNASeq analysis experiment was repeated twice independently.
Run1 included a total of 7 samples. Samples 1-3 were isolated from V0 – control tumors (3 different tumors), samples 4-5 were isolated from KPC1-expressing tumors (2 different pools of 3 tumors each due to small tumor size), and samples 6-7 were isolated from p50-expressing tumors for (2 different pools of 2-3 tumors each, due to very small tumor size).
Run2 included a total of 5 samples. Samples 8-10 were isolated from V0 (control) tumors (3 different tumors), samples 11-12 were isolated from KPC1 tumors (2 different pools of 3 tumors each due to small tumor size). Several repeated attempts to extract RNA from the p50-expressing tumors did not yield any results, as the tumors were miniscule.
Contributor(s) Ciechanover A, Kravtsova-Ivantsiv Y, Shomer I, Cohen-Kaplan V, Snijder B
Citation(s) 25860612
Submission date Aug 19, 2014
Last update date May 15, 2019
Contact name Aaron Ciechanover
Phone +972-48295427
Organization name Technion-Israel Institute of Technology
Department Faculty of Medicine, Cancer and Vascular Biology Research Center
Street address Efron Street, Bat Galim, POB 9649
City Haifa
State/province Haifa
ZIP/Postal code 31096
Country Israel
Platforms (1)
GPL16791 Illumina HiSeq 2500 (Homo sapiens)
Samples (12)
GSM1481719 V0 (control) rep1- run1
GSM1481720 V0 (control) rep2- run1
GSM1481721 V0 (control) rep3- run1
BioProject PRJNA258473
SRA SRP045611

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Supplementary file Size Download File type/resource
GSE60530_final_results_U87_run1+run2.xlsx 15.6 Mb (ftp)(http) XLSX
GSE60530_genes.fpkm_tracking.gz 1.4 Mb (ftp)(http) FPKM_TRACKING
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Raw data are available in SRA
Processed data are available on Series record

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